Growth media

Br J Cancer, 1982 Feb, 45(2), 256 - 64
Oxygen tensions in multicell spheroids of two cell lines; Mueller-Klieser WF et al.; O2 tensions (Po2) were measured with microelectrodes in multicellular spheroids from EMT6/Ro and V-79-171-B cells . The measurements were performed in spheroids kept in flowing growth medium that was equilibrated with 5% CO2 and air at a temperature of 37 degrees C and contained 5.5 mM glucose . The recorded Po2 profiles are characterized by a diffusion-depleted zone surrounding the spheroids and by a steep drop in Po2 within the spheroids over mean distance of 220 and 188 micrometer from the surface of EMT6/Ro and V-79-171B spheroids over mean distance of 220 and 188 micrometer from the surface of EMT6/Ro and V-79-171B spheroids respectively . Smaller spheroid exhibit parabolic Po2 profiles, larger ones show a central plateau . The region of the steep decrease in Po2 corresponds to the thickness of the viable rim: the plateau region is created by the absence of O2 consumption in the central necrotic area . Po2 in the centre of EMT6/Ro spheroids decreased from 66 mmHg at a diameter of 400 micrometer to 13 mmHg at a diameter of 1000 micrometer . Under the present conditions during growth and in the experiments, values below 5 mmHg were recorded only in spheroids 1200 micrometer . Comparably low Po2 was recorded in V-79 spheroids with diameters of 650 micrometer + . In spheroids of this cell type with a diameter of 400 micrometer, Po2 was 42 mmHg . The findings provide evidence that necrosis may arise at average Po2 of 57 and 42 mmHg in EMT6/Ro and V-79-171B spheroids, respectively, grown under the conditions described.

Cancer Treat Rep, 1982 Feb, 66(2), 327 - 38
Isolation and properties of Chinese hamster lung cells resistant to ellipticine derivatives; Salles B et al.; Chinese hamster lung cells resistant to 9-OH-ellipticine (9-OH-E) were selected in vitro by adding stepwise increasing drug concentrations to the cell growth medium . This selection procedure resulted in the isolation of two sublines, about tenfold and 12-fold resistant to 9-OH-E . This level of resistance did not increase even after about 30 months of drug exposure . Cytogenetic studies revealed that the resistant cells carry several discrete karyotype modifications, the most striking being the absence of one No . 5 chromosome and the presence of a marker chromosome characterized by peculiar G-banding . Development of 9-OH-E resistance was also associated with some changes in cell properties such as modifications of morphology and growth parameters, lower oncogenic potential, and cross-resistance to a variety of antitumor agents . Although these results suggest that the resistance is associated with a modification of cell membrane properties, drug uptake studies did not show any significant difference between the parental sensitive cells and the resistant sublines.

Biokhimiia, 1982 Feb, 47(2), 284 - 9
{Changes in proteinase A and B activity during glucose repression in the yeast Saccharomyces cerevisiae}; Novikova LA et al.; The divisible and indivisible yeast S . cerevisiae were subjected to glucose repression by increasing the glucose content in the growth medium up to 10% . Under these conditions the total activities of both proteinases did not change significantly, while those of free (i.e . not bound to natural inhibitors) proteinases were increased manyfold . This effect is probably due to liberation of the proteinases from the vacuoles and digestion of cytosolic proteinase inhibitors.

Biochem Genet, 1982 Feb, 20(1-2), 17 - 28
Biochemical evidence for diverse etiologies in biotin-responsive multiple carboxylase deficiency; Packman S et al.; Biotin-responsive multiple carboxylase deficiency can be categorized by clinical criteria into a neonatal-onset disorder and distinct syndrome of infantile onset . Pedigrees in each instance are consistent with autosomal recessive inheritance . For a neonatal-onset proband, the sensitivity to relative biotin deprivation and the rapid clinical response to biotin supplementation are reflected by in vitro studies . Specific activities of biotin-dependent pyruvate carboxylase, propionyl CoA carboxylase, and 1-methylcrotonyl CoA carboxylase are 0.8 to 16% of mean control values after growth of fibroblasts in intermediate and very low biotin concentrations . Following relative biotin depletion, pyruvate carboxylase activity returns to normal after only 14 hr of growth in biotin-supplemented medium . In contrast, carboxylase activities in fibroblasts of an infantile-onset proband remain normal at very low biotin concentrations, even when avidin is added to the growth medium . The clinical heterogeneity, taken together with the distinct responses of cultured skin fibroblasts to biotin deprivation in vitro, probably reflect fundamentally different etiologies for the two categories of biotin-responsive multiple carboxylase deficiency.

J Bacteriol, 1982 Feb, 149(2), 469 - 78
Molybdenum cofactor requirement for biotin sulfoxide reduction in Escherichia coli; del Campillo-Campbell A et al.; The bisC gene of Escherichia coli is tentatively identified as the structural gene for biotin sulfoxide reductase by the isolation of bisC(Ts) mutants that make thermolabile enzyme . The products of four other E . coli genes (chlA, chlB, chlE and chlG) are also needed for enzymatic activity . Mutations previously assigned to the bisA, bisB, and bisD genes belong to genes chlA, chlE, and chlG, respectively . The biotin sulfoxide reductase deficiency of a chlG, mutant is partially reversed by the addition of 10 mM molybdate to the growth medium . Mutational inactivation of the chlD gene reduces the specific activity of biotin sulfoxide reductase about twofold . This effect is reversed by the addition of 1 mM molybdate to the growth medium . The specific activity of biotin sulfoxide reductase is decreased about 30-fold by the presence of tungstate in the growth medium, an effect that has been observed previously with nitrate reductase and other molybdoenzymes . The specific activity of biotin sulfoxide reductase is not elevated in a lysate prepared by derepressing a lambda cI857 chlG prophage . Whereas biotin sulfoxide reductase prepared by sonic extraction of growing cells is almost completely dependent on the presence of a small heat-stable protein resembling thioredoxin, much of the enzyme obtained from lysates of thermoinduced lambda cI857 lysogens does not require this factor.

Eur J Biochem, 1982 Feb, 122(1), 169 - 74
Phospholipid interconversions in Mycoplasma capricolum; Gross Z et al.; Mycoplasma capricolum cells increase their phospholipid content by incorporating exogenous phospholipids from the growth medium . Growing the cells in media with increasing serum concentrations resulted in a massive incorporation of phosphatidylcholine and sphingomyelin (up to about 50% of total phospholipids) into the cell membrane . The incorporation of the exogenous phospholipids had essentially no effect on the rate of cell growth and did not decrease the overall phospholipid biosynthesis of the cells . Thus, the ratio of phospholipid to protein in membranes from cells grown with 5% horse serum was 0.5 (mumol/mg) compared to 0.3 (mumol/mg) in cells grown without serum, and the relative content of charged polar lipids was apparently decreased . The consequence of the incorporation of exogenous phosphatidylcholine was an alteration in the relative amount of the major end-products of the de novo phospholipid biosynthesis; a marked increase in the ratio of diphosphatidylglycerol to phosphatidylglycerol was observed . The possibility that the increase in the ratio of diphosphatidylglycerol to phosphatidylglycerol is part of a control mechanism to maintain a mixture of bilayer and non-bilayer lipids is discussed.

Biochim Biophys Acta, 1982 Jan 22, 684(2), 241 - 8
Cell surface of the fish pathogenic bacterium Aeromonas salmonicida . I . Relationship between autoagglutination and the presence of a major cell envelope protein; Evenberg D et al.; A comparison was made of membrane protein patterns of various Aeromonas salmonicida strains, initially isolated from different habitats with respect to fish species affected, pathological entity, and geographic location of the outbreak of the disease . A major protein with a molecular weight of 54 000 was found in all autoagglutinating strains, whereas this protein is present in low amounts, or not at all, in non-autoagglutinating strains . Evidence for a causal relationship between the presence of this protein and the phenomenon of autoagglutination came from the observation that a change of the growth medium led simultaneously to an almost complete loss of the additional cell envelope protein and the property of autoagglutination . As it has already been reported that autoagglutination is correlated with the presence of an additional cell surface layer, we hypothesize that the additional cell envelope protein is the (major) subunit of this layer . The application of the gel immuno radio assay, an immunological technique suited to detect antigens in a gel, revealed that the additional cell envelope proteins of all tested strains are immunologically related . The possibility to the use of this protein as a component of a vaccine against A . salmonicida infections is discussed.

Biochim Biophys Acta, 1982 Jan 12, 714(1), 93 - 100
Regulation of putrescine metabolism in Ehrlich ascites carcinoma cells exposed to hypotonic medium; Kapyaho K et al.; When exposed to hypotonic growth medium, Ehrlich ascites carcinoma cells showed a rapid stimulation of ornithine decarboxylase (EC 4.1.1.17) activity in 4 h, followed by a rise in their putrescine content . This effect was totally abolished by addition of a slightly hypertonic concentration of sodium chloride or sucrose to the medium . The general protein synthesis was unaffected by the hypotonic treatment . The uptake of putrescine and, to a lesser extent, spermidine was enhanced, the conversion of the radioactive putrescine into spermidine appeared partially inhibited during later stages of the hypotonic treatment . As a result, the half-life of putrescine increased from 2.8 h under isoosmotic conditions to 7.3 h in hypoosmotic medium . Both exogenous ({14C}-putrescine-derived) and endogenous ({14C}ornithine-derived) putrescine degraded at similar rates in control and hypotonic cells, yet the putrescine taken from the medium degraded preferably to nonpolyamine products, while the putrescine synthetized in the cell was converted evenly to spermidine and to other metabolites . Adenosyl-methionine decarboxylase activity (EC 4.1.1.50), which provides the second precursor for spermidine and spermine synthesis, was distinctly inhibited in the hypotonic medium . Inhibition was likewise observed in spermidine synthase activity, while spermine synthase was marginally stimulated . It appears that the hypotonic treatment serves a special condition under which not only the formation of putrescine is enhanced dramatically but the cells also attempt to converse the diamine by preventing its further metabolism to higher polyamines.

Pharmacology, 1982, 25(3), 170 - 6
Response of mouse liver tumor cells to the differentiation-inducing agent dimethylsulfoxide; Higgins PJ; The effects of the differentiation-inducing agent dimethylsulfoxide (DMSO) on the growth and protein composition of murine hepatic tumor cell (BW77-1) cultures were investigated using various concentrations of the polar solvent in the growth medium . A 4-day exposure to DMSO in concentrations of 0.5, 1 and 3% reduced final hepatocyte population density to 63, 43 and 15% of control values, respectively, while the amount of transferrin in the cellular and extracellular compartments increased in a dose-dependent fashion . The biochemical basis for elevated transferrin extractable per 10(6) BW77-1 cells in DMSO-stimulated cultures was due to increases in both the transferrin contribution to total cellular protein and the mean protein content per cell . This stimulation in cellular protein levels was reflected in the incorporation of {3H}-amino acids into cell-associated 10% trichloroacetic acid-insoluble material . Cultures of transformed liver epithelial cells thus appear to respond to in vitro DMSO exposure by increasing their accumulation of a late-stage hepatocyte gene product.

Z Allg Mikrobiol, 1982, 22(6), 379 - 88
{Cytochrome pattern of methylotropic acetic acid bacterium MB 58 as dependent on growth conditions}; Steudel A et al.; In contrast to methylotrophic bacteria investigated up to now the facultative methylotrophic Bacterium MB 58 (Acetobactersp . MB 58) does not possess a cytochrome aa3-complex, but we could find out cytochrome, cytochrome cco, cytochrome a1, and moreover cytochrome d in dependence on the growth conditions . Cytochrome d was found only in stationary phase of heterotrophic growth . Under methylotrophic growth conditions cytochrome d could be demonstrated only by lowering of the aeration rate during the fermentation, by variation the pH-value of the growth medium from 4.0 to 6.5 and with low growth rates (low dilution rates) during continuous fermentation . The addition of cyanide to the oxidized suspension of bacteria during the registration of the cytochrome-redox-difference spectrum allowed the selective representation of cytochrome d under all conditions even if no identification was possible in the spectrum normally . The oxidation of cytochrome d of Acetobacter sp . MB 58 in the presence of cyanide is an indication of its cyanide insensitivity . The low level of b-type cytochromes could be represented by a special technique for registration of spectra . In this connection a unknown absorption peak at 570 nm was registered . The cyanide insensitivity of cytochrome d from Acetobacter sp . MB 58 and the occurrence of several terminal oxidases is appreciated as a hint for a branched respiratory chain.

Int Arch Allergy Appl Immunol, 1982, 69(3), 224 - 30
Isolation and characterization of the allergen Dpt 12 from Dermatophagoides pteronyssinus by chromatofocusing; Stewart GA; The allergen Dpt 12, from the house dust mite Dermatophagoides pteronyssinus, was isolated from spent growth medium by chromatofocusing . The allergen was isolated in the pI range 5.86-6.95, indicating heterogeneity with regard to pI . However, on sodium dodecylsulphate-polyacrylamide gel electrophoresis the allergen appeared homogeneous with an apparent molecular weight of 27,000 daltons . Radioallergosorbent scores utilizing paper discs coupled with Dpt 12 correlated significantly (p less than 0.0005) with those obtained using discs coupled with whole mite extract, although the corresponding titres were higher with Dpt 12 (p less than 0.0005).

Mol Cell Biol, 1982 Jan, 2(1), 76 - 81
Identification and partial characterization of an extracellular acid phosphatase activity of Leishmania donovani promastigotes; Gottlieb M et al.; An extracellular acid phosphatase was detected in the growth media of Leishmania donovani promastigotes . The enzyme was released at all stages of the growth cycle and in amounts which accounted for 90% of the total amount of this enzyme in the culture . The exoenzyme exhibited a pH optimum of 4.5 to 5.0 and was active with a variety of organic phosphates . The enzymatic activity was excluded from Sephacryl S-300 and was retained by ultrafilters with nominal molecular weight cutoffs of up to 300,000 . The results of comparative studies indicated that the extracellular enzyme was distinct from a surface membrane-bound acid phosphatase of L . donovani promastigotes which has been previously described.

J Cell Biochem, 1982, 18(1), 121 - 33
Differential reactivation of zinc-mediated metallothionein induction in ultraviolet-irradiated normal and repair-deficient human cells; Hildebrand CE et al.; The ubiquitous, low-molecular-weight, thiol-rich, metal-binding protein, metallothionein (MT), can be induced in cultured normal human fibroblasts (NF) and xeroderma pigmentosum (XP) cells by exposure to ZnCl2 . Both NF and XP cells tolerate up to 200 microM ZnCl2 in the growth medium, upon addition of ZnCl2 (200 microM) to monolayer cultures, both NF and XP cells showed similar kinetics for the induction of MT synthesis: Within 7 hours the MT synthesis rate rose from a low, marginally detectable rate to a maximal rate at least 50-fold greater than the basal rate . The induction of MT synthesis in both cell types was inhibited by actinomycin D (5 microgram/ml), indicating that the induction process is controlled at the level of transcription . Exposure of NF and XP cells to far ultraviolet light (UV) followed by induction with ZnCl2 resulted in a UV dose-dependent decrease in the he maximal rate of MT synthesis measured 8.5 hours postirradiation . The UV sensitivity of the MT induction was greater in XP cells than in NF cells . However, considerations of the differential repair capacities of NF and XP cells superimposed upon the kinetics of MT induction were invoked to explain the apparent differential UV sensitivity of MT induction . Liquid holding recovery experiments showed that NF cells possess the capacity to reactivate this inducible gene function rapidly while XP cells are deficient in the reactivation capacity . These results are discussed in the context of both UV transcriptional mapping of this inducible gene function and development of techniques for measuring repair of transcription-blocking lesions.

J Bacteriol, 1982 Jan, 149(1), 338 - 45
Possible association of segregated lipid domains of Mycoplasma gallisepticum membranes with cell resistance to osmotic lysis; Rottem S et al.; Freeze-fracturing of cholesterol-rich Mycoplasma gallisepticum membranes from cells grown in a medium containing horse serum revealed particle-free patches . The patches appeared in cells quenched from either 4 or 37 degrees C . Particle-free patches also occurred in membranes of cells grown in a serum-free medium supplemented with egg-phosphatidylcholine but not in membranes of cells grown with dioleoylphosphatidylcholine . The appearance of particle-free patches was attributed to the presence of disaturated phosphatidylcholine (PC) molecules in M . gallisepticum membranes, which were synthesized by the insertion of a saturated fatty acid at position 2 of lysophosphatidylcholine derived from exogenous PC present in the growth medium . Consequences of the synthesis of the disaturated PC also included a decrease in osmotic fragility and the ability of the cells to be permeated by K+ . Electron paramagnetic resonance and fluorescence polarization measurements revealed that the fluidity of the lipid domain in the protein-rich M . gallisepticum membranes was almost identical to that of an aqueous dispersion of M . gallisepticum membrane lipids . Furthermore, the electron paramagnetic resonance spectra of the membranes were single-component spectra showing no indication of immobilized regions . The possibility that the osmotic resistance of M . gallisepticum cells is associated with the particle-free patches rather than with a restricted membrane fluidity caused by membrane proteins is discussed.

J Bacteriol, 1982 Jan, 149(1), 131 - 5
Iron transport in Mycobacterium smegmatis: Uptake of iron from ferric citrate; Messenger AJ et al.; In mycobacterial growth medium 40 to 400 microM citrate was required to solubilize 2 microM 55Fe . This solubilized 55Fe was taken up into both iron-deficient and iron sufficient washed cell suspensions of Mycobacterium smegmatis and Mycobacterium bovis BCG . Although the 55Fe was taken up into the cell, the citrate was not . The uptake system with M . smegmatis was not inhibited by electron transport inhibitors, uncouplers of oxidative phosphorylation, or thiol reagents and was saturable with iron at approximately 35 microM . The system was independent of the iron transport systems already known to exist in M . smegmatis: i.e., the two exochelin routes of assimilation as well as the mycobactin-salicylate system . It was not induced by the presence of 400 microM citrate in the growth medium, nor did the presence of citrate in the medium affect the production of either exochelin or mycobactin.

Mol Gen Genet, 1982, 186(2), 157 - 63
Characterization of a yeast regulatory mutant constitutive for synthesis of inositol-1-phosphate synthase; Greenberg ML et al.; The enzyme inositol-1-phosphate synthase is repressed at least 50-fold in wild type yeast grown in inositol-supplemented media . Mutants which synthesize this enzyme constitutively have been isolated using a selection procedure based on excretion of inositol into the growth medium by putative mutants . Biochemical analysis of one of the mutants (opi1-1) confirmed that the nature of the mutations is regulatory, and not in the structural gene for the enzyme . Immunoprecipitation of crude extracts with antibody directed against purified inositol-1-phosphate synthase showed that a protein which reacts with the antibody is present in the mutant grown under both repressing and derepressing conditions, in contrast to the wild type which synthesizes the enzyme only when derepressed . Assay of inositol-1-phosphate synthase activity in crude extracts of the mutant verified synthase activity in cells grown under both repressing and derepressing conditions . Synthase purified from this mutant was characterized with respect to molecular weight, thermolability and affinity for substrates glucose-6-phosphate and NAD . These analyses indicated that purified mutant synthase was similar to the wild type enzyme.

Comp Biochem Physiol B, 1982, 71(3), 397 - 402
Fatty acids of Trypanosoma cruzi; Timm SL et al.; 1 . The fatty acid pattern of total lipids from T . cruzi is different from one of its growth medium . 2 . The distribution of major fatty acids in phospholipid fraction was linoleic (50.4%), oleic (25.6%), stearic (10.1%) and palmitic (6.3%) and in neutral lipid fraction oleic and linolelc (about 29% each), palmitic (18.3%) and stearic (9.8%) . 3 . In each of the individual phospholipids a particular fatty acid pattern was observed . 4 . For phosphatidylcholine: linoleic (55.7%), oleic (16.6%), stearic and palmitic (about 8% each) and polyunsaturated acids (4%) . 5 . For phosphatidylethanolamine: linoleic (41.5%), oleic (27.6%), stearic (9.9%) and an unidentified fatty acid (9.2%) . 6 . For phosphatidylinositol: stearic and linoleic (about 30% each) and oleic (22.1%).

Biosci Rep, 1982 Jan, 2(1), 15 - 30
Calcium ions and the control of proliferation in normal and cancer cells; Durham AC et al.; Several lines of evidence suggest tha Ca2+ ions control cell proliferation: Ca2+ entry into cytoplasm acts as a general mitogen; serum and serum-replacements induce Ca2+ influx; the Ca2+ concentrations in growth media required to support the proliferation of normal cells are much higher than those required for cancer cells; serum and growth factors reduce the Ca2+ requirements of normal cells; tumour promoters alter Ca2+ fluxes via a mechanism used principally by growth factors . Minor supporting evidence includes the effects of various drugs and viruses, and the behaviour of tumour cell mitochondria and intercellular junctions . It is still not possible to decide exactly where and when inside cells the critical effect of Ca2+ on proliferation occurs, but we discuss at length the practical problems of understanding Ca2+ movements in tissue-culture cells . Carried to its logical conclusion, present evidence suggests that an overridden or bypassed Ca2+ control process may be the key, common determinant of unrestrained proliferation in cancer cells.

J Cell Biochem, 1982, 20(1), 41 - 50
Involvement of multiple subcellular compartments in intracellular processing of epidermal growth factor; Miskimins WK et al.; The intracellular translocation and processing of epidermal growth factor (EGF) in 3T3 cells has been studied utilizing Percoll density gradients . EGF is internalized and rapidly becomes associated with two types of intracellular compartments . The extent to which EGF is delivered to these two compartments is apparently regulated depending upon the cell's physiological condition . In growth medium, an increased proportion of EGF is taken up into a Golgi-like element . Uptake through this pathway correlates with a decrease in degradation of the ligand . In the absence of serum and amino acids, an increased proportion of EGF is taken up into a component which has a density of 1.05 . Uptake through this pathway correlates with increased degradation of the ligand . The ligand taken up through both pathways is transferred to dense vesicles which comigrate with lysosomes . In the presence of growth medium, however, dense vesicles containing EGF can be shown to be lysosomal enzyme-deficient upon further fractionation . In addition, in the presence of serum, a portion of the internalized EGF is apparently released from the cells, intact, and then re-bound . The processes described may be important in the production of a mitogenic response and the ability of cells to self-regulate their responsiveness to the growth factor.

Arch Virol, 1982, 74(1), 11 - 20
The effect of cytochalasin D and monensin on enveloped vaccinia virus release; Payne LG et al.; Both monensin and cytochalasin D reduced the production of infectious cell associated virus and infectious extracellular virus with the latter clearly being the more sensitive . The difference in yields were even more clearly seen if the yield of virus particles was monitored instead of yield of infectious virus . Addition of 1 microgram/ml cytochalasin D or 1 microM monensin to the growth medium of vaccinia virus-infected cells inhibited the appearance of extracellular enveloped vaccinia virus (EEV) in the growth medium without affecting the production of intracellular naked vaccinia virus (INV) particles . Although EEV was not released into the medium of cytochalasin D treated cells, EEV was, nevertheless, detectable in CsCl gradients of cell associated virus . Monensin treatment did not affect the synthesis of vaccinia glycoproteins but did significantly reduce the transport of these glycoproteins to the cell surface and also reduced the secretion of proteins . Monensin had the additional effect of causing the appearance of numerous large vacuoles in the cytoplasm . Vaccinia is normally wrapped by cytoplasmic membranes in preparation for release . The monensin induced conversion of cytoplasmic membranes to large vacuoles is presumably the basis for the block in virus wrapping and subsequent release . Cytochalasin D did not alter any of the steps in protein synthesis, transport or secretion . Electronmicroscopic studies confirmed the existence of EEV on the surface of infected cells treated with cytochalasin D . This drug which specifically affects cellular microfilament organisation thus imposes a block on the final release of EEV from the cell surface.

J Cell Physiol, 1982 Jan, 110(1), 9 - 16
Effects of isolation and culture on prostaglandin synthesis by porcine aortic endothelial and smooth muscle cells; Ager A et al.; Freshly isolated neonatal porcine aortic tissue (smooth muscle with or without endothelium present) produced approximately 30 ng/mg wet tissue of 6-oxo-prostaglandin F1 alpha (the stable hydrolysis product from prostacyclin) and approximately 15 ng/mg of prostaglandin E2, as measured by radioimmunoassay after 24 h incubation in culture medium . Primary cultures of porcine endothelial and smooth muscle cells (isolated by enzymic digestion of aortic tissue) exhibited the same pattern of prostaglandin production, but absolute values were greater than for fresh tissue, particularly in the case of endothelium . Subcultures of endothelium produced smaller amounts of prostaglandins, although the pattern remained similar . In contrast, subcultures of smooth muscle cells produced a greater total amount of prostaglandins than did primary cultures, and the main product was prostaglandin E2 . Experiments with {14C} prostaglandin H2 or {14C}arachidonic acid confirmed that aortic tissue, cultured endothelium, and primary cultures or aortic smooth muscle cells synthesized prostacyclin, and demonstrated that subcultured smooth muscle cells enzymically isomerised prostaglandin H2 to prostaglandin E2 . Kinetic studies showed that prostaglandin production by cultured vascular cells was transiently increased by subculture or changing the growth medium, and that production per cell declined with increasing cell density . The change in pattern of prostaglandin production during culture was shown to be due to a rapid decline in the rate of prostacyclin production (which apparently began immediately after tissue isolation), together with a more gradual rise in prostaglandin E2 production . These results indicate that the amounts and ratios of prostaglandins produced by vascular endothelial and smooth muscle cells are greatly affected by the conditions used to isolate and culture the cells; vascular cells in vivo may similarly alter their pattern of prostaglandin production in response to local changes in their environment.

J Bacteriol, 1982 Jan, 149(1), 361 - 3
Accumulation of poly-beta-hydroxybutyrate in Spirulina platensis; Campbell J 3rd et al.; Poly-beta-hydroxybutyrate has been identified in the cyanobacterium Spirulina platensis . The addition of reduced carbon compounds to the growth medium was not required for poly-beta-hydroxybutyrate accumulation . Poly-beta-hydroxybutyrate accumulated during exponential growth to 6% of the total dry weight and then decreased during the stationary phase.

Microbios, 1982, 33(133-34), 149 - 60
Growth studies on Mycobacterium BCG: establishment of growth curves and measurement of the oxygen tension of the growth medium; Moore DF et al.; Mycobacterium BCG grew exponentially in shallow, static volumes of culture medium for approximately 10 days; the oxygen tension of the medium at all stages of growth was 100% saturation . Higher yields were obtained from Dubos than from glycerol-free medium . In static cultures, the oxygen tension of the culture and consequently the growth rate of BCG was dependent on the depth of the medium; active growth ceased at an oxygen tension of less than 40% saturation . BCG grew actively in a cell sediment, while cells growing in suspension made a negligible contribution to the yield.

Genetika, 1982, 18(8), 1245 - 54
{Escherichia coli K-12 mutants with enhanced resistance to ionizing radiation . II . Study of DNA damage and repair in gamma-irradiated cells}; Bresler SE et al.; Formation and repair of single-stranded breaks (ssb) and double-stranded breaks (dsb) in DNA of gamma-irradiated cells of three Escherichia coli strains (the parental strain AB1157 and radiation-resistant mutants Gamr444 and Gamr445) were studied by centrifugation in alkaline or neutral sucrose gradients . The initial yield of ssb and the kinetics of their repair during post-irradiation incubation in a growth medium are similar for Gamr mutants and the wild-type strain . The yield of dsb in the chromosome of Gamr mutants is significantly lower than in chromosomal DNA of the parental strain, both immediately after gamma-irradiation and after three hours of post-irradiation incubation in a growth medium . The decreased yield of dsb in the Gamr mutants correlates with their relative radioresistance . It was found also that the level of DNA degradation is significantly lower in UV- or gamma-irradiated cells of Gamr mutants, as compared with the wild-type strain . It is suggested that enhanced radioresistance of the Gamr mutants is due to decreased formation of enzymatically induced dsb, as a consequence of moderated DNA degradation under the action of repair exonucleases and also to enhanced efficiency of dsb repair.

EMBO J, 1982, 1(7), 801 - 4
An ion-channel forming protein produced by Entamoeba histolytica; Lynch EC et al.; We have identified a remarkable ion-channel forming material in virulent strains of Entamoeba histolytica that may be responsible for many of the symptoms associated with amoebic dysentery . A polypeptide that we refer to as amoebapore is shed into the growth media and is also found within the amoeba in a high speed sedimentable fraction . Amoebapore has the distinctive property of spontaneously incorporating into lipid bilayers, liposomes, and cells, leading to progressive and irreversible changes in the ion conductance of the target membranes . Exposure of planar lipid bilayers to amoebapore -containing fractions under voltage clamp conditions results in an almost immediate and progressive incorporation of ion channels which continues in an irreversible manner leading to a fall in membrane impedance of up to five orders of magnitude . The ion-channel conductance is moderately cation-selective, voltage dependent, and displays a unit size of 1.6 +/- 0.2 nanoSiemens in 1 M KCl at -10 mV . In the bilayer, the amoebapore -induced conductance exhibits an in situ sensitivity to protease . Amoebapore is mainly concentrated in a fraction sedimenting at 150 000 g . It is insoluble in Triton X-100 but can be dissociated in an active state in 1% SDS . Under these conditions it has an apparent mol . wt . of 13 000 daltons.

EMBO J, 1982, 1(3), 333 - 7
Regulation of adenylate cyclase in E . coli; Gstrein-Reider E et al.; The intracellular concentrations of cAMP in Escherichia coli are regulated mainly by control of the activity of adenylate cyclase . Withdrawal of the carbon source from the growth medium causes a gradual reduction of cellular energy and a dramatic stimulation of cyclase activity . Manipulations of the proton gradient at the cell membrane of ATP synthase-deficient E . coli (unc-) revealed that this part of the energy compartment is not responsible for the starvation-induced stimulation of cyclase . Neither is the ATP pool involved in regulation of the activity of the cyclase . The intracellular concentrations of ATP were experimentally lowered by purine starvation of auxotrophs, by inhibition of purine synthesis using amethopterin, or by affecting ATP synthesis using arsenate . None of these conditions led to stimulation of cyclase activity . The control of cyclase is exerted not via the energy pools but via uptake systems of energy substrates independent of whether the substrate can be metabolized or not, or how the transport is energized . The stringent coupling between these transport systems and cyclase activity enables the cell to react instantaneously to changes in its environment.

Folia Biol (Praha), 1982, 28(5), 311 - 6
Inhibitory activity of cytosine arabinoside on Marek's disease virus; Benda V; The presence of ara-C in growth medium at concentrations of 10(-4) to 10(-7) M completely or partially suppressed the formation of plaques specific for MDV or HVT and decreased proportionally the growth of HPRS line 1 lymphoblastoid cells . Administration of ara-C to chickens immediately after infection with MDV (1 mg/chicken/day i.p . for 5 days) reduced the incidence of Marek's disease by 50% . Thus ara-C appears to be an inhibitor of Marek's disease.

Mol Gen Genet, 1982, 186(4), 482 - 7
Negative control of ornithine decarboxylase and arginine decarboxylase by adenosine-3':5'-cyclic monophosphate in Escherichia coli; Wright JM et al.; The polyamine biosynthetic enzymes, ornithine decarboxylase (EC 4.1.1.17) (ODC) and arginine decarboxylase (EC 4.1.1.19) (ADC), are negatively controlled by cAMP in Escherichia coli . The specific activities of ODC and ADC were determined in crude extracts prepared from E . coli strains carrying a mutation in the adenylate cyclase (EC 4.6.1.1) structural gene (cya) and wildtype strains . These strains were cultured on various carbon sources in the presence and absence of cAMP . In wild-type strains, ODC and ADC activities were diminished in cells grown on glycerol compared to these strains cultured on glucose . When cya strains were grown on glucose or glycerol, ODC and ADC activities were the same . Addition of 1 mM cAMP to glucose-based medium repressed ODC and ADC activities in both the wild-type and cya strains . Furthermore, cAMP exerts its negative control through the cAMP receptor protein, since strains carrying a mutation in the crp structural gene fail to repress ODC and ADC activities in response to increased cAMP obtained by carbon source manipulation or cAMP supplementation of the growth medium . This evidence suggests that negative control of ODC and ADC by cAMP occurs at the level of transcription.

J Cancer Res Clin Oncol, 1982, 103(1), 17 - 29
Induction of differentiation in human promyelocytic leukemia cells by tumor promoters; Nakayasu M et al.; 12-)-Tetradecanoylphorbol-13-acetate (TPA), the prototype polyfunctional diterpene ester tumor promoter of two-step carcinogenesis in mouse skin, induced differentiation of human promyelocytic leukemia cells (HL-60) in culture . Differentiation of HL-60 cells was characterized by increased phagocytosis, increased lysozyme activity (EC 3.2.1.17) in the growth medium, and changes in morphology to those characteristics of more mature cells resembling macrophages . Many of the cells treated with TPA became aggregated, attaching firmly to culture flasks . The average intracellular myeloperoxidase activity (EC 1.11.1.7) per cell decreased during induction of differentiation by TPA . It was also found that TPA enhanced, rather than inhibited, differentiation of HL-60 cells induced by DMSO . In addition to TPA, several polyfunctional diterpene esters of the tigliane, ingenane, and daphnane type have been tested for their ability to induce morphological and functional changes of HL-60 cells . The activities of the compounds to induce these changes correlated well with their activities as tumor promoters in two-step carcinogenesis in mouse skin . In particular, half the concentrations required for induction of adhesion of the cells to flasks were roughly correlated to the potency of these compounds as tumor promoters . Among the compounds tested, phorbol-12,13-didecanoate (PDD), ingenol-3-hexadecanoate, Pimelea factor P1 and Pimelea factor P2 were as active as TPA, while 4-O-methyl-TPA and 4 alpha-PDD were much less active . Phorbol and ingenol were totally inactive up to a concentrations 10,000-fold higher than that of TPA.

Vopr Virusol, 1982 Jan-Feb, (1), 34 - 8
{Hormone-dependent process of the maturation of the major structural proteins of the mouse mammary tumor virus}; Beriashvili MM et al.; The following findings were obtained by the radio-immunoprecipitation method with antisera to gp52 and p27 . When the cells were cultivated in a hormone-free medium, they contained precursors of proteins of gene gag (Pr 73gag) and gene evn (gPr 70env) . No mature structural MTV proteins were found . The addition of insulin to the growth medium had no effect on maturation of protein precursors . When dexamethazone was added to the medium, gPr 70env "maturated" to gp52 of the main envelope glycoprotein of virion coat whereas Pr 73gag was not cleaved into final products . When the cells were cultivated in the presence of insulin and dexamethazone, there was a marked stimulation of Pr 73gag (although its maturation did not occur), g Pr 70env, and, especially, gp 52 . Extracellular particles were found to contain p27 which was precipitated by homologous monospecific antiserum, that is, in the system of clone F2 cells processing of Pr 73gag occurred mainly in extracellular particles . Thus, expression of all three known genes (gag, pol, and env) occurred in the cells of cloned F2 culture.

Folia Microbiol (Praha), 1982, 27(2), 76 - 80
Relationship between the composition of phospholipids and respiratory activity of choline-deficient mutants of Neurospora crassa; Fecikova H et al.; Phosphatidylcholine is one of the most frequent phospholipid components of the inner mitochondrial membrane of Neurospora crassa . Quantitative analysis of phosphlipids of the wild strain of Neurospora crassa and of its two cho mutants showed that these strains did not significantly differ in the content of phosphatidylcholine . Mutants cultivated in a medium without choline contained, as compared with the wild strain, an increased amount of phosphatidylserine and a decreased quantity of phosphatidic acid . Respiratory activity increased and sensitivity to inhibitors of respiration changed . It is likely that the presence of choline in the growth medium exerts a regulatory effect on the cell metabolism of these mutants.

Arch Dermatol Res, 1982, 273(1-2), 9 - 14
Pemphigus antibodies fixation and keratinocyte differentiation in organ cultures . Effects of some agents influencing the intracellular content of cyclic AMP; Sbano E et al.; The effect of some agents, influencing the cyclic adenosine 3', 5'-monophosphate (cAMP) content of human cells, on the ability of the keratinocytes of binding pemphigus antibodies was studied by using tissue cultures of rabbit esophagus . As demonstrated by immunofluorescence (IF) for IgG, the bound antibodies appeared markedly decreased on esophagus explants grown under standard conditions, that is without test agents, when compared to ones fixed on fresh esophagus . But the IF reaction was remarkably more intense when methylxanthines or epinephrine were added to the growth medium of the cultures . Following the addition of these agents to the cultures some histologic modifications appeared in the explants, indicating that the keratinization process had probably been stimulated . This temporal relationship of immunofluorescence and histologic findings seems to suggest the hypothesis that keratinocyte differentiation, regulation of cAMP intracellular content, and pemphigus antibodies fixation are related processes.

Histochemistry, 1982, 76(1), 113 - 21
Flow cytometric analysis of chromosomes and cells using a modified BrdU-Hoechst method; Severin E et al.; Chromosomes and interphase cells were harvested from cultures of the Chinese hamster line B14 F28 grown in medium containing BrdU up to four cell cycles and stained with the fluorescent dye 33342 Hoechst for flow cytometry . The newly synthetized BrdU-DNA is not stainable by the Hoechst dye which is highly specific for thymidine . The temporal development of the DNA fluorescence after addition of BrdU to the growth medium has been investigated . The chromosomal fluorescence intensity is reduced one step per generation . The extent of the intensity decrease by BrdU incorporation is proportional to the amount of new DNA and it is realized by repeated measurement following an UV-exposure . This UV-illumination stops the quenching by BrdU of the Hoechst stain induced DNA fluorescence . Therefore, the entire DNA content of these chromosomes now becomes measurable . The obtained intensity gain serves as a measure of the extent of the previous BrdU caused intensity shift . In this way we could establish 3 successive mitoses . Principally, this method is suitable also for measurement of whole cells in order to obtain both the number of generations in the experimental period and the phase distribution of the cell cycle.

Arch Virol, 1982, 71(4), 311 - 22
Characterization of a hamster brain cell line persistently infected with measles virus; Vainionpaa R et al.; A persistent infection of measles virus in hamster brain cell cultures was established . Hamster brain cells were transformed with a human papovavirus strain BK and infected with a wild-type measles virus in order to get the cell line persistently infected with measles virus . About 75 per cent of these M-HB/MVB-cells were measles virus-infected . The cells released only small amount of infectious virus and produced low levels of interferon-like activity into the growth medium . During the first 50 passages no large syncytia typical of a lytic measles virus infection were seen . The products of measles virus infection in the cells and in cell culture fluids were followed at two temperatures . Another cell line persistently infected with measles virus (Lu-carrier-cells, 28) was investigated in parallel . In both cell lines measles antigens were cytoplasmic, but during the observation period large amounts of measles nucleocapsid accumulated in the nuclei of the M-HB/MVB-cells . The virus-specific protein synthesis in M-HB/MVB-cells was weak and the intracellular amount of immunoreactive measles nucleocapsid was only 50 per cent (600 ng/10(5) infected cells) of the (1200 ng/10(5) cells) found in Lu-carrier-cells . Also the release of nucleocapsid was minimal in hamster brain cells . The decreased temperature had no clear-cut effect on virus-specific protein synthesis or on the release of the virus-specific products.

J Natl Cancer Inst, 1982 Jan, 68(1), 27 - 33
In vitro reversal of depressed T-lymphocyte function in the peripheral blood of brain tumor patients; Wood GW et al.; Peripheral blood mononuclear cells from some brain tumor patients exhibited depressed T-lymphocyte responses to the polyclonal mitogen phytohemagglutinin (PHA) . These responses, initially depressed, were restored to near normal by a 24-hour preculture in growth medium . The effect of preculture was mimicked by mild trypsinization before addition of PHA . In addition, cells treated with deaggregated antihuman IgG resulted in greatly reduced responses to mitogen by mononuclear cells from all brain tumor patients . Anti-IgG had no effect if cells were precultured for 24 hours . The results suggest that, in brain tumor patients, depressed immune function associated with tumor progression was caused by suppressor cells which were activated by humoral factors that express IgG antigenic determinants.

J Cell Biochem, 1982, 20(4), 369 - 80
Influence of growth conditions on the composition of the plasma membrane from yeast and on kinetic properties of two membrane functions; Stadtlander K et al.; The influence of different growth conditions on the phospholipid composition and on two membrane functions, the Mg-ATPase and the purine transport system, was investigated . Addition of cholinechloride to the growth medium led to a certain rise in the amount of phosphatidylcholine, whereas supplementation with ethanolamine resulted in a considerably higher portion of phosphatidylethanolamine . When yeast cells were cultured at lower temperatures we found more short-chain fatty acids with a higher content of monounsaturated chains as compared to higher growth temperatures . Addition of paraquat, a herbicide which enhances lipid peroxidation by free radicals, reduced the amount of unsaturated fatty acids without influencing their chain length . The altered membrane composition had no influence on the basic mechanism of interaction between ATPase, MgATP, and free Mg2+ ions . However, several kinetic constants such as Km, Vmax, Ka, and especially Ki were influenced to some extent . Whereas the affinity of the purine transport system to its substrate was not significantly changed by the growth conditions, an effect on Vmax could be seen . Lower growth temperatures clearly led to higher maximal uptake velocities . The presence of paraquat during growth resulted in a considerable decrease of Vmax.

Mol Gen Genet, 1982, 186(1), 33 - 9
Regulation of L-amino acid oxidase and of D-amino acid oxidase in Neurospora crassa; Sikora L et al.; Neurospora crassa possesses an inducible L-amino acid oxidase that is expressed only when cells are derepressed for nitrogen in the presence of an amino acid . Enzyme synthesis requires both induction by an amino acid and simultaneous nitrogen catabolite derepression . Carbon limition in the presence of an amino acid does not permit induction of L-amino acid oxidase . The nit-2 gene is a major regulatory locus which is believed to mediate nitrogen catabolite repression in Neurospora . Mutants of nit-2 are repressed for L-amino acid oxidase activity under conditions which lead to good enzyme induction in wild type and nit-2 revertants . The loss of the enzyme in nit-2 mutants does not result from inducer exclusion, which suggests that the nit-2 gene product has a direct role in controlling the expression of this enzyme . Substantial amounts of L-amino acid oxidase were detected in the growth medium as well as in cell extracts of the wild type strain . Biochemical data indicates that the intracellular and the extracellular L-amino acid oxidases are identical . Inhibitors of protein and of RNA synthesis block accumulation of L-amino acid oxidase, suggesting that enzyme expression is controlled at the level of transcription . D-amino acid oxidase can be detected in cell extracts of Neurospora grown in the presence of a D-amino acid . The enzyme is present in cys-3 mutants and is not repressed by high concentrations of sulfate or nitrogen indicating that D-amino acid oxidase is not a member of the sulfur or nitrogen regulatory circuits of this organism.

Rev Esp Oncol, 1982, 29(4), 641 - 7
{Use of purine rescue pathways by L1210 cells exposed to methotrexate in vitro}; Buesa JM; Methotrexate (MTX) blocks the de novo synthesis of purines and pyrimidines due to a diminution of reduced folates, and produces an accumulation of intracellular 5-phosphoribosyl-1-pyrophosphate (PRPP) . Phosphoribosyltransferases, in the presence of PRPP, synthesize nucleotides starting on purine bases, that are activated to triphosphates ("rescue pathway") . The authors study the activity of the rescue pathways in L1210 leukemia cells in vitro to restore the levels of ATP and GTP that have been lowered by MTX . Hypoxanthine (Hpx) was used as a preformed source of purines, high efficiency liquid chromatography was employed to estimate the intracellular concentrations of ATP and GTP . Two hours after 0.1 mM Hpx addition to in vitro L1210 cells in the presence of 0.1 microM MTX, the amount of ATP and GTP increased 5 to 7 times . In former experiments the author has shown that the uptake of PRPP increases when Hpx is added to L1210 cells exposed to MTX . The results of this work show that L1210 leukemia cells in vitro can use the rescue pathways of purine synthesis when Hpx is present in the growth medium, so preventing the anti-purine effect of MTX.

J Biol Chem, 1981 Dec 10, 256(23), 12156 - 63
Ornithine decarboxylase from Saccharomyces cerevisiae . Purification, properties, and regulation of activity; Tyagi AK et al.; Ornithine decarboxylase has been purified 1,500-fold to homogeneity from a spe2 mutant of Saccharomyces cerevisiae which lacks S-adenosylmethionine decarboxylase and is derepressed for ornithine decarboxylase . The ornithine decarboxylase is a single polypeptide (Mr = 68,000) and requires a thiol and pyridoxal phosphate for activity . Addition of 10(-4) M spermidine and 10(-4) M spermine to the growth medium reduces the activity of the enzyme by 90% in 4 h . However, immunoprecipitation studies showed that the extracts of polyamine-treated cells contain as much enzyme protein as normal cell extracts . This loss of ornithine decarboxylase activity is probably due to a post-translational modification of enzyme protein because we found no evidence for any inhibitor of activity in the polyamine-treated cells.

Lab Anim Sci, 1981 Dec, 31(6), 723 - 5
A technique for collection and cultivation of macrophages from Cynomolgus monkeys (Macaca fascicularis); Heisey GB et al.; Several techniques were compared for collection of peritoneal mononuclear phagocytes from cynomolgus monkeys . A multiple-holed cannula proved superior to a hypodermic needle for the collection of peritoneal washings because it was not occluded by the omentum . With the cannula technique 55--70% of the harvest medium could be recovered from the peritoneal cavity with a yield of 2.5--5 x 10(5) macrophages per ml . A closed system for injecting and collecting harvest medium was better than a syringe in preventing bacterial contamination of peritoneal washings . After collection, cells were resuspended in growth medium to a concentration of 5 x 10(5) macrophages per ml, distributed into Leighton tubes, and incubated at 36 degrees C . After 12 hours, cultures were washed to remove non-adherent cells and refed with growth medium . This technique provided a sterile, reliable method for the collection and cultivation of macrophages from monkeys.

Can J Microbiol, 1981 Dec, 27(12), 1298 - 305
A defined growth medium for the production of chloroperoxidase by Caldariomyces fumago; Pickard MA; Ten strains of Caldariomyces fumago and related fungi were found to produce extracellular chloroperoxidase when grown on a glucose - malt extract medium . High enzyme levels and pigment production were observed for C . fumago ATCC 16373 and C . fumago CMI 89362 . Removal of malt extract from the medium and the replacement of glucose by fructose as the carbon source provided a defined medium which, by comparison with the complex medium, produced the following results with both fungal strains . Chloroperoxidase was produced to similar levels, with maximum production after 6 days rather than 12 days of growth; pigmentation of the medium was reduced by 90% and the pH of the medium remained constant, thus stabilizing enzyme activity . Addition or urea or proline as a nitrogen supplement to nitrate enhanced enzyme production by strain CMI 89362 . Comparison of the two strains indicated that CMI 89352 produced higher levels of chloroperoxidase than ATCC 16373.

Eur J Cell Biol, 1981 Dec, 26(1), 154 - 7
Cytochalasin B inhibits polyamine biosynthesis in HeLa cells; Sunkara PS et al.; Cytochalasin B (CB), a potent inhibitor of cytokinesis in mammalian cells, has been shown to cause the rapid inhibition of ornithine decarboxylase and S-adenosyl methionine decarboxylase activities followed by a gradual but marked decline in intracellular levels of both putrescine and spermidine . Further, the multinucleation induced by CB could be reversed to some extent by exogenous addition of polyamines to the growth medium . The results of this study suggested that polyamines might have some indirect role in the CB-induced multinucleation of mammalian cells.

Vet Res Commun, 1981 Dec, 5(2), 183 - 5
Cell lines from sheep and cattle blood; Woldehiwet Z et al.; A simple and reproducible method of establishing cell lines from the blood of sheep and cattle is described . Buffy coat cells were allowed to adhere to plastic culture flasks in media containing 20 per cent autologous plasma overnight . The fluids were then replaced with growth medium supplemented with non-inactivated foetal calf serum, lamb serum or autologous serum . Ovine cell lines were established with any of the serum supplements but bovine cell lines established more readily if unheated autologous serum was used.

J Clin Microbiol, 1981 Dec, 14(6), 623 - 7
Comparison of liquid growth media for Legionella pneumophila; Saito A et al.; Ten liquid media were compared under standard conditions for their ability to support the growth of Legionella pneumophila . Modified gonococcal-ferric cysteine broth (without sodium chloride) supplemented with 1% yeast extract yielded the best overall growth of the one strain of L . pneumophila examined . Growth rates were independent of pH changes which occurred during incubation . The growth rates of 10 different strains of L.pneumophila were compared in this medium . There appeared to be little difference in the growth rates of strains passaged frequently or infrequently, or between environmental and clinical isolates . Moderate aeration resulted in a faster growth rate and in approximately a 1 log10 higher final cell concentration as compared to a static broth culture . These experiments demonstrate that there are moderate to marked differences among the various media described in the literature and that no liquid medium yet developed supports rapid growth of L . pneumophila incubated without shaking.

J Bacteriol, 1981 Dec, 148(3), 762 - 8
Proton translocation coupled to trimethylamine N-oxide reduction in anaerobically grown Escherichia coli; Takagi M et al.; Proton translocation coupled to trimethylamine N-oxide reduction was studied in Escherichia coli grown anaerobically in the presence of trimethylamine N-oxide . Rapid acidification of the medium was observed when trimethylamine N-oxide was added to anaerobic cell suspensions of E . coli K-10 . Acidification was sensitive to the proton conductor 3,5-di-tert-butyl-4-hydroxybenzylidenemalononitrile (SF6847) . No pH change was shown in a strain deficient in trimethylamine N-oxide reductase activity . The apparent H+/trimethylamine N-oxide ratio in cells oxidizing endogenous substrates was 3 to 4 g-ions of H+ translocated per mol of trimethylamine N-oxide added . The addition of trimethylamine N-oxide and formate to ethylenediaminetetraacetic acid-treated cell suspension caused fluorescence quenching of 3,3'-dipropylthiacarbocyanine {diS-C3-(5)}, indicating the generation of membrane potential . These results indicate that the reduction of trimethylamine N-oxide in E . coli is catalyzed by an anaerobic electron transfer system, resulting in formation of a proton motive force . Trimethylamine N-oxide reductase activity and proton extrusion were also examined in chlorate-resistant mutants . Reduction of trimethylamine N-oxide occurred in chlC, chlG, and chlE mutants, whereas chlA, chlB, and chlD mutants, which are deficient in the molybdenum cofactor, could not reduce it . Protons were extruded in chlC and chlG mutants, but not in chlA, chlB, and chlD mutants . Trimethylamine N-oxide reductase activity in a chlD mutant was restored to the wild-type level by the addition of 100 microM molybdate to the growth medium, indicating that the same molybdenum cofactor as used by nitrate reductase is required for the trimethylamine N-oxide reductase system.

Mol Cell Biol, 1981 Dec, 1(12), 1150 - 62
Membrane protein redistribution during differentiation of cultured human erythroleukemic cells; Hunt RC et al.; Human erythroleukemic (K562) cells differentiate along the erythroid differentiation pathway in vitro when 0.05 mM hemin is included in the growth medium . In the presence of the inducer the cells continue to proliferate and, after a delay of 24 to 48 h, start to synthesize hemoglobin . However, during differentiation, no changes in the major cell surface proteins were detected using lactoperoxidase-catalyzed iodination, and no change in the synthesis of spectrin, the major cytoskeletal protein of the mature erythrocyte, was detected by specific immune precipitation . Despite this absence of major changes in cell surface proteins, profound changes take place in the organization of the cell membranes . A process similar but not identical to the enucleation observed in erythroid differentiation in vivo occurs in which a smooth-surfaced cell, about 10 micrometers in diameter, is divided from the nucleus-containing part of the cell . With the exception of ribosomes, these reticulocyte-like cells contain no organelles when examined by transmission electron microscopy, but contain much of the parent cell's hemoglobin, spectrin, and glycophorin.

Acta Pathol Microbiol Scand {C}, 1981 Dec, 89(6), 353 - 8
The effect of uraemic serum and plasma on proliferating cells in vitro; Wessel-Aas T; Phytohaemagglutinin (PHA)-stimulated lymphocytes from healthy individuals and a malignant cell-line (K-562 cells) were cultured in a growth medium of 75 per cent RPMI 1640 (Gibco) supplemented with 25 per cent of uraemic serum or plasma obtained after systemic heparinization . Pooled human A Rh+ serum from 3 different donors served as controls . There was a depression of the DNA synthesis in both cell systems measured as uptake of methyl-3H-thymidine when the cells were cultured in uraemic milieu . Systemic heparinization markedly increased the depressive effect of uraemic plasma compared with controls . Heparin added to the growth media in concentrations much higher than those usually present in plasma after systemic heparinization had little effect on the cell proliferation . Systemic heparinization thus appears to induce production of toxic substances in plasma acting on proliferating cells.

Infect Immun, 1981 Dec, 34(3), 888 - 94
Protection against feline leukemia by vaccination with a subunit vaccine; Lewis MG et al.; An effective vaccine against feline leukemia virus infection has been developed by the collection and concentration of tissue culture medium harvested from a tumor cell line . Lymphoid cells were grown to near saturation density in a normal growth medium and then transferred to a serum-free medium . The serum-free medium was collected, concentrated, and evaluated for its vaccine potential . Cats receiving the vaccine emulsified in complete Freund adjuvant developed high antiviral and antitumor titers and were protected (81%) against virus challenge . Cats receiving the vaccine without an adjuvant developed lower antibody levels and lower protection (53%) from viremia . Age-matched and litter-matched controls developed no antibody to test antigens before the challenge, and 100% became persistently viremic after the challenge . Vaccination with the soluble tumor cell antigen vaccine proved successful in preventing the induction of feline leukemia virus infection.

J Biol Chem, 1981 Nov 10, 256(21), 10973 - 8
Phospholipid accumulation during the cell cycle in synchronous cultures of the yeast, Saccharomyces cerevisiae; Cottrell SF et al.; Phospholipid concentrations have been examined throughout successive cell cycles in synchronously growing cultures of the yeast, Saccharomyces cerevisiae . Total phospholipid phosphorus, as well as lecithin and phosphatidylethanolamine levels, exhibited stepwise increases during the cell cycle with step increments beginning just prior to new rounds of bud formation . Phosphatidylinositol and phosphatidylserine levels, on the other hand, showed what have been interpreted to be peak concentrations near the time of bud formation . Cardiolipin content varied considerably and was dependent upon the carbon source of the growth medium . Glucose-grown cells exhibited peak concentrations of cardiolipin near the time of bud formation, with marked decreases after this time . In contrast, galactose-grown synchronous cells exhibited stepwise increments in cardiolipin content, with step increases occurring near the time of new rounds of bud formation . Step or peak increases in cardiolipin, as well as all other phospholipids, were found to coincide with the time of stepwise increases in cytochrome c oxidase activity in these cells . No correlations were observed between the elaboration of mitochondrial membranes during the synchronous cell cycle and the observed patterns of phospholipid increase.

Mikrobiologiia, 1981 Nov-Dec, 50(6), 1046 - 52
{Effect of the age of the culture and the composition of the medium on alkaloid biosynthesis by Penicillium gorlenkoanum}; Kozlovskii AG et al.; The production of the alkaloids costaclavine and epicostaclavine by Penicillium gorlenkoanum was studied as a function of the culture age and the composition of the growth medium . The alkaloids were found in the mycelium and, in great quantities, in the cultural broth . The production of the extracellular alkaloids started from the first days of growth and run in parallel to the biomass accumulation; the synthesis of costaclavine and epicostaclavine was maximal at the end of the logarithmic-stationary growth phases . The medium supplied with mannitol, succinic acid and 1% KH2PO4 was optimal for epicostaclavine synthesis . A carbohydrate or organic acid substitution as well as a variation in the concentration of phosphate changed the proportion between epicostaclavine and costaclavine contents . The biosynthesis of epicostaclavine was inhibited almost entirely by the glucose deficiency (2%) in the medium . This glucose concentration, phosphate excess (10 g/l), and the presence of citric or tartaric acids created favourable conditions for costaclavine biosynthesis.

Mikrobiologiia, 1981 Nov-Dec, 50(6), 1042 - 5
{Comparative physiological and biochemical characteristics of Aspergillus niger isolated from the mesosphere and of its mutant}; Imshenetskii AA et al.; The aim of this work was to study certain physiological-biochemical characteristics of Aspergillus niger, strain 26, isolated from the mesosphere as well as those of its mutant having light-brown conidia . The parent strain and its mutant were grown in a liquid Chapek medium to study accumulation of the biomass, changes in the pH of the medium, as well as assimilation of glucose, nitrogen (NO3-) and phosphorus (PO4-) . The content of polysaccharides, protein, RNA and DNA was determined in the biomass . The parent culture and its mutant had the same growth dynamics and changes in the pH of the growth medium . They assimilated nitrogen, phosphorus and glucose at the same rate . No significant differences were found in the content of DNA, RNA, protein and polysaccharides . Lipids were an exception: their content was higher by ca . 26% in the mutant as compared with the parent strain . Apparently, the elevated sensitivity of the mutant to UV is due not only to a loss of certain pigments, but also to a damage of other protective mechanisms of the cell.

Mol Cell Biol, 1981 Nov, 1(11), 1007 - 15
Coordinate control of syntheses of ribosomal ribonucleic acid and ribosomal proteins during nutritional shift-up in Saccharomyces cerevisiae; Kief DR et al.; We investigated the regulation of ribosome synthesis in Saccharomyces cerevisiae growing at different rates and in response to a growth stimulus . The ribosome content and the rates of synthesis of ribosomal ribonucleic acid and of ribosomal proteins were compared in cultures growing in minimal medium with either glucose or ethanol as a carbon source . The results demonstrated that ribosome content is proportional to growth rate . Moreover, these steady-state concentrations are regulated at the level of synthesis of ribosomal precursor ribonucleic acid and of ribosomal proteins . When cultures growing on ethanol were enriched with glucose, the rate of ribosomal ribonucleic acid synthesis, measured by pulsing cells with {methyl-3H}methionine, increased by 40% within 5 min, doubled within 15 min, and reached a steady state characteristic of the new growth medium by 30 min . Labeling with {3H}leucine reveal a coordinate increase in the rate of synthesis of 30 or more ribosomal proteins as compared with that of total cellular proteins . Their synthesis was stimulated approximately 2.5-fold within 15 min and nearly 4-fold within 60 min . The data suggest that S . cerevisiae responds to a growth stimulus by preferential stimulation of the synthesis of ribosomal ribonucleic acid and ribosomal proteins.

Br J Ind Med, 1981 Nov, 38(4), 394 - 6
Cotton bacterial endotoxin assessed by electron microscopy; Helander I et al.; A piece of bale cotton was incubated in nutrient broth . Electron microscopic inspection of the cotton and the broth showed Gram-negative bacteria with long flagella, loosely attached to the cotton fibres . Large amounts of endotoxin liberating from these bacteria were visible in the growth medium.

Cancer Res, 1981 Nov, 41(11 Pt 1), 4493 - 8
Effects of deoxynucleosides on cultured human leukemia cell growth and deoxynucleotide pools; Ross DD et al.; We investigated the mechanism of cell growth inhibition caused by the deoxyribonucleosides thymidine (dThd), deoxyguanosine (dGuo), deoxyadenosine (dAdo), and deoxycytidine (dCyd) . Growth of the cultured human leukemic cells HL-60 and K-562 was measured by cloning in soft agar . Of the deoxyribonucleosides, dGuo was the most potent cell growth inhibitor; however, the potency of added dAdo was probably attenuated by the presence of adenosine deaminase in the tissue culture growth medium . The concentrations of nucleoside causing 50% inhibition of HL-60 cloning were: dCyd, greater than 10,000 microM; dAdo, 500 microM; dThd, 5,000 microM; and dGuo, 80 microM . For K-562 cloning, the concentrations causing 50% inhibition of cloning were dCyd, greater 10,000 microM; dAdo, 1,600 microM; dThd, 880 microM;' and dGuo, 100 microM . Measurement of deoxycytidine 5'-triphosphate (dCTP) pool size in HL-60 cells following incubation with 750 microM deoxyribonucleosides revealed that dGuo caused the greatest reduction of dCTP pools, both in early (passage 10)- and late (passage 71)-passage-derived HL-60 cell cultures (35 and 19% of control, respectively), compared to dThd (61 and 26% of control, respectively) and dAdo (39% of control of HL-60 passage 10) . In K-562 cells, reductions in dCTP pool size caused by dAdo, dThd, and dGuo were 68, 46, and 35% of control, respectively . Incorporation of {3H}dCyd into DNA of HL-60 and K-562 cells was enhanced by dThd and dGuo, but the degree of enhancement was greater for dThd than for dGuo . Despite its effect in reducing HL-60 dCTP pool size, dAdo failed to enhance {3H}dCyd incorporation in either HL-60 or K-562 cells . Addition of dCyd to the cultures could only partially rescue the inhibition of HL-60 cloning caused by dThd or dGuo, suggesting that inhibition of cytidine 5'-diphosphate reduction by ribonucleotide reductase is not the only mechanism whereby these nucleosides inhibit leukemic cell cloning . These data suggest that, in addition to inhibiting de novo dCTP production via ribonucleotide reductase, these nucleosides may affect other processes in the salvage pathway such as cellular uptake and phosphorylation or the DNA polymerase reaction itself.

Cancer Res, 1981 Nov, 41(11 Pt 1), 4579 - 87
Induction of the regulatory subunit of type I adenosine cyclic 3':5'-monophosphate-dependent protein kinase in differentiated N-18 mouse neuroblastoma cells; Liu AY et al.; The expression of a adenosine cyclic 3':5'-monophosphate (cAMP)-binding protein, regulatory subunit of the type I cAMP-dependent protein kinase (Rl), and its functional significance in the differentiation of N-18 mouse neuroblastoma cells were examined . 8-Azidoadenosine cyclic 3':5'-{32P}monophosphate, a photoaffinity-labeling analog of cAMP, and high-resolution sodium dodecyl sulfate-polyacrylamide gel electrophoresis were used to identify and quantitate cAMP-binding proteins in cell extracts . The induction of differentiation of N-18 mouse neuroblastoma cells, initiated either by adding dibutyryl adenosine cyclic 3':5'-monophosphate to the growth medium or by culturing cells in medium supplemented with 1% fetal calf serum, led to a 3-fold increase in the amount of 8-azidoadenosine cyclic 3':5'-{32P}monophosphate incorporated into Rl, when assayed in vitro . This increased incorporation was attributable to an increase in the amount of Rl rather than to an increase in the affinity of Rl for 8-azidoadenosine cyclic 4':5'-{32P}monophosphate . The subunit molecular weight, isoelectric point, and immunoreactivity of Rl were found to be identical to that of the regulatory subunit of the type I cAMP-dependent protein kinase purified from bovine skeletal muscle . The increase in Rl was not accompanied by an increase in the cAMP-dependent protein kinase activity . DEAE-cellulose column chromatography confirmed the induction of Rl as a free cAMP-binding protein in the differentiated neuroblastoma cells . The possibility of a growth-dependent regulation of Rl was also examined . Addition of 2% dimethyl sulfoxide to cultures of N-18 mouse neuroblastoma cells inhibited cell growth without increasing the specific activity of Rl . Dimethyl sulfoxide had no effect on neurite outgrowth or acetylcholinesterase activity, two parameters characteristic of differentiated cells . The fact that the induction of Rl coincided with differentiation of the neuroblastoma cells suggests that the expression of Rl may be used as a biochemical index of differentiation in these cells . The presence of a free cAMP-binding protein, not associated with cAMP-dependent protein kinase in neuroblastoma cells, raises important considerations concerning the action of cAMP in the regulation of growth and differentiation.

J Cell Physiol, 1981 Nov, 109(2), 309 - 15
Growth of C6 glioma cells in serum-containing medium decreases beta-adrenergic receptor number; Dibner MD et al.; Rat C6 glioma cells were grown in 5% fetal bovine serum-containing medium and under serum-free, defined conditions . In order to ask whether cells grown in serum-free medium are phenotypically identical to cells grown in serum, we examined effects of cell growth under both conditions on the beta-adrenergic receptor, a cell surface marker that activates adenylate cyclase and thereby regulates these cells . beta-Adrenergic receptors were qualitatively similar in cells grown in serum-containing and serum-free media, but the number of receptors was 30-50% less in cells grown with serum . This effect required several days to occur or to be reversed . The decreased number of receptors appeared to be caused by an inhibitory effect of serum on receptor number and not by a stimulatory action of the constituents of the serum-free medium . Growth medium with 5% fetal bovine serum maximally inhibited beta-adrenergic receptor numbers with 1% serum causing a half-maximal inhibition . The ability of serum to inhibit the expression of beta-adrenergic receptors could be blocked by dialyzing but not by boiling fetal bovine serum . beta-Adrenergic receptor-stimulated cyclic AMP accumulation was also decreased by growth in medium containing serum . These studies demonstrate that compared to growth of cells in serum-free medium, growth in serum-containing medium can inhibit expression of cell surface beta-adrenergic receptors . These results imply that the presence of serum in medium in which cells are grown alters properties in the plasma membrane and may alter hormonal responses in such cells.

Proc Natl Acad Sci U S A, 1981 Nov, 78(11), 6907 - 11
Epibolin: a protein of human plasma that supports epithelial cell movement; Stenn KS; Earlier studies suggested that there is a specific activity in mammalian serum and plasma that supports epidermal (epithelial) cell movement . This activity was shown to be nondialyzable and heat labile . In the studies reported here, using standard biochemical procedures--i.e., ammonium sulfate fractionation, ion-exchange and gel filtration chromatography, isoelectric precipitation, and preparative polyacrylamide gel electrophoresis--we have purified a factor from human plasma that supports epidermal cell movement . The factor travels as an apparent single band on disc gel electrophoresis and corresponds to a glycosylated single-chain protein of approximately 65,000 +/- 3,000 daltons . The purified fraction is necessary and sufficient for dissociated epidermal cells, (ii) outgrowth of epithelial sheets from skin explants, and (iii) epiboly, epithelial sheet movement over a floating skin explant . The purified fraction is active at a concentration of 1-2 micrograms/ml of growth medium . It is destroyed by trypsin and its activity is augmented more than 10-fold by a second, as yet unpurified, fraction of plasma . These studies support the notion that a single protein of plasma supports epidermal cell movement and that this protein may play an important role in wound closure . Because it supports epiboly, the most biologically relevant of the assays, it has been named epibolin.

J Biol Chem, 1981 Oct 25, 256(20), 10623 - 7
The mannose 6-phosphate receptor of Chinese hamster ovary cells . Compartmentalization of acid hydrolases in mutants with altered receptors; Robbins AR et al.; The localization of acid hydrolases was examined in Chinese hamster ovary cells with defective mannose 6-phosphate receptors; these mutants had been shown to exhibit reduced uptake and altered binding of exogenously added acid hydrolase (Robbins, A . R., Myerowitz, R., Youle, R . J., Murray, G . J., and Neville, D . M., Jr . (1981) J . Biol . Chem . 256, 10618-10622) . Cells were grown in the presence of {3H}mannose, alpha-L-iduronidase and beta-hexosaminidase were immunoprecipitated sequentially, electrophoresed on polyacrylamide gels containing sodium dodecyl sulfate, and detected by fluorography . About 55% of the alpha-L-iduronidase and beta-hexosaminidase synthesized by the mutants in 12 h was found in the growth medium; parental cells secreted only approximately 15% . The mutants also secreted 2 to 6 times more alpha-mannosidase, beta-glucuronidase, and alpha-L-fucosidase than the parent as determined by measurements of enzyme activity . Intracellular levels of these enzymes were reduced in the mutants . The mutants secreted acid hydrolases in the precursor forms, within the cells these enzymes resided in lysosomes and were processed normally; thus, the mutants appeared aberrant only with respect to distribution of hydrolases between intracellular and extracellular compartments . {35S}methionine-labeled beta-hexosaminidase and alpha-L-iduronidase secreted by the mutants were taken up normally by both human fibroblasts and wild type CHO cells, and this uptake was inhibited by mannose 6-phosphate . Thus, the elevated secretion of acid hydrolases was not due to alteration of the mannose 6-phosphate recognition marker on the enzymes, but appears to result from alterations in the mannose 6-phosphate receptor.

Biochim Biophys Acta, 1981 Oct 12, 677(2), 269 - 73
Constitutive behavior of methionyl-tRNA synthetase compared to repressible behavior of methionine adenosyltransferase in mammalian cells; Rubnitz JE et al.; Methionine adenosyltransferase, one of the two major enzymes utilizing methionine, is regulated by the levels of methionine in the growth medium (Jacobsen, S.J., Hoffman, R.M . and Erbe, R.W . (1980) J . Natl . Cancer Inst . 65, 1237--1244, and Caboche, M . and Mulsant, P . (1978) Somatic Cell Genet . 4, 407--421) . We report here that methionyl-tRNA synthetase, unlike methionine adenosyltransferase, behaves in a constitutive manner with respect to the concentration of methionine in the culture medium . This behavior is seen in Chinese hamster ovary cells and in normal diploid and SV40-transformed human fibroblasts . Although the kinetics of regulation of methionine adenosyltransferase and methionyl-tRNA synthetase by exogenous methionine are clearly different, the levels of the two enzymes in the human cell lines are similar.

Gann, 1981 Oct, 72(5), 717 - 22
Effect of prolonged exposure of HELA S3 cells to low concentrations of misonidazole; Ohizumi Y et al.; The cytotoxic and radiosensitizing effects of low concentrations of misonidazole were examined by measurement of the colony-forming ability and growth of HeLa S3 cells . Under hypoxic conditions, cytotoxicity depended on the misonidazole concentration and the exposure time . After 24 hr exposure, the surviving fraction of cells treated with 0.1 mM misonidazole decreased to 0.2 . The ability of cells to adhere to the growth surface of plastic dishes was reduced by 0.1 mM misonidazole upon 24 hr hypoxia and this effect persisted even during subsequent euoxia, irrespective of whether the drug was removed from the original growth medium . The growth of aerobic cells was not affected when 0.1 mM misonidazole containing medium, obtained from the hypoxia-treated dishes, was applied, nor when those cells were exposed for 72 hr to freshly prepared 1.0 mM misonidazole . Upon 24 hr exposure, 0.1 mM misonidazole exerted no obvious radiosensitizing effect on hypoxic cells . Based on these results, it is suggested that continuous administration of low doses of misonidazole may be efficacious as a specific chemotherapeutic in the treatment of hypoxic cells in tumor tissue.

Cancer Res, 1981 Oct, 41(10), 4093 - 100
Growth of human mammary epithelial cells on collagen gel surfaces; Yang NS et al.; We have developed a method for sustained growth of human mammary epithelial cells in monolayer cultures . Epithelial organoids derived from solid breast tissues were grown on the surface of thin (approximately 1 mm) collagen gel layers in an enriched growth medium supplemented with hormones, growth factors, fetal calf serum, and horse serum . To transfer the cultures, the collagen layers were dislodged and digested with collagenase . The monolayers of cells released into suspension were then dissociated into single cells using trypsin-ethylene-diaminetetraacetate . Dissociated single cells were repleted with 75 to 95% efficiency onto collagen layers or tissue culture plastic surfaces . The dissociated cells could also be cryopreserved and reactivated with greater than 80% plating efficiency on collagen layers . Normal human mammary epithelial cells grown under these conditions progressed through 12 to 15 population doublings . The population-doubling times for normal and malignant mammary cells on collagen layers were 34 and 65 hr, respectively . After reaching confluence, cells in some cultures, derived from either normal or malignant tissues, penetrated the gel surface and grew into the collagen . Within the gels, the cells became organized into three-dimensional tubular structures . The use of collagen layers eliminates a major problem in growth of human mammary epithelial cells in culture, difficulty in efficient dissociation, and cell transfer from monolayers.

J Cell Physiol, 1981 Oct, 109(1), 171 - 9
Failure of hydrocortisone or growth factors to influence the senescence of fibroblasts in a new culture system for assessing replicative lifespan; Didinsky JB et al.; It has been reported that the replicative lifespan of human fibroblasts can be substantially extended by supplementing the growth medium with hydrocortisone or increased levels of serum proteins . These observations have been made only on cell populations transferred many times at high cell density, and cumulative population doublings have been recorded, rather than a more direct measure of cell division potential . We have measured the replicative potential of human fibroblasts cultured so as to avoid conditions of high cell density, medium depletion, and departure from exponential growth . Two fetal lung and two newborn foreskin fibroblast strains were serially passaged in the presence or absence of hydrocortisone (HC), epidermal growth factor (EGF), and fibroblast growth factor (FGF) until they senesced . At each passage cells were plated at densities sufficiently low that colony-forming efficiency could be calculated . We determined cumulative population doublings and also estimated the number of cell generations attained under each condition . FGF caused small but possibly significant changes, while HC and EGF failed to substantially alter replicative lifespan . The reported effect of HC on the doubling potential of fetal lung fibroblasts is therefore not an inevitable action of this hormone on the senescence mechanism, but may instead depend for its apparent activity on the passage regimen used . The fibroblast's insensitivity to EGF as a modulator of replicative potential, as compared with the keratinocyte, whose lifespan can be tripled by EGF, implies that the mechanisms limiting the replicative potential of these two cell types are not identical.

J Cell Physiol, 1981 Oct, 109(1), 1 - 15
Isolation of the major serine protease inhibitor from the 5-day serum-free conditioned medium of human embryonic lung cells and demonstration that it is fetuin; Rohrlich ST et al.; Human embryonic lung (HuEL) cells in culture produce large amounts of the enzyme, plasminogen activator, and thus generate substantial amounts of active plasmin . Despite the presence of plasmin, however, HuEL cells grow in ordered, flattened, adherent sheets . It seemed of interest to characterize protease inhibitors that might be present in HuEL cultures and which might account for this apparent contradiction . This paper reports the isolation and purification of the major serine protease inhibitor in 5-day serum-free conditioned medium (CM) from HuEL cells, and the purification of an identical molecule from fetal bovine serum (FBS) . Both the CM-derived inhibitor and the FBS-derived inhibitor are identical with fetuin, the major glycoprotein of FBS . The CM-derived inhibitor is apparently derived from the FBS used to supplement the growth medium of HuEL cells between serum-free CM collection periods . It is not labeled metabolically with 3H-leucine . Its electrophoretic behavior is indistinguishable from that of standard fetuin in SDS-PAGE, non-SDS basic pH,PAGE, and isoelectric focusing . The CM-derived inhibitor and standard fetuin inhibit trypsin and plasmin with similar efficiencies, but neither inhibits chymotrypsin, pancreatic elastase, or plasminogen activator . They are immunologically indistinguishable . The suggestion is made that fetuin, and possibly other protease inhibitors present in HuEL cell cultures, may be concentrated locally by HuEL cells and gradually released back into the medium in the absence of serum . These molecules may serve to protect HuEL cells against proteases they generate.

Biull Eksp Biol Med, 1981 Oct, 91(10), 460 - 2
{Interferons produced by the cells of newborn mice in the presence of sera from newborn and adult animals}; Malinovskaia VV et al.; Fibroblasts of newborn mice produced far less amount of interferon in the presence of sera from newborn animals than in the presence of sera from adult animals . The interferons obtained were purified by adsorption chromatography on porous glass and were analyzed by electrophoresis in polyacrylamide gel . It has been shown that antiviral activity of interferon preparations obtained in the presence of sera from newborn mice was associated with the fraction of 45 Kd . Addition into the growth medium of sera from adult animals led to the production by the same cells of interferon activity associated with 41 and 28 Kd fractions . It is assumed that the sera of newborn mice contained the components influencing the molecular content of interferon produced by the cells of newborn animals.

Hoppe Seylers Z Physiol Chem, 1981 Sep, 362(9), 1219 - 27
{Degradation of L-phenylalanine and of aromatic carboxylic acids by chloridazon-degrading bacteria . Combination of side chain degradation and dioxygenase pathway}; Wegst W et al.; Strain N of Chloridazon-degrading bacteria degrades phenylalanine via cis-2,3-dihydro-2,3-dihydroxyphenylalanine,2,3-dihydroxyphenylalanine aspartate and 4-hydroxy-2-oxovalerate {Hoppe-Seyler's Z . Physiol . Chem . 360, 957--969, (1979); Biochem . J . 194, 679--684 (1981)} . cis-2,3-Dihydro-2,3-dihydroxyphenylalanine and 2,3-dihydroxyphenylalanine as well as phenylpyruvate, cis-2,3-dihydro-2,3-dihydroxyphenylpyruvate, 2,3-dihydroxyphenylpyruvate, cis-2,3-dihydro-2,3-dihydroxyphenylacetate, 2,3-dihydroxyphenylacetate and 2,3-dihydroxybenzaldehyde are detectable in the medium of strain E during growth on phenylalanine . Incubation with phenylacetate, 3-phenylpropionate or 4-phenylbutyrate leads to the accumulation of the corresponding cis-2,3-dihydro-2,3-dihydroxyphenyl derivatives . These compounds are transformed with dihydrodiol dehydrogenase to 2,3-dihydroxyphenylacetate, 3-(2,3-dihydroxyphenyl)propionate and 4-(2,3-dihydroxyphenyl)-butyrate, 3-(2,3-dihydroxyphenyl)propionate is attacked by a catechol 2,3-dioxygenase and the meta-cleavage product is again cleaved by a hydrolase yielding succinate . In a similar reaction sequence the degradation of 4-phenylbutyrate leads to the formation of glutarate . From the growth medium of strain E on phenylacetate also small amounts of 2-, 3- and 4-hydroxyphenylacetate were isolated . Resting cells were shown to metabolize 3- and 4-hydroxyphenylacetate via homogentisate and 3,4-dihydroxyphenylacetate . In the culture medium of strain K2AP benzoate could be detected . Pathways for the degradation of phenylalanine and aromatic carboxylic acids in chloridazon degrading bacteria are proposed.

J Virol, 1981 Sep, 39(3), 903 - 13
Characterization of intracellular and extracellular vaccinia virus variants: N1-isonicotinoyl-N2-3-methyl-4-chlorobenzoylhydrazine interferes with cytoplasmic virus dissemination and release; Hiller G et al.; Infectious vaccinia virus can be purified from whole cells by experimentally induced lysis (intracellular virus) or from supernatant growth medium (extracellular virus) . Extracellular virus and intracellular virus differed by buoyant density (1.237 versus 1.272 g/cm3), phospholipid content and composition, and polypeptide pattern . Differences in structural polypeptides on the virus surface could be detected by lactoperoxidase-catalyzed radioiodination or Brij treatment . Characteristic of extracellular virus was an additional polypeptide, with a molecular weight of 37,000 (37K), which represented 5 to 7% of the total particle protein . Antibodies to the 37K protein detected only some of the cell-associated particles late in normal infection . Upon treatment of infected cultures with N1-isonicotinoyl-N2-3-methyl-4-chlorobenzoylhydrazine, a drug which prevents vaccinia virus release, no particle-associated 37K protein could be detected . In all other properties tested so far, except for a slight difference in phospholipid composition, the virus obtained in the presence of the drug resembled the normal intracellular virus . N1-Isonicotinoyl-N2-3-methyl-4-chlorobenzoylhydrazine prevented vesicularization of intracellular viral particles . Lack of vesicularization was accompanied by the absence of particle-associated 37K viral protein and seemed to correlate with an inhibition of virus dissemination to the cell periphery.

Chem Biol Interact, 1981 Sep, 36(3), 259 - 74
Binding of methylmercury(II) by HeLa S3 suspension-culture cells: intracellular methylmercury levels and their effect on DNA replication and protein synthesis; Gruenwedel DW et al.; HeLa S3 cells were exposed to varied concentrations of methylmercury over varied periods of time and its binding by the cells was studied using 203Hg-labeled methylmercuric chloride as radioactive marker . Also studied was the effect of cell-bound methylmercury on DNA replication and protein synthesis and on the growth rate of the cells . The results show that methyl-mercury binding is a rapid process, with much of the organomercurial bound within the the first 60 min of incubation, and that considerable quantities of organic mercury become affixed to the cells . The amounts of bound methylmercury, {CH3Hg(II)}bound, given in mol/cell, range from 2 X 10(-16) (at 1 h of incubation and at 1 microM CH3Hg(II) in the medium) to almost 4 X 10(-14) (at 24 h of incubation and at 100 microM CH3Hg(II) in the medium) . A {CH3Hg(II)}bound value of about 30 X 10(-16) mol/cell appears to be the threshold below which cells display a normal growth pattern and below which metabolic events such as DNA replication or protein synthesis are affected only to a minor degree but above which major changes in cell metabolism and cell growth take place . Methylmercury binding by the cells is tight so that only 20% of the bound material is released from the cells over a 3-h incubation period when the cells are placed into fresh, methylmercury-free growth medium . Analysis of the binding data in terms of binding to identical and completely independent sites yields an association constant K of 7.92 X 10(4) l/mol and for the maximum concentration of cellular binding sites the value 2.40 X 10(-14) mol/cell or 1.45 X 10(10) sites/cell . Evidence is presented which shows that cellular sulfhydryl groups do not suffice to provide all the sites taken up by methylmercury and that binding, in all likelihood, involves basic nitrogen, too . The levels of cell-bound methylmercury are such that binding to HeLa DNA and HeLa chromatin, for instance, can readily take place . Methylmercury binding data obtained by using the technique of particle-induced X-ray emission (PIXE) are in good agreement with the data obtained via isotope dilution.

Cancer Res, 1981 Sep, 41(9 Pt 1), 3592 - 6
Growth and differentiation of human and murine erythroleukemia cell lines in serum-free synthetic medium; Pessano S et al.; Only two chemicals (transferrin and selenium dioxide) are required to supplement serum-free Roswell Park Memorial Institute Medium 1640 for long-term growth and for spontaneous and induced differentiation of established lines of human and mouse erythroleukemia cells . We describe here two serum-free media (a minimal synthetic medium and a high-density synthetic medium) that support the growth and differentiation of human K562(S) and mouse clones 745, 707, and 3TCl 12 erythroleukemia cell lines in long-term culture . The doubling times of the erythroleukemic cell populations are longer in minimal synthetic medium that in serum-containing medium . Cell saturation density in minimal growth medium is one-half that obtained in serum-containing medium for clone 745, whereas for K562(S) it is approximately the same . Cell saturation density in high-density medium (containing albumin) is greater than that achieved in serum-containing medium for K562(S), whereas for clone 745 cell saturation density increases for cells in midlogarithmic growth, although not to the density of cells grown in serum-containing medium . The differences in saturation density are due to a decreased doubling time as well as to better survival of the cells 3 or 4 days after plating . The cells can grow in the synthetic media and be passaged for as many generations as desired without impairment of growth capabilities . In the minimal synthetic medium, spontaneous differentiation of erythroleukemia cells continues to occur, indicating that spontaneously differentiating cells are the result of intracellular mechanisms controlling the expression of a genetic program of some of the cells at any given time . Hemoglobin synthesis can be induced in cells growing in synthetic medium by using lower concentrations of the same inducers that are effective in serum-containing medium, indicating that these chemicals do not depend on serum factors to initiate the process of differentiation . The percentage of benzidine-positive cells and the concentration of hemoglobin per cell, however, are less in the synthetic medium than in serum-containing medium, suggesting that serum factors do play a role in modulating the extent of hemoglobin synthesis . The types of hemoglobins synthesized by cells in synthetic medium are identical to those reported in serum-containing medium.

Cell, 1981 Sep, 25(3), 773 - 82
Growth-rate-dependent regulation of ribosome synthesis in E . coli: expression of the lacZ and galK genes fused to ribosomal promoters; Miura A et al.; Hybrid transducing phages were constructed in vitro that carry the galK gene fused to each of three ribosomal promoters: the promotor for an rRNA operon (rrnE); the promoter for the spec r protein operon and the promotor for the alpha r protein operon . We also constructed hybrid transducing phages that carry the IacZ gene fused to the promoter for the rrnE operon or to the promoter for the spc r protein operon . The amounts of galactokinase (or beta-galactosidase) were analyzed in lysogens carrying these various transducing phages grown in several different growth media . The synthesis rate of galactokinase (or beta-galactosidase) from the fused rrn-gal (or rrn-lac) operon relative to the total protein synthesis rate increased with increasing growth rate, as expected from the transcriptional activity of rRNA operons . In contrast, the relative synthesis rate of galactokinase (or beta-galactosidase) from the operon fused to alpha or spc r protein promoter remained approximately constant with increasing growth rate . These results were interpreted to mean that the characteristic increase in the relative synthesis rate of r protein with increasing growth rate is determined not by transcription regulatory mechanisms, but by posttranscriptional mechanisms, which presumably involve the feedback inhibition of r protein mRNA translation by free r proteins.

J Neuropathol Exp Neurol, 1981 Sep, 40(5), 512 - 25
Growth characteristics of human glioma-derived and fetal neural cells in culture; Icard C et al.; Growth characteristics of human fetal neural cells (CH) and human glioblastoma multiforme-derived cells (12-18) in culture were compared . Cells were grown to confluent densities of 38,000 to 42,500 cells/cm2 for CH and 85,800 to 87,100 for 12-18 . Population doubling times were 40.0 +/- 5.1 hr and 66.5 +/- 9.8 hr for CH and 12-18 cells, respectively . The mean DNA content per cell of the glioma-derived cells was twice that of the fetal brain cells at sparse, log, and confluent cell densities . High concentrations (40%) of serum in growth medium increased DNA contents in confluent CH, but not 12-18, cells . The amount of protein per cell also was consistently higher in glioma cells than CH cells, but, as cell densities increased, protein contents decreased for both: 1200 to 700 pg/cell in glioma cells, and 840 to 560 pg/cell in CH cells . In each cell line, initial rates of {3H}ThdR incorporation into TCA precipitable material decreased as cell density increased, but confluent glioma-derived cells incorporated 10 times more {3H}ThdR than confluent fetal cells . Almost all CH cells had a normal diploid chromosome number of 46 . A histogram showing the relative frequencies of chromosome numbers of glioma-derived cells had peaks of 52, 79, and 105 chromosomes per metaphase, indicating a haploid number of 26 for most cells . Lengths of cell cycle phases, determined using autoradiographic techniques, indicate that glioma-derived cells had a longer generation time and S period than fetal neural cells . These data demonstrate several biological differences between glioblastoma-derived cells and non-neoplastic fetal neural cells, indicating that this system is of potential value for comparative studies on growth control and contact inhibition.

Acta Virol, 1981 Sep, 25(5), 257 - 65
Resistance of Semliki forest virus protein synthesis to high salt treatment; Erdei S; Selective translation of Semliki Forest virus-specific mRNA occurred in virus-infected cells exposed to hypertonic growth medium . The selective resistance of the virus-specific protein synthesis could be detected at a wide NaCl concentration range and was more significant at lowered incubation temperature (28 degrees C) . It is suggested that the translation of the structural proteins encoding subgenomic 26 S RNA is more resistant to the hypertonic initiation block than the translation of the genomic 42 S RNA which codes for the non-structural viral proteins.

Biochemistry, 1981 Sep 1, 20(18), 5336 - 40
Biosynthesis of o-succinylbenzoic acid in a men- Escherichia coli mutant requires decarboxylation of L-glutamate at the C-1 position; Meganathan R et al.; A men- mutant of Escherichia coli, AN 209, which accumulates o-succinylbenzoic acid, has been used for a direct study of the biosynthesis of this benzenoid compound . Samples of labeled glutamic acids were added to growth media, and the o-succinylbenzoic acid was isolated and converted to a dimethyl derivative . This dimethyl derivative was purified on thin-layer chromatograms and by gas chromatography . When the glutamic acid used as precursor contained 14C at position 5, or was uniformly labeled, the dimethyl o-succinylbenzoate contained radioactivity (as shown by radiogas chromatography) . However, from {1-14C}glutamate, the dimethyl o-succinylbenzoate was without radioactivity . Hence, in the biosynthesis of o-succinylbenzoate, carbon atom 1 of glutamate is lost, and carbon atoms 2-5 are retained . It was also shown that this mutant lacked the enzyme dihydroxynaphthoic acid synthase . It should, therefore, continue to be classified as a menB mutant, rather than as a member of the newly created menE group (lacking o-succinylbenzoate-CoA synthetase).

Brain Res, 1981 Aug 31, 219(2), 407 - 21
Growth of adult superior cervical ganglion explants in serum-free media; Dombrowski AM et al.; Neurite outgrowth from explants of superior cervical ganglion from adult rats can be achieved in a serum-free medium . Extensive neurite outgrowth occurred from ganglion explants maintained in Eagle's minimum essential medium supplemented with either 10% (V/V) fetal calf serum or 1% (W/V) bovine serum albumin and nerve growth factor . After one week in culture, the ATP content of explants maintained in the serum-free medium was slightly higher than that noted in explants cultured in the presence of fetal calf serum and amounts of phosphocreatine were significantly lower . Despite these differences in high energy phosphate content, the abundance and morphology of neuritic outgrowth were essentially the same from explants cultured in the two types of media . Comparable activities of a number of NADP+-dependent dehydrogenases were noted in explants maintained in the two types of media . Increases in the activities of the oxidative enzymes of the pentose pathway, which occur in axotomized ganglia in vivo, were observed in the cultured ganglion explants . NADP+-dependent isocitrate dehydrogenase activity remained constant in ganglion explants in vitro, and measurements of this activity were employed in a new method to quantitate neurite outgrowth . The activity of isocitrate dehydrogenase in lyophilized neurite processes that had grown out onto a Millipore filter substrate correlated well with visual estimates of neuritic outgrowth . Substitution of delipidated for normal bovine serum albumin in the growth medium resulted in a significant decrease in neuritic outgrowth from ganglion explants from both adult and weanling rats . Addition of fatty acids to media containing delipidated bovine serum albumin enhanced neuritic outgrowth in explants of weanling rats . Thus, lipophilic substances bound to bovine serum albumin including fatty acids appear necessary for optimal growth of neurites from explants of the rat superior cervical ganglion.

Biochim Biophys Acta, 1981 Aug 17, 676(2), 221 - 5
High performance liquid chromatographic analysis of catecholamines in growing and non-growing Tetrahymena pyriformis; Goldman ME et al.; Tetrahymena pyriformis, strains NT-1 and W, harvested in logarithmic (growing) and stationary (non-growing) phases, were found by high-performance liquid chromatography to contain considerable quantities of dopamine . In addition, small amounts of epinephrine and norepinephrine were detected . Logarithmic-phase strain NT-1 cells contained 249 +/- 44 pg dopamine/10(6) cells compared to 477 +/- 42 pg/10(6) cells for logarithmic-phase strain W cells for logarithmic-phase strain W cells . The dopamine content of stationary-phase cells was approximately half the value of the logarithmic-phase cells . There was a significant amount of dopamine in the growth medium from stationary-phase cultures and, to a lesser extent, logarithmic-phase cells.

Biochim Biophys Acta, 1981 Aug 13, 660(2), 371 - 4
Regulation of enterochelin synthetase in Escherichia coli K-12; Greenwood KT et al.; Enterochelin synthetase activity is controlled by both repression and feed-back inhibition mechanisms . Inclusion of iron in growth media results in synthesis of all four (D, E, F and G) components of enterochelin synthetase being repressed . The specific inhibition of L-serine activation (partial reaction catalyzed by the F component) by the end products, ferric-enterochelin and 2,3-dihydroxybenzoylserine, is shown to inhibit overall enterochelin synthetase activity.

Biochim Biophys Acta, 1981 Aug 5, 676(1), 91 - 100
Cyclic nucleotide metabolism in differentiated and undifferentiated epithelial thyroid cells in culture; Mandato E et al.; A highly differentiated thyroid cell line (FR-RL) was compared with a less differentiated (FR-T Cl1) and an undifferentiated (1-5G) cell line . FR-TL is modulated in vivo and in vitro by thyrotropin and has the lowest adenylate cyclase and guanylate cyclase and the highest phosphodiesterase activities . In contrast, 1-5G tumor cells do not respond to thyrotropin and have the highest adenylate cyclase guanylate cyclase and lowest hydrolyzing enzyme activities . Intermediate enzyme activities were found in FR-T Cl1 cells . The differences between the two normal rat thyroid cell lines are not due to differences in the composition of the growth medium.

J Invest Dermatol, 1981 Aug, 77(2), 221 - 4
Induction of lymphocyte differentiation by epidermal cultures; Rubenfeld MR et al.; Human and murine lymphoid cell populations were induced to express terminal deoxynucleotidyl transferase, a marker of early lymphoid differentiation, by exposure to allogeneic or syngeneic epidermal cells . Control growth medium, fibroblasts, or a mammary epithelial cell line did not induce this marker . These findings suggest that epidermal cells can induce lymphoid cell differentiation in vitro.

Br J Cancer, 1981 Aug, 44(2), 219 - 35
Cytotoxic and clastogenic effects of soluble and insoluble compounds containing hexavalent and trivalent chromium; Levis AG et al.; Cr(III) and Cr(VI) compounds of varying solubilities have been tested in vitro for their ability to inhibit cell growth and nucleic acid and protein syntheses in BHK cells, to induce alterations in the mitotic cycle in HEp cells, and to increase the frequency of chromosomal aberrations and sister chromatid exchanges (SCE) in CHO cells . All Cr(VI) compounds, and particularly those containing soluble Cr(VI), such as potassium dichromate and zinc yellow, differentially inhibit macromolecular syntheses in BKH cells, that of DNA being always the most affected . Among Cr(III) compounds, which generally have very low cytotoxicity, chromite is particularly active, and inhibits cell growth and DNA synthesis even more than the poorly soluble Cr(VI) compounds . Preincubation in growth medium, with or without metabolizing cell cultures, solubilizes considerable amounts of Cr(VI) from zinc yellow and chromite, but significant amounts are also obtained from the most insoluble Cr(VI) pigments . When BHK cells are treated with such preincubated solutions, reduction of soluble Cr(VI) to Cr(III) by cell metabolites is seen with all Cr(VI) compounds, accompanied by decreased cytotoxicity . The same differences between Cr(VI) and Cr(III) compounds apply to the cytotoxic effects on mitosis of HEp cells and the clastogenic effects on CHO cells . The activity of chromite, the only Cr(III) pigment capable of significantly increasing the frequency of SCE, is due to contamination with soluble Cr(VI) . In contrast to the very low cytotoxicity of Cr(III), much higher chromium levels are detected in the cells incubated with soluble Cr(III) than with the same concentrations of soluble Cr(VI) . 50% and 75% of chromium accumulated in the cells during treatments with Cr(VI) and Cr(III) respectively remains firmly bound to the cells, even when they are incubated for up to 48 h in normal growth medium . Chromium accumulated in the cells after treatment with Cr(III) is most probably bound to the cell membrane, whereas some of the Cr(VI) is transported through the cell membrane and reduced in the cell nucleus . The results of the present investigation are in agreement with those obtained with the same Cr(VI) and Cr(III) compounds in mutagenicity assays in bacteria and carcinogenicity tests in rodents . A re-evaluation of the mechanisms of chromium carcinogenisis is proposed.

Am J Hosp Pharm, 1981 Aug, 38(8), 1144 - 7
Effect of laminar air flow and clean-room dress on contamination rates of intravenous admixtures; Brier KL et al.; The effect of laminar air flow conditions and clean-room dress on the microbial contamination rates of intravenous admixtures was investigated . Intravenous admixtures were prepared by one investigator using aseptic technique under four environmental conditions: laminar air flow conditions with clean-room dress; laminar air flow without clean-room dress; clean table top with clean-room dress; and clean table top without clean-room dress . In each environmental condition, 350 admixtures were compounded . Negative-control samples (n = 150) were also tested, as were 10 positive-control samples . Samples were tested in each of two growth media and incubated at 35 degrees C for 14 days or until growth occurred . The incidence of contamination of admixtures compounded in laminar air flow conditions was significantly less than the contamination of those compounded on a clean table top (p less than 0.05) regardless of the operator's dress . The incidence of contamination of admixtures compounded while wearing clean-room dress was not significantly different from those prepared while not wearing clean-room dress regardless of the environment in which the admixture was prepared . The overall low level of contamination {0.79% (11/1400)} was inconclusive regarding the effect of dress on the incidence of contamination when admixtures were prepared under LAF conditions . It is concluded that, when one adheres to aseptic technique, the environment in which admixtures are compounded is the most important variable affecting the microbial contamination rate.

J Bacteriol, 1981 Aug, 147(2), 373 - 81
Dihydroxy and monohydroxy fatty acids in Legionella pneumophila; Mayberry WR; Five strains of Legionella pneumophila were examined for the presence of hydroxy fatty acid . The cellular distribution of the fatty acids was also determined, as was the variation of hydroxy acid production on five growth media . The strains tested all produced approximately 5 mol% of hydroxy fatty acid, most of which was found in the nonextractable, alkali-stable, acid-labile (wall-associated, amide-linked) fraction . Three major hydroxy acids were found, along with several minor components . The major hydroxy acids were analyzed by thin-layer chromatography, gas-liquid chromatography, mass spectrometry, and infrared spectrophotometry . These compounds were tentatively identified as 3-hydroxy-12-methyltridecanoate, 3-hydroxy-n-eicosanoate, and a novel dihydroxy acid, 2,3-dihydroxy-12-methyltridecanoate . The total amount of hydroxy acid produced, as well as the profile of the hydroxy acids, remained relatively unchanged with respect to strain and growth medium.

Appl Environ Microbiol, 1981 Aug, 42(2), 200 - 3
Serum substitute in epithelial cell culture media: nonfat dry milk filtrate; Fassolitis AC et al.; A number of milk types and milk fractions were investigated as possible substitutes for serum in cell culture media . A filtrate of reconstituted nonfat dry milk showed promise . Culture fluids containing 5% of the nonfat dry milk filtrate were used to propagate primary and continuous cell cultures, and the cell growth from these cultures was compared with that of cells grown in a serum-containing medium . The nonfat dry milk filtrate-supplemented medium supported the growth of all epithelial cells tested, but two fibroblast-type cultures failed to replicate . Cells grown in the medium containing the milk filtrate grew slowly for 2 to 3 days and then propagated to confluency in 6 to 8 days . Viable cell counts of 9 days were comparable to those of serum-grown cells that had been propagated for 7 days . Cells grown in the milk filtrate could be split 1 to 4 when subcultures were prepared . Cell growth could be stimulated by refeeding on days 2 to 3 or by the addition of 30 microM 2-mercaptoethanol to the growth medium . Virus susceptibility and titer comparisons with poliovirus 1, coxsackievirus B2, echovirus 7, and herpes simplex virus indicated that approximately the same data were obtained when either the nonfat dry milk filtrate-treated or the serum-treated cells were studied . The nonfat dry milk filtrate is inexpensive, is easily prepared, and is a substitute for serum in epithelial cell culture media.

J Biol Chem, 1981 Jul 25, 256(14), 7628 - 37
The cyclic nucleotide phosphodiesterase inhibitory protein of Dictyostelium discoideum . Purification and characterization; Franke J et al.; We have purified the glycoprotein inhibitor of the extracellular cyclic nucleotide phosphodiesterase of Dictyostelium discoideum to apparent homogeneity . The inhibitor has a molecular weight of 47,000 measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis . The interaction of the inhibitor and the cyclic nucleotide phosphodiesterase occurs with 1:1 stoichiometry and with a dissociation constant of about 10(-10) M . Periodate oxidation of the inhibitor or of the enzyme destroys concanavalin A binding ability but does not affect the formation of the enzyme-inhibitor complex . Inhibitor is not produced by cells during logarithmic growth but appears in quantity during stationary phase and after transfer from growth medium to phosphate buffer.

Int J Cancer, 1981 Jul 15, 28(1), 51 - 7
Characterization of clones of tumorigenic feline cells transformed by a chemical carcinogen; Rhim JS et al.; Tumorigenic clonal lines derived from soft agar colonies induced by DMBA-transformed feline embryo cells were isolated and characterized . The morphologically altered clonal cells formed large aggregates, growing in this aggregate form when suspended in liquid growth medium above an agar base and forming colonies in soft agar with high efficiency . When inoculated into athymic nude mice, chemically altered clonal cells produced progressively growing sarcomas . Cells established from the tumors morphologically resembled the DMBA-transformed feline embryo cells and were characterized as cat cells by karyological analysis . The tumorigenic lines were negative for feline oncornavirus-associated cell membrane antigen (FOCMA), and for "gag-X" the transformation-related polyprotein which is encoded by the replication defective feline sarcoma virus.

Mol Cell Biol, 1981 Jul, 1(7), 652 - 60
Sensitivity of a mutator gene in Chinese hamster ovary cell to deoxynucleoside triphosphate pool alterations; Meuth M; The Thy- mutants of Chinese hamster ovary cells have a 5- to 10-fold elevated pool of deoxycytidine 5'-triphosphate (dCTP) and are auxotrophic for thymidine as an apparent consequence of a single mutation . thy is also a mutator gene, elevating the spontaneous rate of mutation 5- to 200-fold for at least two genetic markers . Previous experiments suggested that this mutator activity was caused by the elevated pool of dCTP in Thy- cells . To test this, the dCTP and deoxythymidine 5'-triphosphate (dTTP) pools were manipulated by altering the external concentration of thymidine in the growth medium . The rate of mutation at one genetic locus, ouabain resistance, was directly related to cellular dCTP content . At the highest level of dCTP the rate in one Thy- strain was approximately 200 times that of wild-type cells . However, the relationship between dCTP content and the rate of mutation at the ouabain locus was different for two mutator strains and wild-type cells . The rate of mutation at a second locus, thioguanine resistance, was increased approximately 10-fold over wild type regardless of the dCTP-dTTP pools . These experiments suggest that the mutator activity of thy is clearly related to dCTP content, but the dCTP level alone does not appear to be the cause of the mutator.

Vopr Virusol, 1981 Jul-Aug, (4), 452 - 6
{Acute and chronic Lassa virus infection of Vero cells}; Lukashevich IS et al.; Acute and persistent infections of Vero cells with a cloned Lassa virus were studied . After acute infection at a multiplicity of 2 PFU/cell the latent period was 12-16 hrs and the maximum yield of virus particles in the growth medium was obtained 28-32 hours postinfection . A persistently infected cell line was established by inoculating Vero cells with the cloned Lassa virus followed by subcultivation of the infected cells . The infected cells underwent 36 passages . Lassa virus production by the infected cells followed a cyclic pattern varying from 10(2) to 10(5) PFU/ml . Neither the plating efficiency of the virus nor its growth curves changes at 35 degrees C and 39 degrees C . The replication of the cloned virus in infected cells (superinfection) was markedly depressed in comparison to that of normal Vero cells infected with Lassa virus . The role of defective interfering particles in the establishment of persistent infection is discussed.

Chem Biol Interact, 1981 Jul, 36(1), 45 - 59
Ultraviolet radiation-induced neoplastic transformation of normal human cells, in vitro; Milo GE et al.; Human foreskin cell cultures in scheduled DNA synthesis (S phase) of the cell cycle were exposed to UV irradiation at a dose of 10 J . m-1 in the presence of insulin . These treated cell populations, when selectively passaged in a high amino acid supplemented complete growth medium (CM) after 20 Dulbecco's phosphate buffered saline (pH 6.8) (PDL), were able to be grown in soft agar . These treated cell populations were also grown in 1% serum supplemented growth medium and at 41 degrees C in 10% serum supplemented growth medium . Cell populations 4--5 PDL after treatment exhibited altered colony morphology and altered lectin agglutination profiles but would not grow in soft agar . These events appeared to be associated with the early stages in the expression phase of the transformed phenotype . After 20 PDL, we observed that these cells would grow in soft agar at a frequency of 20 colonies/10(5) cells seeded in soft agar . The cell populations derived from these colonies, when propagated and injected into the nude mice, formed myxofibromas at the injection sites rather than the type of tumor (fibrosarcoma) previously described for chemical carcinogen-induced neoplasms.

Can J Microbiol, 1981 Jul, 27(7), 670 - 4
Lipid level and total fatty acid composition for selected developmental stages of Entomophthora egressa; Dunphy GB et al.; The major fatty acids (greater than 10%) of Entomophthora egressa were C16:0 and C18:1 . Minor fatty acids, which varied with the stage of fungal development, included C11:0, C12:0, C13:0, C14:0, C15:0, C16;1, C17:0, C18:0, C18:2, C18:3 C:201, C20:2, C20:3, C20:4, C20:5 and two unidentified unsaturated fatty acids . Differences were observed between the total fatty acid levels of C12:0, C14:0, C17:0, C18:0, and C20:5 and the degree of unsaturation of the fatty acids of 37-h protoplasts grown in modified Grace's medium and a simplified growth medium (SGM) . The levels of C12:0, C14:0, C18:1, C20:4, and C20:5 decreased and the levels of C18:0 and C20:2 increased with the formation of spherical hyphal body (shb)initials . With the production of mature shb increased levels of C12:0, C14:0, C15:0, C18:1, C20:4, and C20:5 were detected . During the germination of the shb the levels of C14:0, C16:1, C18;1, and C20:4 increased, whereas C15:0 and C20:5 levels declined . The fatty acid levels, except for C12:0, C13:0, and C20:2, remained constant during the mycelial stage . The degree of fatty acid unsaturation decreased during early stages of development (protoplasts through shb initials) . In SGM the degree of fatty acid unsaturation was lowest during the shb initial stage and highest during the shb stage . The total lipid level increased during shb maturation and declined during shb germination.

Strahlentherapie, 1981 Jul, 157(7), 468 - 73
{Action of x-rays and neuraminidase on pinocytotic activity and microtubule-microfilament system of Ehrlich ascites tumor cells in monolayer culture (author's transl)}; Pfab R et al.; Monolayer cultures of Ehrlich ascites tumor cells in exponential growth phase were treated with X-rays and neuraminidase alone or in combination . A radiation dose of 2 Gy (200 rd) and higher effected a significant inhibition of DNA synthesis and cell proliferation . Neuraminidase treatment in addition to irradiation did not modify the growth-inhibitory irradiation effect . Cells pretreated for 0.5 to 1.0 hours with neuraminidase (in Earle's salt solution) and cultured thereafter (4h) in serum-containing growth medium exhibited an enhanced development of the microtubule-microfilament-system . Morphometric analysis of microtubules on electronmicroscopic sections revealed a nearly 4-fold increased number in neuraminidase treated cells while the average length of microtubules was similar to controls . Pinocytosis as measured by the ultrastructural uptake of ferritin appeared to be increased following neuraminidase treatment . A connection between neuraminic acid containing components (glycoproteins or glycolipids?) of the cell membrane or cell surface, on the one hand, and the cytoskeleton and the endocytotic process of these tumor cells, on the other hand, is suggested.

Biochim Biophys Acta, 1981 Jul, 675(2), 178 - 87
Comparative role of polyamines in division and plastid differentiation of Euglena gracilis; Schuber F et al.; Regulation of polyamine biosynthesis during growth and differentiation of Euglena gracilis was investigated . Increased activity of L-ornithine decarboxylase (EC 4.1.1.17), the enzyme which catalyzes the initial step in polyamine synthesis in Euglena, and accumulation of polyamines were observed prior to DNA replication in synchronous cultures of heterotrophically or photoautotrophically grown cells . In photoautotrophic cells three maxima of polyamine synthesis were observed during the light period of the cell cycle . The transition form quiescence to active growth was accompanied in heterotrophic Euglena by a very large stimulation of ornithine decarboxylase activity and polyamine synthesis; the decrease in growth potential of these cells was correlated with a decrease in polyamine levels . In contrast, differentiation of Euglena i.e . a shift from heterotrophic to photoautotrophic mode of living in the absence of division, led only to a minor stimulation of polyamine biosynthesis . Alpha-Methylornithine, an inhibitor of ornithine decarboxylase, blocked the growth of heterotrophic Euglena, and depletion of intracellular polyamines decreased the differentiation rate . Both events could be reversed by addition of putrescine to the growth medium . This study suggests that Euglena requires a minimal intracellular level of polyamines to grow and differentiate under optimal conditions . This requirement seems to be more stringent for cell division.

Cancer Res, 1981 Jul, 41(7), 2611 - 5
Release of glycosyltransferase and glycosidase activities from normal and transformed cell lines; Klohs WD et al.; The release of galactosyltransferase, sialyltransferase, and several glycosidase activities into the growth media from several normal and transformed cell lines was examined . Six of the seven cell lines released galactosyltransferase into their culture media . Only the human leukemia CCRF-CEM cells failed to release demonstrable galactosyltransferase activity . Release of galactosyltransferase activity into the media closely paralleled the growth curves for all but the BHKpy cells . These cells continued to release peak levels of galactosyltransferase activity into the culture media after their growth had plateaued . Media galactosyltransferase activity was unaffected by Triton X-100 treatment had remained in the supernatant fraction of a 100,000 X g, 12-hr centrifugation, suggesting that the cells release galactosyltransferase in a soluble form . In contrast to galactosyltransferase activity, only one of the cell lines (L1210) released sialyltransferase activity in appreciable amounts . Even this level of activity was 20-fold less than that observed for galactosyltransferase in the media from L1210 cells . Of the nine glycosidase activities assayed, only N-acetylglucosaminidase was observed in significant amounts in the media from all but the CCRF-CEM cells . However, N-acetylglucosaminidase release did not correlate closely with cell growth . These findings suggest a relatively specific release of galactosyltransferase and N-acetylglucosaminidase activities by cells in tissue culture . Moreover, the release of galactosyltransferase closely parallels cell growth . The significance of these released enzymes, especially to cell growth, has yet to be determined.

Eur J Biochem, 1981 Jul, 117(2), 431 - 7
Citrate-dependent iron transport system in Escherichia coli K-12; Hussein S et al.; Induction of the citrate-dependent iron transport system of Escherichia coli K-12 required 0.1 mM citrate and 0.1 micrometer iron in the growth medium . Five--ten-times more iron than citrate was taken up into the cells which suggests that citrate was largely excluded from the transport . Fluorocitrate and phosphocitrate induced the citrate-dependent iron transport system although they supported iron uptake only very poorly . An outer membrane protein (FecA), belonging to the transport system, was induced in fecB mutants which were devoid of citrate-dependent iron transport . The intracellular citrate and iron concentrations were 10--100-times higher than the external concentrations required for induction of the transport system . It is concluded that only exogenous ferric citrate induced the transport system, and that citrate did not have to enter the cytoplasm . The Tn10 transposon, conferring tetracycline resistance, was inserted near the fec gene region which controls the expression of the citrate-dependent iron transport system . The determination of the cotransduction frequencies of Tn10 with the fecA and fecB markers suggested the gene order fecA fecB Tn10.

Thromb Haemost, 1981 Jun 30, 45(3), 219 - 24
Secretion of plasminogen activators by cultured bovine endothelial cells: partial purification, characterization and evidence for multiple forms; Laug WE; Endothelial cells were obtained from the aortae of newborn calves and cloned . High plasminogen activator (PA) activity was detected in the supernatant medium and the cell lysates of confluent cultures . The PA activity in the growth medium increased steadily during 12 hrs of incubation, indicating active enzyme secretion by these cells . Sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis of the concentrated medium demonstrated the presence of four plasminogen activators with apparent molecular weights of 77,000 (+/- 3000), 43,000 (+/- 2000), 26,000 (+/- 1500) and 14,500 (+/- 1500) respectively . The 43,000, 26,000 and 14,500 molecular weight forms could be converted to radioactive derivatives by active site labeling with 3H diisopropyl fluorophosphate (3H DFP) while the 77,000 Dalton form took up only traces of this radioactively labeled compound . The 43,000 molecular weight form was partially purified by means of salt precipitation and gel filtration . This enzyme preparation activated plasminogen by proteolytic cleavage with maximum activity at pH 7.5-8.5 and demonstrated a specific activity of 80,000 CA (Committee on Thrombolytic Agents) units/mg protein when tested on 125I-fibrin in the presence of plasminogen . This PA was rapidly and irreversibly inhibited by diisopropyl fluorophosphate (DFP), suggesting that it was a serine protease . The partially purified enzyme was extremely labile at temperatures from 0-60 degrees C, but could be stabilized by lowering the pH to 3 or by the addition of albumin.

J Biol Chem, 1981 Jun 25, 256(12), 6304 - 10
Evidence for coordinate expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase ad low density lipoprotein binding activity; Chin J et al.; A Chinese hamster ovary cell mutant that requires both cholesterol and unsaturated fatty acids for growth (Limanek, J . S., Chin, J., and Chang, T . Y . (1978) Proc . Natl . Acad . Sci . U . S . A . 75, 5452-5456) has been further characterized with respect to its dependence on cholesterol . Upon removal of serum lipids from the growth medium, the activity of the important cholesterogenic enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and the low density lipoprotein (LDL) binding activity both increase significantly in the normal cell . Both these increases were much less in the mutant cell . Studies in vitro with NaF indicate that the differences in reductase activities between normal and mutant cells are not due to differences in activation by a dephosphorylation mechanism . Heat inactivation profiles and Km for HMG-CoA of both cell reductases were found to be identical, thus reducing the possibility that the mutant cell contains a mutation in the polypeptide chain of reductase . The fact that in lipid-deficient medium both reductase and LDL binding activities are low in the mutant strongly suggests that the expression of these activities is controlled in a coordinate manner . This conclusion is supported by parallel studies on a spontaneous revertant of the mutant in which the expression of reductase and LDL binding activities have both reverted to normal . These results indicate that the phenotypic abnormalities seen in the mutant are probably caused by a single mutation . A common factor is postulated to mediate this coordinate expression, and the function of such a factor is altered in the mutant cell.

J Biol Chem, 1981 Jun 25, 256(12), 6174 - 80
Evidence indicating that inactivation of 3-hydroxy-3-methylglutaryl coenzyme A reductase by low density lipoprotein or by 25-hydroxycholesterol requires mediator protein(s) with rapid turnover rate; Chang TY et al.; The half-life (t 1/2) of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase of Chinese hamster ovary cells grown in fetal calf serum medium is approximately 2 h . When cells are switched to grow in delipidated serum medium (DeL-M) for more than 24 h, the t 1/2 of the enzyme is found to be drastically altered to approximately 13 h . Exposure of low density lipoprotein (LDL) (100 micrograms of protein/ml) or 25-hydroxycholesterol (1 microgram/ml) to cells grown in DeL-M suppresses reductase activity more rapidly than would be expected solely if reductase synthesis were suppressed, showing that inactivation of reductase activity by sterols, previously demonstrated using only analogs of cholesterol, is a normal mechanism for regulation of HMG-CoA reductase activity by the physiologically important sterol source (LDL) . This inactivation effect by LDL or by 25-hydroxycholesterol is shown to be at least in part due to acceleration of reductase degradation rate . Furthermore, the inactivation effect by sterols is shown to be largely abolished if cycloheximide (250 micrograms/ml) is added simultaneously to the growth medium, indicating that continuous synthesis of a class of mediator protein(s) is necessary in mediating the effect of LDL or 25-hydroxycholesterol . Two different protein synthesis inhibitors (emetine and puromycin) were used and gave essentially identical results . Preincubation of cell culture with cycloheximide for 2 h essentially completely abolishes the effect of 25-hydroxycholesterol, indicating that the mediator protein(s) turns over rapidly, with t 1/2 less than 3 or 4 h.

Eur J Biochem, 1981 Jun, 117(1), 147 - 53
The organization of cholesterol esters in membranes of Mycoplasma capricolum; Melchior DL et al.; The organization of cholesterol esters in Mycoplasma capricolum membranes was studied by differential scanning calorimetry . Cells grown in the presence of horse serum incorporated large amounts of cholesterol esters into their membranes . The cholesterolester-containing membranes after incubation at low temperature showed an endotherm characteristic of a cholesterol ester crystalline leads to isotropic liquid transition that was identical in membranes both before and after thermal protein denaturation . This transition was not observed in membranes of cells grown in medium in which the horse serum was replaced by bovine albumin, fatty acids and unesterified cholesterol unless cholesterol esters were added to the growth medium . In membrane preparations obtained both from cells grown in horse serum and from cells grown with bovine albumin plus cholesterol and fatty acids, the free cholesterol content was sufficient to eliminate the bilayer order/disorder transition observed in isolated membrane phospholipids . Our studies indicate that the majority of cholesterol esters in M . capricolum membranes is not present in attached serum lipoprotein particles, nor is intimately associated with membrane protein, but exists as relatively large cholesterol ester droplets or pockets tightly associated with the membrane . The cholesterol esters in these pockets appear relatively pure, although the presence of small amounts of other membrane components is likely.

J Cell Physiol, 1981 Jun, 107(3), 427 - 38
The biotin requirement of HeLa cells; Dakshinamurti K et al.; We have examined the effect of alterations in the biotin content of the medium on the growth, viability, biotin content, and the activities of biotin-independent and biotin-independent enzymes of the HeLa cells . The inclusion in the growth medium of avidin, which almost irreversibly binds with biotin (Kd, 10(-15) M), results in an increase in cellular biotin content and biotin enzyme activity over that seen when the cells are grown in a biotin-depleted medium . The addition of avidin-bound biotin to the growth medium led to a forty-fold increase in cellular biotin when compared to the inclusion of an equivalent amount of free biotin in the medium . HeLa cells are able to internalize avidin-bound biotin . Biotin is released from this complex to function as the prosthetic group of biotin enzymes . HeLa cells do have a nutritional requirement for biotin.

J Invest Dermatol, 1981 Jun, 76(6), 474 - 9
In vitro alterations of epidermal cell adhesion induced by temperature, substrate, and cations; Patel H et al.; Epidermal cell-to-cell or basal cell-to-substrate adhesion in vitro, involves as yet, unknown mechanisms . The rate of attachment of newly dissociated neonatal mouse epidermal cells and the rate of keratinocyte detachment from preformed epidermal monolayers was investigated under a variety of experimental conditions . In the present investigation, we obtained the following results: (1) variables such as temperature, nature of substrate, and presence of cations in the growth medium were important in the initial cel-to-substrate attachment; (2) removal of Ca++ and Mg++ from the growth medium was associated with a very low attachment rate (less than 5% at 24 hr and 48 hr); (3) the initial cell-to-substrate attachment of epidermal cells decreased about 50% when maintained in medium deficient in either Mg++ or Ca++ indicating that both cations are important in the cell-to-substrate attachment; (4) keratinocyte-detachment from preformed monolayers increased in medium deficient in Ca++, Mg++ as well as in medium only deficient in Ca++ . However, detachment in cultures maintained in Mg++ deficient medium was similar to controls . In conclusion, Ca++ is one factor that may regulate epidermal cell-to-cell and cell-to-substrate interactions in vitro . Mg++ appears to influence also in the initial attachment process of epidermal basal cells.

Br J Pharmacol, 1981 Jun, 73(2), 333 - 40
Down-regulation of the sodium pump following chronic exposure of HeLa cells and chick embryo heart cells to ouabain; Aiton JF et al.; 1 HeLa cells and primary cultures of embryonic chick heart cells were grown in medium containing low concentrations of ouabain for 24 h . 2 Compared with normal cells, cells grown in ouabain have fewer free sodium pump sites, an increased intracellular sodium concentration and a decreased intracellular potassium concentration . The cells are able to maintain their intracellular ion contents because the remaining pump sites have an increased turnover rate . 3 When cells that have been chronically exposed to ouabain are returned to normal growth medium, the sodium pump site numbers increase; the recovery process begins within 6 to 8 h and is complete within 24 h . Recovery of pump site numbers is primarily dependent upon de novo protein synthesis since the protein synthesis inhibitor, cycloheximide, prevents recovery.

J Bacteriol, 1981 Jun, 146(3), 1124 - 34
Long-chain fatty acid perturbations in Mycoplasma pneumoniae; Leon O et al.; The fatty acid content of Mycoplasma pneumoniae increased 2.5- to 9.6-fold when the growth medium was supplemented with a saturated, unsaturated, or beta-hydroxy fatty acid, the greatest increase occurring with palmitic acid . The amount of each supplemented fatty acid found within this organism was 2.8 to 5.5% of the total fatty acid content; the exception was palmitic acid . Up to 57% of the palmitic acid was utilized from the supplemented medium, whereas only 0.2 to 10% of the other fatty acids was utilized . Chromatographic and isotopic analyses revealed that 22% of the labeled palmitic acid incorporated from the palmitic acid-supplemented medium remained free in this organism . Also, even though complex lipid synthesis increased a minimum of 3.8-fold under these conditions, this mycoplasma continued to incorporate intact complex lipids from the growth medium . Bacteriostatic and bactericidal studies which used high concentrations of various long-chain fatty acids showed that only palmitic, myristic, and beta-hydroxydecanoic acids were not bactericidal . The addition of palmitic acid to the growth medium resulted in the formation of exceedingly long, filamentous cells in approximately 25% of the population . Osmotic fragility and electron spin resonance spectroscopy studies showed a correlation among this increased fatty acid content, decreased membrane fluidity, and the increased osmotic fragility of palmitic acid-grown cells . In addition, these cells had a lowered cholesterol content . The effect of such compositional changes on osmotic fragility is discussed in this paper . Finally, the profound increase in the total fatty acid content of palmitic acid-grown cells altered neither sensitivity to tetracycline or erythromycin nor the amount of hydrogen peroxide secreted.

Gann, 1981 Jun, 72(3), 395 - 402
Potentiation of bleomycin cytotoxicity toward cultured mouse cells by hyperthermia and ethanol; Mizuno S et al.; The effects of hyperthermia and ethanol on bleomycin cytotoxicity toward mouse mammary carcinoma FM3A cells in culture were followed in terms of clonal growth in a soft agar medium . Heating the cells at 43 degrees during or before bleomycin treatment sensitized them to bleomycin . Heat treatment after drug treatment was less effective . Bleomycin cytotoxicity was also greatly potentiated by exposing the cells to ethanol at a concentration of more than 4% (v/v) at 37 degrees before or after drug treatment . However, when the cells were exposed to ethanol during bleomycin treatment, the potentiation of cytotoxicity was reduced, and a higher concentration of ethanol was required for inducing equivalent sensitization . The cytoxicity of pepleomycin, a potent antitumor agent derived from bleomycin, was also greatly potentiated by hyperthermia and ethanol . The cellular uptake of 3H-pepleomycin was not increased by either treatment, and thus the increased cytotoxicity was not correlated with a change in membrane permeability to the drug . The sensitizing effect of hyperthermia or ethanol was maximum when the drug treatment was carried out immediately after the cells were exposed to heat or ethanol . Sensitization decreased linearly as the time interval between the two treatments was increased, and after a 6 hr interval the sensitizing effect of either agent disappeared . This recovery from the sensitized state was not inhibited by cycloheximide or by incubation in phosphate-buffered saline instead of complete growth medium, but it was reversibly inhibited at 0 degrees . It is suggested that the modes of action of hyperthermia and ethanol in inducing cell sensitization of bleomycin are similar.

Biochim Biophys Acta, 1981 May 18, 674(3), 372 - 82
Purification of a high-affinity discoidin I-binding proteoglycan from axenic Dictyostelium discoideum growth medium; Bartles JR et al.; The axenic Dictyostelium discoideum growth medium HL-5, prepared using Difco proteose peptone No . 2, contains an extremely potent inhibitor of the binding of 125I-labeled discoidin I to glutaraldehyde-fixed, cohesive D . discoideum cells . Axenic strain A3 D . discoideum cells bind or internalize the inhibitor during growth in HL-5 medium and subsequently shed or excrete it while differentiating in suspension . The inhibitor has been purified from Difco proteose peptone No . 2 by sequential gel filtration on Sepharose 4B and affinity adsorption using discoidin I-Sepharose . The inhibitor is heterogeneous in molecular weight (4 . 10(5)--2 . 10(6)), but is relatively homogeneous in density on CsCl density gradients . The size and activity of the inhibitor are resistant to periodate, reduction and maleylation, proteases, nucleases and heating in the absence or presence of sodium dodecyl sulfate . Mild alkali causes a partial reduction in activity and converts the higher molecular weight fraction of the inhibitor to a lower molecular weight . The purified inhibitor contains neutral hexose, hexosamine and amino acid in an approximate molar ratio of 4 : 3 : 2 . These and other properties suggest that the inhibitor is an unusual proteoglycan . Certain well-characterized glycosaminoglycans are relatively potent inhibitors of discoidin I binding . The proteoglycan reported here is the most potent discoidin I-binding inhibitor ever identified.

Biochem J, 1981 May 15, 196(2), 403 - 10
Growth-rate-dependent adjustment of ribosome function in the fungus Mucor racemosus; Orlowski M; The dimorphic fungus Mucor racemosus was grown at rates between 0.043 and 0.434 doubling/h while maintained as yeasts or at rates between 0.21 and 0.50 doubling/h while maintained as hyphae by altering the composition of the growth medium or the gaseous environment of the cells . Yeasts at the higher growth rates contained many more ribosomes than did yeasts at the lower growth rates . They also had a higher percentage of ribosomes active in protein synthesis and a faster rate of polypeptide-chain elongation than did the slower-growing cells . Hyphal cells at faster growth rates also contained many more ribosomes and showed a faster rate of polypeptide-chain elongation than did slower-growing cells . However, the faster-growing cells had a substantially lower proportion of ribosomes active in protein synthesis than did the slower-growing hyphae . Pulse-chase experiments failed to provide any evidence of protein turnover, which might otherwise invalidate the values calculated for the peptide-chain elongation rates.

J Biol Chem, 1981 May 10, 256(9), 4146 - 9
First observation of amino acid side chain dynamics in membrane proteins using high field deuterium nuclear magnetic resonance spectroscopy; Kinsey RA et al.; We have obtained the first deuterium NMR spectra of an individual membrane protein, bacteriorhodopsin in the purple membrane of Halobacterium halobium R1 . Biosynthetic isotopic enrichment with {gamma-2H6}valine and high field Fourier transform operation permitted rapid data acquisition on intact membranes, including measurement of relaxation times . At some temperatures high quality spectra could be obtained in less than 1 s . {U-14C}Valine tracer studies indicate that less than or equal to 2% of valine added to the growth medium is broken down and incorporated into other membrane constituents . The NMR results indicate that the valine side chain is a rather rigid structure . Motion about C alpha-C beta is slow (less than 10(5) s-1) at growth temperature, While motion about C beta-C gamma is as expected fast (much greater than 10(5) s-1) at all accessible temperatures . The activation energy for methyl group rotation from spin-lattice relaxation data between -75 and 53 degrees C is approximately 2.4 kcal/mol, in good agreement with previous 1H NMR studies on solid alkanes . Preliminary data on {gamma-2H6}valine-labeled Acholeplasma laidlawii B (PG9) cell membranes are also presented . Our results strongly suggest that it should now be possible to observe in great detail the motions of any type of amino acid side chain in membrane proteins, including the effects of lipid composition on protein dynamics.

Mikrobiologiia, 1981 May-Jun, 50(3), 453 - 7
{Endo-N-acetylglucosaminidase formed by Streptomyces levoris}; Shchelkova NS et al.; Endo-N-acetylglucoseaminidase (EC 3.2.1.30) was prepared from the cultural broth of Streptomyces levoris 96 using precipitation with ammonium sulfate, gel filtration on Sephadex G-25 and ion exchange chromatography on DEAE-cellulose . The isoelectric point of the enzyme is 4.2 Str . levoris produces various quantities of the enzyme depending on the composition of the growth medium . A medium in which the enzyme is produced in maximal amounts has been selected . The effect of peptidoglycan, cell walls and chitin added to the medium on the enzyme biosynthesis was studied . Chitin induced biosynthesis of the enzyme by 35--45%.

Arch Microbiol, 1981 May, 129(3), 216 - 20
Concentration of metabolites and the regulation of phosphofructokinase and fructose-1,6-bisphosphatase in Saccharomyces cerevisiae; Foy JJ et al.; The intracellular levels of adenosine triphosphate and several glycolytic intermediates were determined in Saccharomyces cerevisiae in relation to the presence of the metabolically antagonistic enzymes phosphofructokinase and fructose-1,6-bisphosphatase . Phosphofructokinase is synthesized constitutively in cells grown in the presence of glucose and fructose-1,6-bisphosphatase derepression occurs upon the exhaustion of glucose from the growth medium . Transcriptional regulation of fructose-1,6-bisphosphatase was suggested based on experiments with wild type cells using 8-hydroxyquinoline, a known inhibitor of nuclear transcription, and with the S . cerevisiae mutant strain A364A (ts-136) blocked in the transport of nuclear RNA at non-permissive temperature . The level of phosphofructokinase was reduced more than 25-fold under conditions of high citrate accumulation in an aconitase-less, glutamate requiring mutant strain, MO-1-9B . There was a rapid decrease in the levels of adenosine triphosphate and fructose-1,6-bisphosphate at the end of log-phase of culture growth when both fructose-1,6-bisphosphatase and phosphofructokinase were present in the cells simultaneously . The changes in the levels of key glycolytic intermediates, but not the changes in adenosine triphosphate, during the simultaneous presence of these two enzymes, can be explained without involving any futile cycling.

J Cell Physiol, 1981 May, 107(2), 295 - 302
Regulation of glucose utilization in chick embryo fibroblasts by bicarbonate ion; Miller MH et al.; The amount of glucose consumed by chick embryo fibroblasts in primary culture is strongly influenced by the presence of bicarbonate ion in the culture medium . Cells grown on glucose at physiologic concentration (5.5 mm) and in the absence of bicarbonate ion have a reduced rate of glucose utilization when compared to their counterparts cultivated in medium containing the usual 25 mM bicarbonate . The presence or absence of bicarbonate is without effect on chick embryo fibroblast proliferation over a 6-day growth period . Both lactic acid accumulation per mole of glucose consumed and the utilization of glutamine increase as a function of bicarbonate ion in the growth medium.

Biochim Biophys Acta, 1981 Apr 27, 653(2), 248 - 58
Dependence of mammalian DNA synthesis on DNA supercoiling . III . Characterization of the inhibition of replicative and repair-type DNA synthesis by novobiocin and nalidixic acid; Mattern MR et al.; Novobiocin and nalidixic acid, inhibitors of the bacterial enzyme DNA gyrase, inhibit DNA, RNA and protein synthesis in several human and rodent cell lines . The sensitivity of DNA synthesis (both replicative and repair) to inhibition by novobiocin and nalidixic acid is greater than that of protein synthesis . Novobiocin inhibits RNA synthesis about half as effectively as it does DNA synthesis, whereas nalidixic acid inhibits both equally well . Replicative DNA synthesis, as measured by incorporation of {3H}thymidine, is blocked by novobiocin in a number of cell strains; the inhibition is reversible with respect to both DNA synthesis and cell killing, and continues for as long as 20--30 h if the cells are kept in novobiocin-containing growth medium . Both novobiocin and nalidixic acid inhibit repair DNA synthesis (measured by BND-cellulose chromatography) induced by ultraviolet light or N-methyl-N'-nitro-N-nitrosoguanidine (but not that induced by methyl methanesulfonate) at lower concentration (as low as 5 micrograms/ml) than those required to inhibit replicative DNA synthesis (50 micrograms/ml or greater) . Neither novobiocin nor nalidixic acid alone induces DNA repair synthesis . Incubation of ultraviolet-irradiated cells with 10--100 micrograms/ml novobiocin results in little, if any, further reduction of colony-forming ability (beyond that caused by the ultraviolet irradiation) . Novobiocin at sufficiently low concentrations (200 micrograms/ml) apparently generates a quiescent state (in terms of cellular DNA metabolism) from which recovery is possible . Under more drastic conditions of time in contact with cells and concentration, however, novobiocin itself induces mammalian cell killing.

Thymus, 1981 Apr, 2(6), 339 - 53
A 35 000 dalton T-cell supernatant protein associated with suppression of mitogenic responses; Demartino JL et al.; Synthesis of a CEM T-lymphoblast supernatant protein of 35 000 daltons (p35), which is scarcely expressed on CEM membranes, increased 8-fold when cells were grown in crowded cultures . Synthesis of p35 was more dependent upon crowding of cells on the flask bottom (maximum at 2.0 X 10(6) cells/cm2) than upon cell density in the culture medium (maximum at 3.0 X 10(6) cells/ml) . Growth medium containing p35 inhibited the response of normal peripheral blood lymphocytes to PHA and to Con A . Supernatant media lacking p35, collected from low-density CEM cultures of from RAJI B lymphoblastoid cells which were grown under low or high-density conditions, did not suppress mitogenic responses of normal lymphocytes . These experiments support the view that the p35 T-cell supernatant protein plays a regulatory role in cell proliferation and possibly in the immune system.

J Bacteriol, 1981 Apr, 146(1), 163 - 9
Alternate mechanism for amino acid entry into Neurospora crassa: extracellular deamination and subsequent keto acid transport; DeBusk RM et al.; The growth of the pm nbg mutant strain of Neurospora crassa was inhibited by the amino acid analog para-fluorophenylalanine despite the fact that none of the three constitutive amino acid permeases is functional in this strain . This observation led to the detection of both a deaminase which was released into the growth medium in response to para-fluorophenylalanine and a keto acid transport system which allowed entry of the resulting keto acid into the cell . The transported keto acid was recovered in cellular protein, suggesting its regeneration as the amino acid . The cooperative activity of these two systems represents an additional mechanism for the intracellular accumulation of amino acids, which is distinct from the known amino acid permeases.

Proc Natl Acad Sci U S A, 1981 Apr, 78(4), 2616 - 9
Herpes simplex virus (type 1) thymidine kinase gene does not transform cells morphologically; Hampar B et al.; BALB/c-derived 10E2 cells were made thymidine kinase(TK)-negative and one isolated clone (B2) was used for studying morphological and biochemical transformations by herpes simplex virus (HSV) type 1 (strain 10412) . The B2 cells displayed a "normal" flat appearance and were nontumorigenic in nude mice when tested at frequent intervals over a period of 45 subcultures . B2 cells infected with UV-irradiated HSV (UV-HSV) and maintained in normal growth medium showed foci of spindle-shaped cells after one subculture . The cells from these morphologically transformed foci were tumorigenic in nude mice and were TK negative . B2 cells infected with UV-HSV or transfected with the HSV-1 TK gene and maintained in TK-selective medium showed discrete colonies of cells which displayed a normal flat appearance and expressed the viral TK enzyme . These biochemically transformed B2 cells were nontumorigenic in nude mice . The findings with B2 cells indicate that biochemical and morphological transformations by HSV-1 are independent events and suggest that the HSV-1 TK gene is a suitable vehicle for introducing non-TK genes into cells to assess their transforming potential.

Biochemistry, 1981 Mar 31, 20(7), 1826 - 31
Structure of Escherichia coli membranes . Glycerol auxotrophs as a tool for the analysis of the phospholipid head-group region by deuterium magentic resonance; Gally HU et al.; Glycerol selectively deuterated at various positions was synthesized and supplied to the growth medium of Escherichia coli strain T131 GP, which is defective in endogenous glycerol synthesis as well as glycerol degradation and lacks the ability to synthesize cardiolipin . The procedure enables the stereospecific labeling of the membrane phospholipids (approximately 80% phosphatidylethanolamine, approximately 20% phosphatdylglycerol) . Deuterium magnetic resonance spectra were obtained for cell membranes and lipid dispersions either from total lipid extractions or from purified phosphitidylglycerol or -ethanolamine . When glycerol deuterated at various positions was used, all resonances of the phospholipid glycerol backbone and the terminal glycerol moiety in phosphatidylglycerol could be assigned . The results indicate that the molecular conformation of the glycerol backbone is independent of the phospholipid species investigated and is also not altered by the presence of high amounts of membrane proteins . For the quantitative interpretation of the deuterium magnetic resonance splittings, a model is proposed which assumes essentially free rotation around the glycerol C(2)-C(3) bond combined with an asymmetric and restricted jump process around the C(1)-C(2) bond . This model is compatible with known X-ray structures of phospholipids molecules . The two deuterons of both the glycerol backbone C(1) and C(3) segments were found to be magnetically inequivalent . Stereoselective monodeuteration eliminated one set of quadrupole splittings in both cases.

Biochem J, 1981 Mar 15, 194(3), 707 - 11
Intracellular ionic changes in normal and transformed human fibroblasts after extracellular Ca2+ deprivation; Hazelton BJ et al.; The lowering of extracellular Ca2+ concentration in the growth medium reversibly blocks normal, but not SV40-transformed WI38 diploid fibroblasts in the early G1/G0 phase of the cell cycle . This growth response is characterized by specific changes in ionic content and transport . Ca2+ deprivation (0.03 mM) has little effect on the K+ content of either normal or transformed cells . Na+ content, however, is increased nearly 2-fold in the normal cells . This increase is presumably due to a 3-fold increase in unidirectional Na+ influx in Ca2+-deprived cells . The increased intracellular Na+ also gives rise to a nearly 3-fold enhancement of the active (ouabain-sensitive) Na+ efflux . Ca2+ deprivation causes only slight increases in Na+ influx, ouabain-sensitive Na+ efflux and intracellular Na+ in the transformed cell . In contrast, the transformed cells lose nearly 60% of their intracellular Ca2+ on deprivation, whereas normal WI38 cells lose only 10% . The data suggest that the growth arrest exhibited by the normal cell but not the transformed cell may be related to different membrane-transport and permeability changes in response to Ca2+ deprivation.

Can J Microbiol, 1981 Mar, 27(3), 350 - 7
Growth characteristics af low Na+ concentration and the stability of the Na+ requirement of a marine bacterium; Gow JA et al.; Studies of the marine bacterium Alteromonas haloplanktis 214 (formerly referred to as marine pseudomonad B-16) showed that as the Na+ concentration in the growth medium decreased from 230 to 34 mM, the lowest concentration permitting growth, the length of the lag period preceding exponential growth increased . Once growth had begun, except for a slight reduction in rate of growth at 34 mM Na+, the generation time and extent of growth remained essentially constant over the range of Na+ concentrations tested . Plate counts showed that during the lag period the numbers of viable cells introduced as inoculum into a complex medium containing 33 mM Na+ decreased exponentially before increasing . Repeated subculture of the cells at 33 mM Na+ failed to eliminate the lag period or reduce the loss of viability of the cells . The viability loss and the lag period could be eliminated either by raising the NaCl concentration to 130 mM or by adding sufficient sucrose to make the osmotic pressure of the medium equal to that obtained by adding 130 mM NaCl . In a chemically defined medium, sucrose added to maintain tonicity reduced but did not eliminate the lag periods obtained at suboptimal Na+ concentrations . Increasing the number of cells plated on trypticase agar medium reduced the Na+ concentration required to permit growth . Evidence was obtained of a requirement of A . haloplanktis for Ca2+ for growth . Ca2+ spared to a small extent the requirement for Na+ for growth . Some 10(10) cells of a histidine-requiring, streptomycin-resistant mutant of A . haloplanktis 214, still viable after treatment with N-methyl-N'-nitro-N-nitrosoguanidine, were screened for capacity to grow in the absence of Na+ . Since no non-Na+-requiring mutants were isolated, the requirement of this organism for Na+ would appear to be extremely stable.

Invest Urol, 1981 Mar, 18(5), 364 - 70
Environmental alteration and two distinct mechanisms of E . coli adherence to bladder epithelial cells; Avots-Avotins AE et al.; We used an in vitro model to investigate Escherichia coli attachment to transitional epithelial cells obtained from bladders of female rats . Adhesive abilities and sensitivity to mannose inhibition differed among the isolates for both epithelial cells and erythrocytes . Adherence of some strains could be modulated by bacterial washes and growth media . Variations in adhesiveness were related to bacterial piliation as determined by transmission electron microscopy . With two strains, mannose inhibition of adherence to epithelial cells was dose-related; however, with a maximal inhibitory dose, adherence was reduced by approximately 80 per cent even when the bacteria-to-epithelial cell ratio was varied . These studies show that adhesiveness and piliation of certain adhesive E . coli strains are either reduced or enhanced by environmental alterations . We conclude that E . coli strains adhere to epithelial cells by at least two distinct mechanisms and that a single isolate may utilize both mechanisms . The more efficient process is pili-mediated and inhibited by mannose whereas undetermined surface components mediate the less efficient but mannose-resistant mechanism.

J Biol Chem, 1981 Feb 10, 256(3), 1167 - 71
Investigations on myelination in vitro . Regulation of sulfolipid synthesis by thyroid hormone in cultures of dissociated brain cells from embryonic mice; Bhat NR et al.; L-3,5,3'-Triiodothyronine (T3) has been shown to influence the synthesis of myelin-associated lipids in cultures of cells dissociated from brains of embryonic mice (Bhat, N . R., Sarlieve, L., Subba Rao, G., and Pieringer, R . A . (1979) J . Biol . Chem . 254, 9342-9344) . This culture system was used in the present study to gain additional information on the regulation of the synthesis of myelin lipids by thyroid hormone . The rate of synthesis of the myelin associated sulfolipids remained drastically diminished throughout a 70-day developmental period when cells were grown in the presence of hypothyroid calf serum (T3 < 25 ng/100 ml; thyroxine (T4), 1.2 microgram/ml) . However, the activity could be restored to normal levels after 72 h of exposure to deficient medium supplemented with exogenous T3 . Half-maximal effects were obtained with 2 X 10(-9) M T3 and 6.25 X 10(-7) M T4 . T3 does not alter the synthesis of sulfated mucopolysaccharides, which share adenosine 3'-phosphate, 5'-phosphosulfate (PAPS), as a common precursor, with sulfolipids . This observation argues against the hormone altering the entry of sulfate or the synthesis of PAPS . Rather, T3 acts by changing the activity of the glycolipid:PAPS sulfotransferase(s) in direct proportion to the concentration of T3 in the growth medium . The activity of 2',3'-cyclic nucleotide 3'-phosphohydrolase, another myelin marker was also found to be T3 dependent . The response of sulfolipid synthesis to varying amounts of T3 was also observed in a serum-free medium, which suggests that T3 can function independently of other hormones and serum factors in exerting a relatively specific effect on the regulation of myelination.

Lipids, 1981 Feb, 16(2), 146 - 8
Chaulmoogric acid: assimilation into the complex lipids of mycobacteria; Cabot MC et al.; Lipid analysis of Mycobacterium vaccae, grown in the presence of chaulmoogric acid, demonstrates that this cyclopentenyl fatty acid is taken up by the organism and incorporated into cellular phospholipids and triacylglycerols . As cell growth is retarded by the addition of chaulmoogric acid to the growth medium, it is possible that the antimicrobial properties of this compound results from a perturbation of membrane processes.

Endocrinology, 1981 Feb, 108(2), 573 - 83
Normal mammary cells in long term culture . I . development of hormone-dependent functional monolayer cultures and assay of alpha-lactalbumin production; Ray DB et al.; Mammary cells from normal tissue of virginal, pregnant, and lactating rats have been adapted to long term monolayer culture in plastic culture dishes with retention of hormone-responsive functional activity . The addition of PRL, insulin, and corticosterone resulted in an increase in the proportion of epithelial cells . the development of intracellular lipid droplets, and the ordered aggregations of these cells . In the presence of these hormones, the milk protein, alpha-lactalbumin (a-LA), was secreted into the growth medium at rates of 20-100 ng/mg cellular protein . 24 h . A double antibody RIA for a-LA capable of measuring 0.1 ng a-LA/100 microliter growth medium was developed in our laboratory for these studies . Both intracellular and extracellular a-LA fell below detectability within 2-3 weeks after hormone withdrawal . Intracellular a-LA accounted for less than 3% of the total a-LA accumulated in each culture in 24 h . the production rate of cells continuously given hormones increased 4- to 7-fold over a period of several months in culture, and their output was greater than 100-fold above that of cells not given hormones . These cells were obtained by overnight digestion and dispersion of tissue using selected batches of collagenase in the presence of 5% fetal calf serum . Plating densities of at least 3 X 10(4) cells/cm2 in Minimum Essential Medium supplemented with 14% fetal calf serum were required for optimal functional activity . Despite several months without added hormones, these cultures can retain their hormone responsiveness, since subsequent hormone addition resulted in detectable a-LA production beginning within 7-14 days . Our studies demonstrate for the first time that normal mammary cells can be maintained in a functional hormone-responsive state for extended periods in primary cell culture . These long term cell cultures provide a system with which the effects of these and other hormones on milk production and cell differentiation can be assessed under conditions which minimize the influence of the prior in vivo hormonal millieu . (Endocrinology 108: 573, 1981)

Biophys J, 1981 Feb, 33(2), 211 - 23
A biophysical study of protein-lipid interactions in membranes of Escherichia coli . Fluoromyristic acid as a probe; Gent MP et al.; Fluorine-19 nuclear magentic resonance spectroscopy and transport assays have been used to investigate and compare the membrane properties of unsaturated fatty acid auxotrophs of two strains of Escherichia coli, K1060B5 and ML 308-225-UFA-8 . A fluorinated analog of myristic acid, 8, 8-difluoromyristic acid, can be incorporated into the membrane phospholipids by substitution for oleate in the growth medium . Growth for one generation on 8, 8-difluoromyristate results in a 20% content of fluorinated fatty acid in the membranes, changes in the protein to lipid ratio, and altered transport of methyl beta-D-thiogalactopyranoside . The differences in membrane composition and transport behavior seen in oleate supplemented E . coli K1060B5 relative to ML 308-225-UFA-8 are enhanced by the incorporation of 8, 8-difluoromyristate . The phase transition behavior becomes distinctly different and some differences in lipid organization persist above the transition temperature . Concomitantly, the rate and extent of concentration of methyl beta-D-thiogalactopyranoside are reduced two-fold more in E . coli K1060B5 compared to ML 308-225-UFA-8 . Such behavior suggests that these fluorinated fatty acid supplemented strains of E . coli are useful to study subtle differences in protein-lipid interactions and their effects on the function of membrane-bound enzymes.

Arch Ophthalmol, 1981 Feb, 99(2), 305 - 8
Epidermal growth factor receptors on corneal endothelium; Fabricant RN et al.; If a growth factor could bind to and stimulate human endothelial healing, corneal disease could be minimized . To this end, primary cultures of feline and human corneal endothelium were tested in receptor binding assays for radiolabeled epidermal growth factor (EGF) . Both of these cells bound ten times as much 125I-EGF as did the negative control cell lines . The time course of association of 125I-EGF to cat corneal endothelium was found to be complete after approximately 120 minutes at 22 degrees C . The 125I-EGF was shown not to dissociate greatly when fresh binding buffer was added to endothelial cultures that had bound the radiolabeled peptide . The pH optimum for binding was determined to be approximately 6.4 . The receptor number per cell and the affinity constant for binding were determined to be 40,000 receptors per cell and 1.1 x 10(9) L/mole, respectively, using a Scatchard plot . Parallel cultures of human fetal corneal endothelium grew in vitro only when the growth medium was supplemented with low concentrations of EGF . These studies provide evidence that EGF is specifically bound to the corneal endothelium.

J Virol, 1981 Feb, 37(2), 721 - 9
Independent expression of avian sarcoma virus in doubly infected chicken embryo fibroblasts; Humphries EH et al.; Infection of a chicken cell with avian sarcoma virus requires division of the infected cell before synthesis of infectious progeny is initiated . This requirement for a cell division for the complete expression of avian sarcoma virus has been examined further with chicken embryo fibroblasts infected with two distinct viruses . Chicken cells infected with and producing a mutant of Rous sarcoma virus temperature sensitive for transformation (tsLA24PR-A) were arrested in G0 by depletion of serum factors from growth medium . These stationary cells continued to produce infectious progeny in the absence of further cell division . Superinfection of the stationary cells with the wild-type Prague strain of Rous sarcoma virus (PR-RSV-C) produced a stable double infection in these cells . Progeny of the superinfecting PR-RSV-C, however, were not detected until these cells underwent division after stimulation with fresh serum-containing medium . The addition of colchicine to these serum-stimulated cells, although not affecting production of the tsLA24PR-A, inhibited the appearance of progeny of the superinfecting PR-RSV-C . These experiments indicate that each avian sarcoma virus infection of a chicken embryo fibroblast requires division of the infected cell for production of that virus regardless of whether or not the cell is already producing a similar virus . The results suggest, therefore, that the requirement for a cell division represents a requirement for an event that controls virus expression in a "cis-acting" fashion specific for the provirus.

J Biol Chem, 1981 Jan 25, 256(2), 1015 - 22
Characterization of a hyaluronic acid-dermatan sulfate proteoglycan complex from dedifferentiated human chondrocyte cultures; Oegema TR Jr et al.; Cells grown from human articular cartilage by an explant method were shown to be dedifferentiated chondrocytes by the failure to respond to treatment with 5-bromodeoxyuridine, vitamin A, or the addition of hyaluronic acid . The major glycosaminoglycan produced by these cells was high molecular weight hyaluronic acid . The majority of the 35S-sulfated glycosaminoglycan was copolymeric chondroitin sulfate-dermatan sulfate (Mr = 2 to 2.4 X 10(4) . The dermatan sulfate was present as the 4-sulfated isomer . Labeled proteoglycans were isolated from the growth media by centrifugation in either associative or dissociative cesium chloride density gradients in the presence of protease inhibitors or by precipitation with cetylpyridinium chloride and column chromatography on Sepharose CL-2B . The proteoglycans secreted into the medium by these dedifferentiated chondrocytes were large molecular weight proteoglycans (Kav = 0.23 on Sepharose CL-2B) that were able to form specific complexes with hyaluronic acid . The material isolated by the cetylpyridinium chloride precipitation was an aggregated proteoglycan (78% aggregate) that was not displaced by exogenous proteoglycan which after reduction and alkylation gave a large proteoglycan monomer (Kav = 0.23 on Sepharose CL-2B) . These findings are in direct contrast to the report of small molecular proteoglycans isolated from cultures of dedifferentiated chrondrocytes obtained from other species.

Biochem J, 1981 Jan 15, 194(1), 299 - 307
Removal of glycosaminoglycans from cultures of human skin fibroblasts; Gill PJ et al.; Early-passage human skin fibroblasts were grown as monolayers for 2-3 days in minimum essential medium containing {35S}sulphate, {3H}glucosamine, {3H}fucose, {3H}proline or {3H}leucine to label proteoglycans, glycoproteins or collagen and other proteins . A crude enzyme preparation obtained from a supernatant from sonicated freeze-dried Flavobacter heparinum was added to the cell monolayers . This treatment removed most of the 35S-labelled glycosaminoglycans, with no appreciable removal of the 3H-labelled proteins or 3H-labelled glycoproteins . The cells remained attached and viable as a monolayer . The formation of 35S-labelled glycosaminoglycans was examined after pretreating cultures with crude F . heparinum enzyme, followed by addition of fresh growth medium containing {35S}sulphate . The F . heparinum enzyme did not significantly alter the amount or type of 35S-labelled glycosaminoglycans produced . Thus F . heparinum enzyme can be used to provide cultured-cell monolayers depleted of surface glycosaminoglycans . These cells remain attached, viable and subsequently synthesize normal amounts and type of glycosaminoglycans.

Biochim Biophys Acta, 1981 Jan 15, 657(1), 222 - 31
Angiotensin converting enzyme in cultured endothelial cells and growth medium . Relationships to enzyme from kidney and plasma; Ching SF et al.; We have previously reported that cultured cells from swine aorta possess angiotensin converting enzyme (peptidyldipeptide hydrolase, EC 3.4.15.1) and release it into serum-free culture medium . The present work compares enzyme from these two sources, and from swine kidney and serum, with respect to antibody and lectin binding . Purified enzyme from swine kidney, and the activity in swine serum, cultured endothelial cells and culture medium bind similarly to rabbit antibodies prepared against the kidney converting enzyme . Enzyme from each of these sources was allowed to bind to an immobilized lectin (Ricinus communis), which binds to terminal galactose residues of glycoproteins . Increasing concentrations of galactose were used to remove enzyme from the lectin column and the distribution of enzyme activity in the galactose eluates was determined . The elution pattern was similar for kidney and endothelial cell enzyme, and different from the pattern found for both serum and medium enzymes . Neuraminidase treatment of either serum or medium enzyme altered the distribution of activity eluted to that found for endothelial cell or kidney enzymes . The effects of neuraminidase suggest that the difference in lectin binding between cell and medium enzyme reflects differences in the number of terminal sialic acid residues that cover galactose residues.

J Biol Chem, 1981 Jan 10, 256(1), 87 - 91
Effect of cholesterol on macromolecular synthesis and fatty acid uptake by Mycoplasma capricolum; Dahl JS et al.; The rates of protein and lipid synthesis of Mycoplasma capricolum were essentially synchronous during growth and depended on the sterol supplement in the media increasing in the order cholesterol (0.5 microgram/ml) < lanosterol (10 microgram/ml) < lanosterol (10 microgram/ml) + cholesterol (0.5 microgram/ml) < cholesterol (10 microgram/ml) . The effect of lanosterol plus low cholesterol on macromolecular synthesis was synergistic . Whereas protein and lipid synthesis were brought virtually to a halt by cholesterol starvation, DNA synthesis continued for about 8 h . Increasing the palmitate and elaidate concentrations 4-fold in the lanosterol-supplemented media raised the growth rate even in the absence of the small amount of cholesterol (0.5 microgram/ml) needed otherwise for the synergistic effect on growth . Studies of the kinetics of fatty acid uptake by resting cells showed that the apparent Km (17 microM) of oleate uptake in lanosterol-grown cells was specifically lowered to 3 microM, a value equal to that seen in cholesterol-grown cells, by the inclusion of a synergistic amount of cholesterol in the growth media . By contrast, the apparent Km for palmitate uptake was the same (2 microM) for all three cell types . The results are consistent with the membrane cholesterol serving in a dual role, one as a bulk component and another more specific function involving the regulation of unsaturated fatty acid uptake and thereby phospholipid biosynthesis.

J Bacteriol, 1981 Jan, 145(1), 122 - 8
Cadaverine is covalently linked to peptidoglycan in Selenomonas ruminantium; Kamio Y et al.; Cadaverine was found to exist as a component of cell wall peptidoglycan of Selenomonas ruminantium, a strictly anaerobic bacterium . {14C}cadaverine added to the growth medium was incorporated into the cells, and about 70% of the total radioactivity incorporated was found in the peptidoglycan fraction . When the {14C}cadaverine-labeled peptidoglycan preparation was acid hydrolyzed, all of the 14C counts were recovered as cadaverine . The {14C}cadaverine-labeled peptidoglycan preparation was digested with lysozyme into three small fragments which were radioactive and were positive in ninhydrin reaction . One major spot, a compound of the fragments, was composed of alanine, glutamic acid, diaminopimelic acid, cadaverine, muramic acid, and glucosamine . One of the two amino groups of cadaverine was covalently linked to the peptidoglycan, and the other was free . The chemical composition of the peptidoglycan preparation of this strain was determined to be as follows: L-alanine-D-alanine-D-glutamic acid-meso-diaminopimelic acid-cadaverine-muramic acid-glucosamine (1.0:1.0:1.0:1.0:1.1:0.9:1.0).

Environ Health Perspect, 1981 Jan, 37, 117 - 23
Bioassaying for ozone with pollen systems; Feder WA; Sensitivity to ozone of pollen germinating in vitro is closely correlated with ozone sensitivity of the pollen parent . Ozone-sensitive and tolerant pollen populations have been identified in tobacco, petunia, and tomato cultivars . The rate of tube elongation can be reversibly slowed or stopped by exposure to low concentrations of ozone . Tube growth rates in the presence of a range of ozone dosages, of pollen populations exhibiting differing ozone sensitivity can be measured and different growth rates can be correlated with ozone dosages . The performance of selected pollen populations can then be used to bioassay ozone in ambient air by introducing the air sample into a growth chamber where ozone-sensitive pollen in growing . Petunia and tobacco pollen are especially useful because they store well at ordinary freezer temperatures and do not require special preparation prior to storage . Modified Brewbacker's growth medium is suitable for growth of both these pollen types . Four useful cultivars are Bel W-3, ozone-sensitive and Bel B, ozone-tolerant tobacco, and White Bountiful, ozone-sensitive and Blue Lagoon, ozone-tolerant petunia . Observations can be made directly by using a TV scanner, or by time lapse or interval photography . Year-round pollen production can be achieved in the greenhouse . Harvested pollen can be tested, packaged, and transported to user facilities without loss of vigor . Pollen populations are inexpensive to produce, respond reliably, and are simple to use as a bioassay for air quality.

Cancer Detect Prev, 1981, 4(1-4), 239 - 47
Chemical transformation of cultured skin fibroblasts from humans genetically predisposed to cancer; Rhim JS et al.; Adenomatosis of the colon and rectum (ACR) is an inherited form of cancer . Assuming that phenotypic expressions that appear in cell strains reflect its biological abnormalities, the study of cultured skin fibroblasts derived from individuals with an inherited form of cancer such as ACR provides a unique study for analysis of the oncogenic process . Growth disorders and increased susceptibility to tumor promoters and to transformation by an oncogenic RNA tumor virus have been demonstrated in these skin fibroblasts . We found that human skin fibroblasts (PF) derived from ACR individuals were sensitive to a chemical carcinogen . Cells treated only with various levels of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) underwent morphological alteration . The morphologically altered cells formed large cell aggregates when suspended in liquid growth medium above an agar base and grew to high saturation densities but did not form colonies in soft agar . Transformed cells were resistant to rechallenge of MNNG (1 microgram/ml) and showed prolonged life span compared to those untreated cells . However, no tumors were produced when cells were inoculated subcutaneously into nude mice . Data suggest that neoplastic transformation of these skin cells by chemical carcinogens is a multi-phase process.

Breast Cancer Res Treat, 1981, 1(2), 141 - 8
Potential and problems with growth of breast cancer in a human tumor cloning system; Von Hoff DD et al.; A human tumor cloning system has been utilized to culture 431 patients' breast cancer specimens . Overall, 288 or 67% of the specimens formed colonies in soft agar . Of the primary lesions 188/260 (72%) formed colonies and 100/171 (58%) of the metastatic lesions formed colonies . The median number of colonies per 500,000 nucleated cells plated was 47 for the primary lesions and 30 for the metastatic lesions . Growth from a variety of metastatic sites ranged from 22% for intradermal lesions to 77% for solid visceral metastases . Methods to increase the number of colonies from a specimen are reported including increasing the number of nucleated cells plated and making a variety of changes in the growth media . None of these methods has had a major impact on colony growth . The antitumor activity of standard anticancer agents such as adriamycin and medroxyprogesterone in the assay is presented . In addition, in vitro results with two new anthracene derivatives demonstrate good antitumor activity for the derivatives . The cloning assay represents a new model for both the basic and clinical studies of human breast cancer.

Bioelectromagnetics, 1981, 2(4), 329 - 40
Inhibition and recovery of growth processes in roots of Pisum sativum L . exposed to 60-Hz electric fields; Robertson D et al.; Roots of Pisum sativum L . were chronically exposed in aqueous inorganic nutrient medium to 60-Hz electric fields between 140 and 490 V/m (growth medium conductivity approximately 0.08 S/m) . The growth rate, meristematic mitotic index, and growth rate recovery of the roots were determined . At 140 V/m there was no perturbation in growth rate or mitotic index . At 430 V/m the growth rate and the mitotic index were reduced . The mitotic index had a maximum depression (approximately 55% of control), which occurred at 4 h . The depression in growth rate was immediate and constant over time . When roots were exposed to an electric field at 430 V/m for 2 days, the growth rate was depressed by about 40% . When the field was terminated, the growth rate steadily increased and was almost normal after 5 days . At 490 V/m root growth rate was almost completely arrested . According to these results, there is a narrow range of induced membrane potentials that span the range from slightly altered to almost completely arrested growth rates.

J Anat, 1981 Jan, 132(Pt 1), 1 - 18
Myogenesis in adult mammalian skeletal muscle in vitro; Nag AC et al.; An injury to adult mammalian skeletal muscle is followed by regeneration, which involves a process believed to be similar to the differentiation of muscle fibres in the embryo . The origin of these differentiating myogenic cells is conjectural . The aim of the present study was to examine the source of myogenic cells and the process of myogenesis in adult skeletal muscle . Mononucleated cells were released from adult rat leg muscle mince after incubation with 0.1% pronase for 50-60 minutes at 37 degrees C . The ultrastructural studies revealed that the freshly dissociated mononucleated cells consisted of at least two populations of cells: myogenic satellite cells and non-myogenic fibroblastic cells . These cells were plated in growth media at various densities in cell culture dishes and incubated for 3 weeks in a balanced air atmosphere at 37 degrees C . The culture was routinely examined with a phase contrasted microscope for evidence of myogenic activities of the plated cells . At selected time intervals, the cell cultures were processed for autoradiography and scanning and transmission electron microscopy (SEM and TEM) . Attachment of cells to the dish began soon after plating, with flattening of some non-muscle cells . The round- to spindle-shaped cells, indicative of myoblasts, began to appear within 24 hours . DNA synthesis and cell proliferation were observed in myogenic and non-myogenic cells within 24 hours of culture . SEM revealed that at 72 hours some myoblasts aligned and fused with one another, forming myotubes . Quantitation of autoradiographs indicated that the maximum number of labelled myotubes were present in the 3 days old culture, and thereafter, the labelled myotubes decreased in number and were absent in the 7 days old culture . Within 5-7 days the myotubes became larger and showed contractility . TEM of 6 to 21 day culture revealed that the myotubes contained well differentiated myofibrils, T-tubules and sarcoplasmic reticulum . It was evident from our studies that the mononucleated cells, having satellite cell morphology, were capable of differentiating into fully formed muscle fibres . This study lends support to the satellite cell hypothesis for regeneration of the skeletal muscle.

Radiat Environ Biophys, 1981, 19(3), 197 - 203
Absence of AET protection against fast neutrons: cellular effects; Ferle-Vidovic A et al.; A series of experiments has been undertaken in order to test the biological properties of neutrons produced in the cyclotron of the Institute "Ruder Boskovic" (IRB) in Zagreb . Protective effect of AET (2-amino ethylisothiuronium bromide hydrobromide) on survival of L cells irradiated by fast neutrons generated in the IRB cyclotron were studied by employing the single cell clonal growth method . For comparison the protective effect of AET after gamma irradiation has also been studied . The most important findings that have emerged from these experiments can be summarized as follows: (1) Protective effect of AET was present after gamma irradiation only . (2) The degree of protection was dependent on AET concentration in the growth medium . (3) No protective effect was found after neutron irradiation . These findings are in agreement with the generally less efficient protection of this compounds after high-LET irradiation.

Vopr Onkol, 1981, 27(3), 68 - 71
{Transformation and the kinetic cell growth patterns in vitro after exposure to adenovirus SV20 DNA}; Parkhomenko II et al.; Malignant transformation of the normal rat embryo cells has been obtained by monkey adenovirus SV-20 and its DNA . The appearance of the transformed colonies with a multi-layer cell growth was observed at the 28th day . The influence of the growth medium pH on the process of cell transformation has been studied . The kinetic regularities of normal and transformed cells growth were investigated by a cloning assay.

Z Allg Mikrobiol, 1981, 21(8), 591 - 600
{Use of automated image analysis for screening the efficiency of turimycin-producing Streptomyces strains}; Muhlig P et al.; The growth of the surface colonies on solid media is studied with the image analyzing system Quantimet 720 M controlled by a PDP 11 computer . For the bacterial strain Streptomyces hygroscopicus JA 6599 a new differential equation describing the kinetic behaviour of the area of the colony is suggested . It is shown that this time dependence of area describes more realistically the experiment than the function resulting from Pirt's model . Two new constants arise from the growing curve, alpha describes the profile of density of the colony and k0 is correlated to the growing constant of the colony . Both parameters can be correlated to the amount of antibiotics produced and, therefore, it is possible to use this information for the selection procedure of the colonies . A uniform criterion for selection was found for each colony of two mutants of the turimycin-producing strain Streptomyces hygroscopicus JA 6599, which reads: (Formula: see text) . The quality of the selection criterion of surface colonies with higher productivity is dependent on the mean value of the growing parameters of all colonies under investigation . Limitations of the selection method arise from the need for great uniformity of environmental conditions, growth medium and a homogeneous spore suspension, which have to consist of a large amount of single spores in order to assure a reproducible growing curve of the colonies . For the last condition a special technique is pointed out.

Microbios, 1981, 31(124), 71 - 82
On the uptake of nystatin by Saccharomyces cerevisiae 3 . Electrochemistry of the yeast cell surface; Beezer AE et al.; The electrophoretic behaviour of fresh and liquid nitrogen stored inocula of Saccharomyces cerevisiae has been studied as a function of pH, ionic strength, buffer composition, growth temperature of the inoculum, number of washings of the inoculum prior to electrophoretic investigation, growth medium used in the preparation of the inoculum, interaction of the inoculum with nystatin, and interaction of the inoculum with calcium ions . The results provide a basis for a discussion of the role of the yeast cell surface in the mode of action of nystatin (a polyene antibiotic), on interaction with sensitive Saccharomyces cerevisiae cells . They indicate the profound importance of the cell surface in the interaction with the antibiotic . Moreover, the results of the temperature growth/electrophoresis experiments support the view that a sharp change in membrane fluidity occurs in the yeast membrane . The results are also discussed in the light of those reported in previous papers on the interaction of yeast cells with nystatin.

Microbios, 1981, 30(120), 87 - 96
The surface properties of cells of Mycobacterium BCG; Hardham LE et al.; The surface properties of cells of Mycobacterium BCG, M . phlei, M . Smegmatis and M . microti are identical, irrespective of the growth medium, the age of the cells, and the colonial morphology, and after various vigorous chemical treatments . The negative surface charge for cells of all these species arises from the phosphate groups of phosphodiester linkages between the peptidoglycan and the arabinogalactan of the basic cell wall structure which is common to all species of Mycobacteria.

Carcinogenesis, 1981, 2(9), 863 - 72
Much of spontaneous mutagenesis in Escherichia coli is due to error-prone DNA repair: implications for spontaneous carcinogenesis; Sargentini NJ et al.; The role of DNA repair genes (uvrA, uvrB, uvrD, recA, recB, lexA, and umuC) in spontaneous mutation rate per bacterium per cell division (micro) was determined for the reversion of UAA (his-4 and trpE65), UAG (lacZ53), and frameshift (trpE9777) mutations, and for the occurrence of forward mutations to valine resistance . Rich growth medium enhanced micro in a wildtype strain but not in a uvrB5 strain . In minimal growth medium, the uvrA and uvrB strains had the largest micro (1.9-6.2-fold greater than that for isogenic wild-type strains, depending on the mutation assay) . The uvrB strains carrying lexA, recA, umuC, or both the uvrD and rec B mutations (in combination), i.e., mutations that inhibit error-prone DNA repair, had the lowest micro values (approximately 10-fold less than the uvrB strain) . Teh recA and lexA mutations also reduced micro (by approximately 2-fold) in uvr+ strains . The genetic control of the error prone repair-dependent sector of spontaneous mutagenesis was shown to be qualitatively similar to the genetic control for u.v . radiation mutagenesis . The umuC mutation, which drastically reduced spontaneous mutagensis, had no effect on genetic recombination . It is proposed that the low level of spontaneous mutagenesis observed in the recA, lexA, umuC, and the uvrD recB strains is due to errors made during DNA replication, while the enhanced level of spontaneous mutagenesis observed in the wild type, and especially in the uvrA and uvrB strains, is due to excisable lesions that are produced in the DNA by normal metabolic reactions, and that such unexcised lesions induce mutations via error-prone DNA repair . These results are discussed in terms of their relevance to spontaneous carcinogenesis.

Mol Gen Genet, 1981, 182(2), 288 - 92
Regulation of ferric iron transport in Escherichia coli K12: isolation of a constitutive mutant; Hantke K; The lac genes were inserted with phage Mu(Ap, lac) into the fhuA, fepA, cir and tonB genes which specify components of iron uptake systems . The expression of lac in all these operon fusions was controlled by the availability of iron to the cells, thereby facilitating a quick and simple measurement of the expression of the genes listed above . In an iron rich medium under anaerobic conditions all systems were strongly repressed . fhuA was depressed at higher iron concentration than was fepA or cir, and tonB was repressed only under anaerobic conditions and could be induced by iron limitation . Mutants constitutive for the expression of beta-galactosidase were selected in a fhuA-lac fusion strain . The outer membrane proteins Cir, FhuA, FecA, 76K and 83K were made constitutively in such mutant strains . Therefore, they were termed fur mutants . In these fur mutant strains, the synthesis of a 19K protein was reduced . Furthermore, it was found that transport of ferric enterochelin and ferrichrome was also constitutive in the fur mutant cells, and that ferric citrate uptake could be induced by only 10 microM citrate in the growth medium in contrast to wild-type cells in which at least 100 microM citrate was necessary . The fepA gene was concluded to be under an additional control, because it was not fully derepressed by the fur mutation.

Z Allg Mikrobiol, 1981, 21(3), 255 - 9
Cultural and nutritional studies of zoopathogenic fungi associated with livestock feeds in Nigeria; Ogundero VW; Mycelial growth and nutritional physiology of zoopathogenic Aspergillus fumigatus FRES., Thermoascus aurantiacus MIEHE, and Thermomyces lanuginosus TSIK., obtained from livestock feeds and poultry droppings in Nigeria were studied . An optimal pH-temperature range of 5.5-6.5 and 37 degrees - 45 degrees C was recorded for the growth of these fungi . Various sources of carbon supplied in the growth medium, except sorbose and rhamnose, were utilized for growth . Only Aspergillus fumigatus caused considerable weight losses of the filter papers supplied and hydrolyzed the carboxy-methyl cellulose (CMC) in the medium . The culture filtrates of this fungus contained CM-cellulases which also hydrolyzed CMC to reducing sugar at 45 degrees C . D-glucose and L-asparagine concentrations in the range of 15-20 g/liter and 2.5-5 g/liter, respectively, were best for growth of the test fungi . Various forms of organic and inorganic nitrogen provided were also utilized . The inorganic sources of nitrogen could readily substitute the organic forms at the optimal growth conditions.

Mol Gen Genet, 1981, 182(1), 60 - 4
Nucleic acid metabolism in yeast II . Metabolism of thymidylate during thymidylate excess death; Toper R et al.; A discrete class of strains of Saccharomyces cerevisiae, able to utilize, highly efficiently, exogenous deoxythymidine-5'-monophosphate (dTMP),was found to be sensitive to concentrations greater than 10 Micro M dTMP in an otherwise complete growth medium . Excess dTMP is cytostatic and cytotoxic: 90% of exponentially growing cells lose colony forming ability within 1 h of exposure to excess dTMP is a growth medium . Uptake of dTMP, adenine, histidine, and leucine does occur during this thymidylate excess death (TED) . dTMP is anabolized to higher phosphorylated synthesis is blocked under TED-conditions but not RNA and protein biosynthesis.

Cancer Res, 1981 Jan, 41(1), 237 - 43
Differential expression of the globin genes in human leukemia K562(S) cells induced to differentiate by hemin or butyric acid; Cioe L et al.; Human leukemia K562(S) cells were induced to differentiate by 50 microM hemin or 1.4 mM butyric acid, and the types of hemoglobins synthesized were compared . In both cases, embryonal hemoglobins {Portland, Gower 1, Hb X, and fetal hemoglobin (Hb-F)} were detected . Butyric acid-treated K562(S) cells contained mostly Hb Gower 1 (zeta 2 epsilon 2) and a hemoglobin with the electrophoretic characteristics of Portland (gamma 2 zeta 2) . For hemin-treated K562(S), the most abundant hemoglobin synthesized by Hb X (epsilon 2 gamma 2), and the second most abundant was Bart's (gamma 4) . Traces of Gower 1 were observed in nontreated K562(S) cells . The kinetics of hemoglobin induction as a result of the two treatments differed; increased hemoglobin synthesis was detected after only 24 hr of hemin treatment, whereas 4 days were required in butyric acid-treated cells . Both hemin and butyric acid were able to induce their respective patterns of hemoglobin synthesis independent of the presence of serum in the K562(S) growth medium . Analysis of the globin chains in induced K562(S) cells induced to differentiate indicated that, with both inducers, adult alpha- but not beta-globin chains were present . Karyotype analysis of K562(S) cells revealed a nearly triploid chromosome complement with a modal number of 68 chromosomes . Three copies of chromosome 11 and four copies of chromosome 16 (coding for the beta-like and alpha-like globin genes, respectively) were present . A large marked chromosome, involving chromosome 7, and a Philadelphia chromosome were also seen . These data characterize the K562(S) subline and also indicate that hemin and butyric acid differ in their effects on the expression of embryonal globin genes.

J Cell Biol, 1981 Jan, 88(1), 42 - 50
Membrane-bound ribosomes of myeloma cells . VI . Initiation of immunoglobulin mRNA translation occurs on free ribosomes; Mechler B; Immunoglobulin heavy (Ig H) and light (Ig L) chain mRNA molecules have been released from the endoplasmic reticulum (ER) membranes as free (F) mRNP particles when MOPC 21 (P3K) mouse myeloma cells are exposed to a hypertonic initiation block (HIB) . The subsequent fate of these mRNA sequences has been examined when the cells are returned to normal growth medium . Upon return to isotonicity, all previously translated mRNA molecules reassociate with ribosomes and form functional polysomes . Ig H mRNA is found incorporated first into F polysomes and then into membrane-bound (MB) polysomes . Kinetic studies indicate that the time of passage of Ig H mRNA in F polysomes is approximately 30 s, during which a nascent polypeptide chain of approximately 80 amino acids would have been completed . When the rate of polypeptide elongation is depressed with emetine during the recovery from HIB, both Ig H and L mRNA molecules accumulate in small F polysomes . These results indicate that the formation of Ig-synthesizing polysomes proceeds in the sequence: mRNA leads to F polysomes leads to MB polysomes . With the additional observation that during HIB recovery puromycin completely prevents the reassociation of Ig mRNA with the ER, these findings support a model of MB polysome formation in which the specificity of membrane attachment is determined by the nature of the N-terminal amino acid sequence of the nascent polypeptide chain.

Arch Geschwulstforsch, 1981, 51(7), 615 - 22
{Phorbol ester-induced expression of primate retroviruses (author's transl)}; Wunderlich V et al.; Addition of the tumor promoter 12-0-Tetradecanoyl-phorbol-13-acetate (TPA) to the growth medium of certain human cell cultures persistently infected with simian retroviruses of type C (BaEV, SSV) or type D (MPMV) or a human cell line-derived type D isolate (PMFV), respectively, resulted in a considerable but transient stimulation of virus production . Enhanced virus expression was paralleled by striking morphological alterations of the cells . Among four infected cell types tested so far, ony one (A 204) failed to respond to TPA with significant virus stimulation or altered morphology.

Proc Natl Acad Sci U S A, 1981 Jan, 78(1), 313 - 7
Neoplastic transformation of chimpanzee cells induced by adenovirus type 12--simian virus 40 hybrid virus; Rhim JS et al.; The adenovirus 12--simian virus 40 hybrid virus produced neoplastic transformation of chimpanzee skin fibroblasts in vitro . The transformed fibroblasts showed morphological alteration and became permanent lines . The transformed cells contained both adenovirus 12 and simian virus 40 large tumor antigens and were virus producers . However at passage 9, one line (WES) was found to be a nonproducer, producing neither infectious virus nor virus-specific antigen detectable by the complement fixation test . Virus particles were not detected nor could infectious hybrid virus be rescued from this line by cocultivation with Vero cells . The transformed cells formed large cell aggregates and grew in liquid growth medium above an agar base, formed colonies in soft agar, and grew to high saturation densities; the normal chimpanzee skin fibroblasts did not . One transformed WES line produced tumors when transplanted subcutaneously into newborn nude mice, thus providing an important tool for studying tumor immunity in the chimpanzee.

Dev Biol Stand, 1981, 47, 25 - 33
Serially subcultivated cells as substrates for poliovirus production for vaccine; von Seefried A et al.; Four human diploid cell cultures, WI-38, MRC-5, HEL 299, and IMR-90, and the aneuploid simian cell culture LLC-MK2 were considered in this study as alternatives to primary monkey kidney cells in the production of Inactivated Poliomyelitis Vaccine (IPV) . The composition of the basal media used for cell growth and viral replication was found to be critical . Medium CMRL-1969 required a 50% increase in the concentrations of arginine, cystine, glutamine, isoleucine, leucine, methionine, serine, and threonine to produce optimum cell and viral antigen yields, particularly in the Multi-Surface-Cell-Propagator (MSCP) and in the microcarrier suspension culture systems . Also, Medium 199, with Earle's Basic Salt Solution (BSS) for the human diploid cultures, and Hanks' BSS for the simian aneuploid culture, increased the D-antigen yields when compared with other virus growth media deficient in nucleic acid precursors . Approximately 3--4 cell population doublings were obtained in microcarrier culture systems . The average D-antigen yields for poliovirus Types 1,2 and 3 were about 110, 25 and 35, respectively . The physiological requirements for the four human diploid cell cultures were similar indicating easy interchange . The major advantage of the aneuploid LLC-MK2 cells is the reduced serum requirement . The microcarrier culture system is suitable for these cell substrates in the production of IPV provided the microcarriers can be kept in "single-bead" suspension at low rotational speeds of, initially, 10 rpm, with slight increases as the cell density increases.

Steroids, 1981 Jan, 37(1), 51 - 61
Macromolecular synthesis is required for stimulation of estrogen synthetase activity by dibutyryl cyclic AMP plus theophylline in choriocarcinoma cell culture; Bellino FL et al.; Estrogen secretion and estrogen synthetase (aromatase) activity are stimulated in human trophoblast cells (JAr line) after addition of 1 mM dibutyryl cyclic AMP plus 1 mM theophylline (dbT) to the growth medium . The data given here show that (a) the aromatase specific activity in homogenized cells increases linearly over a 96 hr incubation period after addition of dbT; (b) addition of inhibitors of macromolecular synthesis, cycloheximide or actinomycin D, to the culture medium at the time of addition of dbT abolishes the stimulation of aromatase activity; (c) mixing dbT-grown cells, containing increased aromatase activity, with control cells does not result in an aromatase specific activity higher than the expected average, suggesting that dbT-grown cells do not contain a factor present in excess which serves to stimulate aromatase in control cells; and (d) NADPH, included in vitro in the aromatase assay or incubated with the cells for 48 hr as well as being present in the aromatase assay, has no stimulatory effect on aromatase specific activity in homogenized cells.

J Cell Biol, 1981 Jan, 88(1), 57 - 66
Differentiation of a teratocarcinoma line: preferential development of cholinergic neurons; Pfeiffer SE et al.; A line of embryonal carcinoma cells, PCC7-S, established in vitro from a spontaneous testicular teratocarcinoma, has been studied . Upon removing the cells from a low density monolayer culture system and permitting the cells to form aggregates in suspension, we observed a change of several physical and biochemical parameters: (a) reduction in average cell volume, (b) blockage and accumulation of cells in G1, (c) rise in secreted protease activity, (d) rise in acetylcholinesterase and choline acetyltransferase activities, and (e) disappearance of embryonic antigen F9 . Although PCC7 aggregates did not undergo substantial morphological changes while suspended, when aggregates 4 or more days old were allowed to attach to plastic tissue culture dishes, substantial neurite outgrowth occurred over the next 1-3 d . This process was markedly enhanced by the addition to the growth medium of carboxymethylcellulose and inhibitors of DNA synthesis . Transmission electron microscopy disclosed a neurite ultrastructure consistent with that of neuronal processes . A veratridine-stimulated, tetrodotoxin-blocked sodium influx of 100 nmol/min per mg protein was also observed in these differentiated surface cultures . This cell line is discussed in terms of its utility for the study of early events leading to a commitment to cellular differentiation, as well as for the investigation of terminal differentiation to cholinergic neurons.

J Bacteriol, 1981 Jan, 145(1), 410 - 6
In vivo role of the relA+ gene in regulation of the lac operon; Primakoff P; Under conditions of amino acid limitation, beta-galactosidase was produced at a 70-fold higher rate in a relA+ strain than in an isogenic relA strain of Escherichia coli K-12 . Under identical conditions with the relA+ and relA strains carrying various lac promoter mutations, rates of beta-galactosidase synthesis in relA+ (relative to relA) ranged from 26-fold higher (promoter mutant Pr 13) to only 5-fold higher (promoter mutant PrL8uv5) . This promoter specificity was independent of strain background and the means of eliciting amino acid limitation . Addition of cyclic AMP to the growth medium altered the relA+/relA difference for beta-galactosidase synthesis from the wild-type lac promoter . The experiments suggest that the relA+/relA difference in lac expression arises primarily at the point of transcription initiation . The results are discussed in relation to recent in vitro data showing a promoter-specific guanosine 5'-diphosphate 3'-diphosphate stimulation of lac transcription (P . Primakoff and S . W . Artz, Proc . Natl . Acad . Sci . U.S.A . 76:1726-1730).

Nature, 1980 Dec 25, 288(5792), 715 - 7
Growth-modulating plasma tripeptide may function by facilitating copper uptake into cells; Pickart L et al.; The plasma tripeptide glycyl-L-lysine (GHL), when added at nanomolar concentrations to a wide group of cultured systems, produces a disparate set of responses ranging from the stimulation of growth and differentiation to outright toxicity . Such diverse actions imply that this tripeptide mediates some basic biochemical function common to many types of cells and organisms . During the isolation of GHL we found the compound to co-isolate through a number of steps with approximately equimolar copper and about 1/5 molar iron . Maximal effects on hepatoma cells (HTC4) were seen when the peptide was added with copper and iron to the growth medium . Structure-function studies revealed that several tripeptides with a histidyl-lysyl linkage were nearly as active as GHL . The association of GHL with copper and a homology similarity between the tripeptide and the copper transport sites on albumin and alpha-fetoprotein, where the cupric atom is bound to a histidyl residue adjacent to a basic residue, suggested that GHL may act as a copper transport factor . We report here that the tripeptide readily forms complexes with copper(II) and enhances the uptake of the metal into cultured hepatoma cells.

Mutat Res, 1980 Dec, 73(2), 267 - 77
Yeast mitochondrial DNA characterization after ultraviolet irradiation; Hixon SC et al.; Yeast mitochondrial (mtDNA) 3H-labelled was isolated from exponential phase cells after ultraviolet light irradiation . Both the size and amount of mtDNA were found to be reduced during a 40-h liquid-holding (LH) period in non-growth medium following irradiation as compared to the mtDNA recovered from nonirradiated cells under similar conditions . After the LH period, previously irradiated cells were resuspended in growth medium containing {14C}adenine . Double labelled mtDNA (3H and 14C) was isolated from cell samples removed during new growth . A recovery in the amount and size of mtDNA was observed in irradiated cells during new growth . These biochemical studies agree with the observed loss and recovery of mtDNA genetic markers in UV-irradiated exponential phase yeast after a period of LH and new growth resp.

J Bacteriol, 1980 Dec, 144(3), 1098 - 1112
Messenger ribonucleic acid and protein metabolism during sporulation of Saccharomyces cerevisiae; Kraig E et al.; To investigate differences between growing yeasts and those undergoing sporulation, we compared several parameters of messenger ribonucleic acid (RNA) transcription and translation . The general properties of messenger RNA metabolism were not significantly altered by the starvation conditions accompanying sporulation . The average messenger RNA half-life, calculated from the kinetics of incorporation of {3H}adenine into polyadenylic acid-containing RNA, was 20 min on both cell populations . Furthermore, 1.3 to 1.4% of the total RNA was adenylated in both growing and sporulating cells . However, the proportion of RNA that could be translated in a wheat germ system slowly decreased during sporulation . Within 8 h after the induction of sporulation, isolated RNA stimulated half as much protein synthesis as the equivalent amount of vegetative RNA . There were significant differences in protein synthesis . The percentage of ribosomes in polysomes decreased threefold as the cells entered sporulation . This decrease began within 5 min of the initiation of sporulation, and the steady-state pattern was attained within 120 min . However, the ribosomes were not irreversibly inactivated; they could be reincorporated into polysomes by returning the sporulating cells to growth medium . Though unable to sporulate, strains homozygous for mating type, MAT alpha/MAT alpha, showed a similar decrease in the number of polysomes when placed in sporulation medium . Furthermore, the same shift toward monosomes was observed during stationary phase of growth . We conclude that the redistribution of ribosomes represents a general metabolic response to starvation . Our data indicate that the loss of polysomes is most likely caused by a decrease in the initiation of translation rather than a severe limitation in the amount of messenger RNA . Furthermore, the loss of polysomes is not due to the decreased synthesis of a major class of abundant proteins . Of the 400 vegetative proteins resolved by two-dimensional gel electrophoresis, only 19 were not synthesized by sporulating cells . Approximately 10 to 20% of the cells in a sporulating culture failed to complete ascus formation . We have shown that {35S}methionine is incorporated equivalently into cells committed to sporulation and cells that fail to form asci . Furthermore, the proteins synthesized by these two populations were indistinguishable, on one-dimensional gels . We compared proteins labeled by various protocols, including long-term and pulse-labeling during sporulation and prelabeling during vegetative growth before transfer to sporulation medium . The resulting two-dimensional gel patterns differed significantly . Many spots labeled by the long-term techniques may have arisen by protein processing . We suggest that pulse-labeling produces the most accurate reflection of instantaneous synthesis of proteins.

J Natl Cancer Inst, 1980 Dec, 65(6), 1345 - 50
Glycosaminoglycans synthesized by tumorigenic and nontumorigenic mouse melanoma cells in culture; Heaney-Kieras J et al.; Glycosaminoglycans (GG) synthesized by two tumorigenic cell lines and a nontumorigenic, immunoprotective cell line derived from the B16 mouse melanoma were metabolically labeled with Na235SO4 and {3H}glucosamine . The radioactive GG synthesized at low and high cell densities were prepared from cells and culture media and analyzed by enzymatic and chromatographic methods . The cell-associated and medium GG from both low- and high-density cultures of the nontumorigenic, immunoprotective line contained a significantly higher proportion of heparin and heparan sulfate (85-95% of total) relative to comparable fractions from the tumorigenic lines . In addition to synthesizing less heparin and heparan sulfate and more chondroitin 4-sulfate plus chondroitin 6-sulfate, the tumorigenic lines differed in that the amelanotic line produced significant amounts of dermatan sulfate which remained cell associated . None of the lines produced measurable amounts of hyaluronic acid . In addition, the nontumorigenic immunoprotective line incorporated two to six times more precursor into total GG and released a higher proportion into the growth medium than did the tumorigenic lines.

Proc Natl Acad Sci U S A, 1980 Dec, 77(12), 7306 - 10
Reversible growth arrest in simian virus 40-transformed human fibroblasts; Hoffman RM et al.; A reversible growth arrest of simian virus 40-transformed human fibroblasts has been produced by replacement of methionine in the growth medium by its immediate metabolic precursor, homocysteine, Although these arrested cells exhibit a greatly reduced cloning efficiency when plated in methionine-supplemented medium, they resume rapid proliferation without a lag when subconfluent cells are refed with methionine-supplemented medium . This growth arrest is accompanied by a reduction in the percentage of mitotic cells in the cell population . Furthermore, data obtained using fluorescence-activated cell sorting techniques indicate that the cells are arrested i the S and G2 phases of the cell cycle . This is in contrast to a G1-phase accumulation of cells, which occurs only in methionine-supplemented medium at very high densities and which is similar to the G1 block seen in cultures of normal fibroblasts at high density . The apparent relationship between specific events in the DNA-synthetic and premitotic phase of the cell cycle and methionine dependence in these transformed cultures is discussed.

Arch Microbiol, 1980 Dec, 128(2), 222 - 7
Adenine nucleotide metabolism in Azotobacter vinelandii . Two metabolic pathways of AMP degradation; Yoshino M et al.; AMP-degrading pathways in Azotobacter vinelandii cells were investigated . AMP nucleosidase (EC 3.2.2.4) was rapidly synthesized and reached a maximum at 24 h, while the activity of 5'-nucleotidase (EC 3.1.3.5) specific for AMP, which was negligible during the logarithmic phase of the growth, first appeared in 24 h-cultures, and reached a maximum after complete exhaustion of sucrose from the growth medium (70 h) . Cell-free extracts of A . vinelandii of 48 h-cultures hydrolyzed AMP to ribose 5-phosphate and adenine in the presence of ATP, and adenine was deaminated to hypoxanthine . When ATP was excluded, AMP was dephosphorylated to adenosine, which was further metabolized to inosine, and finally to hypoxanthine . Hypoxanthine thus formed was reutilized for the salvage synthesis of IMP under the conditions where 5-phosphoribosyl 1-pyrophosphate was able to be supplied . These results suggest that the levels of ATP can determine the rate of AMP degradation by the AMP nucleosidase- and 5-'nucleotidase-pathways . The role of ATP in the AMP degradation was discussed in relation to the regulatory properties of AMP nucleosidase, inosine nucleosidase (EC . 3.2.2.2) and adenosine deaminase (EC 3.5.4.4).

J Gen Microbiol, 1980 Dec, 121(Pt . 2), 473 - 6
Regulation of phospho-2-keto-3-deoxy-heptonate aldolase (DAHP synthase) and anthranilate synthase of Pseudomonas aureofaciens; Salcher O et al.; Phospho-2-keto-3-deoxy-heptonate aldolase (DAHP synthase) of Pseudomonas aureofaciens ATCC 15926 was inhibited by L-tyrosine . The inhibition was competitive with erythrose 4-phosphate as the varied substrate but non-competitive with respect to phosphoenolpyruvate . Anthranilate synthase was inhibited by L-tryptophan . The inhibition was competitive with respect to chorismate but non-competitive with L-glutamine or NH4+ as the varied substrate . DAHP synthase and anthranilate synthase were not repressed when aromatic amino acids were included in the growth medium . In bacteria grown in the presence of L-phenylalanine, the anthranilate synthase activity was enhanced about threefold compared with the control . Similar results were obtained with the mutant strain P . aureofaciens ACN, which produces increased amounts of pyrrolnitrin.

Int J Cancer, 1980 Nov 15, 26(5), 565 - 9
Chemical transformation of cultured human skin fibroblasts derived from individuals with hereditary adenomatosis of the colon and rectum; Rhim JS et al.; Chemical transformation of cultured human skin fibroblasts (PF) derived from individuals with hereditary adenomatosis of the colon and rectum is reported . Cells treated only with various levels of N-methyl-N'-nitro N-nitrosoguanidine (MNNG) underwent morphological alteration . The morphologically altered cells formed large aggregates when suspended in liquid growth medium above an agar base and grew to high saturation densities . One altered (MNNG, 1.0 microgram/ml) cell culture formed colonies in soft agar . Transformed cells were resistant to rechallenge of MNNG (l microgram/ml) and showed a more prolonged life-span compared to the untreated cells . Altered cells became heteroploid cells . However, no progressively growing tumors were produced when cells were inoculated subcutaneously into nude mice . The data suggest that chemical carcinogens alone may not induce neoplastic transformation of fibroblasts from humans genetically predisposed to cancer and that neoplastic transformation of these skin cells by chemical carcinogens might require the presence of a tumor promotor and the use of an immuno-privileged site in the nude mouse system.

In Vitro, 1980 Nov, 16(11), 932 - 40
Retention of human skin fibroblast fatty acid modifications during maintenance culture; Spector AA et al.; The fatty acid composition of cultured human skin fibroblasts was modified by adding either oleic or linoleic acid to the growth medium . After the cultures became confluent, they were washed and transferred to different maintenance media in order to determine the stability of the various fatty acyl modifications . Some changes in fatty acid composition occurred under all conditions . When the maintenance medium was supplemented with fatty acid, the cellular neutral lipid and phospholipid fatty acyl composition were altered markedly within 16 to 24 hr . If no supplemental fatty acid was available during the maintenance period, however, the modified fatty acyl compositions were sufficiently retained so that appreciable differences between the cells enriched with oleate and linoleate persisted for at least 48 to 72 hr . This considerable degree of stability occurred when either 10% delipidized fetal bovine serum or 10% fetal bovine serum containing its inherent lipids were present in the maintenance medium . Although the triglyceride content of the fatty acid-modified cells was quite labile, neither the cholesterol nor phospholipid content changed appreciably during culture in any of the maintenance media . Since the fatty acid compositional differences persisted during several days of maintenance under certain conditions, these modified cultures appear to be a useful experimental system for assessing the effect of lipid structure on fairly long-term cellular functions.

Mikrobiologiia, 1980 Nov-Dec, 49(6), 1011 - 13
{Population dynamics and structures of Streptomyces lanatus in 2 soil types}; Mikhailov VV et al.; The dynamics and structure(mycelium--spores) of a population of the soil actinomycete Streptomyces lanatus 21-5 were studied in two types of nonsterile soil using the technique of inoculation on solid growth media in combination with autofluorescence, heat treatment of the culture, and addition of antibiotics, when the population was introduced into soil at a level of density close to the natural one . The dynamics of the population was found to depend on the form (spores or mycelium) in which it was introduced into soil . The population was shown to be stabilized at different levels of density in various types of soil at an identical level of inoculation . The population was found to behave differently in various types of soil . The population was represented in soils preferentially by spores . The structural coefficient K changing from 0 (only mycelium) to 1 (only spores) was proposed in order to estimate quantitatively the structure of the population.

J Protozool, 1980 Nov, 27(4), 474 - 8
Attachment of Entamoeba histolytica to glass in a defined maintenance medium: specific requirement for cysteine and ascorbic acid; Gillin FD et al.; Cysteine and ascorbic acid were previously shown to be required by Entamoeba histolytica trophozoites for attachment to glass, elongation, and ameboid movement as well as for short-term (12-24 h) survival in a balanced salt solution containing bovine serum albumin and a vitamin solution (Maintenance Medium 1) . If the only function of cysteine and ascorbate was to decrease the redox potential, other reducing agents should be effective . However, the requirement for cysteine in the presence of ascorbic acid was highly specific . Equally effective were D- and L-cysteine; however, of many other compounds tested, only thioglycolic acid, ascorbic acid, or L-cystine (in decreasing order) were somewhat active . Under N2 atmosphere, cysteine and ascorbic acid were still required, although their concentrations could be halved . The ability to attach in the maintenance medium was irreversibly lost after only 5 min of cysteine-ascorbic acid deprivation; however, there was no decrease in viability when the amebae were transferred to growth medium within 30 min . Cysteine thiol groups in the medium were oxidized rapidly regardless of the concentration of ascorbic acid or the presence of amebae; however, ascorbic acid prolonged attachment of amebae.

J Biol Chem, 1980 Oct 25, 255(20), 9860 - 9
Insulin-induced loss of the insulin receptor in IM-9 lymphocytes . A biological process mediated through the insulin receptor; Kosmakos FC et al.; Exposure of cultured lymphocytes of the IM-9 line to insulin results in a rapid, time-dependent reduction in the number of insulin receptors to a new steady state concentration . Both the rate of loss and the net loss of receptors were directly related to the ambient insulin concentration . The insulin-induced loss of receptors was mediated by binding of insulin to the receptor itself; insulins, which varied 200-fold in biopotency, produced receptor loss in direct proportion to the ability of each insulin to occupy the receptor . The residual insulin receptors were normal following insulin-mediated receptor loss by a variety of sensitive binding criteria . While insulin binding to its receptors was a necessary condition to induce receptor loss, it was not sufficient . Thus, reduction in the temperature of the preincubation from 37 degrees C to 20 degrees C (which enhanced the total amount of insulin bound to the receptor) abolished the loss of insulin receptors . Likewise, cycloheximide prevented the insulin-induced loss of receptors . Furthermore, turkey erythrocytes, which lack active macromolecular synthesis, had no change in the concentration of insulin receptors when exposed to insulin for similar periods . Interestingly, the turkey erythrocytes, when exposed to insulin or to proinsulin, showed a time- and concentration-dependent increase in the affinity of the insulin receptor over a restricted part of the insulin-binding isotherm, which was reversed over a period of several hours following removal of hormone . The insulin-mediated decrease in receptor number on IM-9 lymphocytes was reversible . Following removal of insulin from the growth medium, about one-half of the receptors were restored within 10 h and the full complement of insulin receptors was restored within 24 h . Cycloheximide prevented restoration of the insulin receptor.

J Cell Physiol, 1980 Oct, 105(1), 51 - 61
Protein synthesis and degradation in growth regulation in rat embryo fibroblasts: role of fast-turnover and slow-turnover protein; Amenta JS et al.; Cultured rat embryo fibroblasts, when stimulated to grow by the addition of fresh medium containing 10% serum, showed an increase in synthesis of slow-turnover proteins while maintaining a uniform degradation rate for these proteins . Slow-turnover proteins with a half-life of 2.4 days accounted for approximately 95% of the cell protein, while the remaining protein could be described in terms of two fast-turnover pools . When we labeled cells to limiting levels over a period of 4 days, the fast-turnover pools became undetectable; with 2-hour labeling periods, however, 25% of the label entered the fast-turnover pools . Fibroblasts, stimulated to grow by fresh growth medium, showed proportionate and coordinate increases in synthesis of both fast-turnover and slow-turnover proteins during the growth period, both returning to baseline levels on reaching the new steady state . No changes could be detected in degradation of either pool during growth . Fibroblasts placed in a serum-free medium showed a decrease in cellular protein and an increased degradation of slow-turnover proteins, while degradation of fast-turnover proteins remained unchanged . We conclude that the slow-turnover protein pool forms the bulk of the cell proteins and turns over at a fairly constant rate . Growth stimulation is effected almost entirely by stimulation of protein synthesis in this pool, while decreasing cellular protein growth is a result of enhanced degradation within this pool.

J Gen Microbiol, 1980 Oct, 120(Pt 2), 545 - 7
Excretion of glutathione by methylglyoxal-resistant Escherichia coli; Murata K et al.; A methylglyoxal-resistant mutant of Escherichia coli B excreted glutathione into the growth medium, especially during growth on medium containing methylglyoxal . In the presence of methylglyoxal, the total amount of glutathione excreted was increased about 50-fold over that of the wild-type strain . The resistant mutant had high activities of two enzyme systems: a glutathione-forming enzyme system (consisting of gamma-glutamylcysteine synthetase and glutathione synthetase) and a glyoxalase system (consisting of glyoxalase I and glyoxalase II) . Methylglyoxal resistance appeared to be due to the simultaneous increase in the activities of these two enzyme systems.

J Bacteriol, 1980 Oct, 144(1), 124 - 30
Effect of sterol side chains on growth and membrane fatty acid composition of Saccharomyces cerevisiae; Buttke TM et al.; Saccharomyces cerevisiae GL7 cells require exogenous sterol and unsaturated fatty acid for growth . When grown in the presence of cholesterol or 7-dehydrocholesterol, the cells incorporated less saturated fatty acid into phospholipids than cells grown with ergosterol, stigmasterol, or beta-sitosterol as the sterol source . This lower saturated fatty acid content was most pronounced in phosphatidylethanolamine, slightly less so in phosphatidylcholine, and least evident in phosphatidylserine and phosphatidylinositol . Growing the cells with the various sterols did not affect the ratios of individual phospholipids . The ability of strain GL7 to use 7-dehydrocholesterol as the only sterol supplement for growth was dependent upon the nature of the unsaturated fatty acids added to the growth medium . In the presence of linoleic, linolenic, or a mixture of palmitoleic and oleic acids, excellent growth was observed with either ergosterol, cholesterol, or 7-dehydrocholesterol . However, when the medium was supplemented with either oleic or petroselenic acid, the cells grew more slowly (oleic) or much more poorly (petroselenic) with 7-dehydrocholesterol than with ergosterol . A specific relationship between sterol structure and membrane fatty acid composition in yeast cells is implied.

Proc Natl Acad Sci U S A, 1980 Oct, 77(10), 5993 - 7
Growth of T-lymphoma cells in serum-free medium: lack of involvement of the cyclic AMP pathway in long-term cultures; Darfler FJ et al.; We have developed a serum-free, chemically defined growth medium containing casein, insulin, transferrin, testosterone, and linoleic acid in Dulbecco's modified Eagle's medium/Ham's F12 medium, 1:1 (vol/vol), for growing murine T lymphomas . This medium supports the growth in suspension of all murine T lymphomas tested, including S49, WEHI 7, EL4, BW5147, and R1.1 . Growth of these cell lines was maintained indefinitely with doubling times approaching those of cells grown in 10% (vol/vol) horse serum . This medium also supports the growth of several of the S49 variants of the beta-adrenergic receptor/adenylate cyclase/cyclic AMP/protein kinase pathway, suggeting little or no involvement of this pathway in the routine growth of S49 cells or in the mechanism of action of the factors in this defined medium . This serum-free medium should prove useful for studies of a variety of metabolic pathways and of differentiated functions of T-lymphoma cells.

Proc Natl Acad Sci U S A, 1980 Oct, 77(10), 5938 - 42
Spontaneous production of human interferon; Pickering LA et al.; Several established lines of human lymphoblastoid cells were evaluated for abilities to produce interferons . Some cell lines were able to produce interferon when induced with either Newcastle disease virus or Sendai virus, whereas others failed to produce detectable interferon when so induced . However, several cell lines were able to spontaneously produce interferon without induction . Spontaneously produced interferon was liberated by cells only during logarithmic growth phase, reaching levels ranging from about 10 reference units/ml of growth medium for some cell lines to 1000 reference units/ml for others . The interferons produced by induced lymphoblastoid cells and the spontaneously produced interferons were all characterized as type I human leukocyte interferon by high levels of cross-species antiviral activities on bovine cells and by neutralizations by antiserum to human leukocyte interferon but not by antiserum to human fibroblast interferon . However, analysis by electrophoresis in sodium dodecyl sulfate/polyacrylamide gels revealed that spontaneously produced interferon was less size heterogeneous than human leukocyte interferon, migrating as a single band of activity with a peak at 20,000 daltons, whereas human leukocyte interferon contained peaks of major activity at 23,000 and 18,000 daltons and virus-induced Namalva lymphoblastoid cell interferon migrated predominantly as the 18,000-dalton form . Also, although neither virus-induced primary leukocyte interferon nor any of the virus-induced lymphoblastoid cell interferons were neutralized by antiserum to mouse interferon, all of the spontaneously produced interferons were neutralized by antiserum to mouse interferons . These data suggest significant structural similarities between the active cores of certain interferons from phylogenetically diverse animal species.

Biochem J, 1980 Sep 15, 190(3), 673 - 83
Evidence of heterogeneity of protein-turnover states in cultured cells; Amenta JS et al.; Previous studies on L-cell cultures {Amenta & Sargus (1979) Biochem . J . 182, 847--859} have suggested: (a) that degradation of slow-turnover proteins occurs in a distinct cell state (D-state); (b) that cells randomly enter the D-state with a first-order transition constant, rapidly degrade cell protein, and return to a quiescent G0-state . In the present study we have tested the hypothesis that the putative D-state exists as a substate within A-state (non-replicating) fibroblasts . Rat-embryo fibroblasts were prelabelled with {14C}leucine and {3H}thymidine, 'chased' for 24 h, and then placed in fresh growth medium containing either vinblastine (10 microM) or colchicine (25 microM) for three successive 24 h periods . Cells trapped in mitosis were separated from the residual non-replicating cells and rates of protein synthesis, degradation and net accumulation were measured in both populations . We observed that significant protein degradation occurred only in the non-replicating population, although both populations showed equally high rates of protein synthesis induced by fresh growth medium . These data support the hypothesis that degradation of slow-turnover protein is heterogeneous, occurring only in A-state cells . A model that proposes a separate D-state within G0-phase successfully accounts for these observations and previous reports on this cell line {Amenta, Sargus & Baccino (1978) J . Cell . Physiol . 97, 267--283} showing no differences in degradation of the slow-turnover protein pool in growth-stimulated and stationary-phase fibroblast cultures.

Biochemistry, 1980 Sep 2, 19(18), 4197 - 201
In vivo effects of intercalating and nonintercalating drugs on the tertiary structure of kinetoplast deoxyribonucleic acid; Benard J et al.; The kinetoplast DNA (kDNA) of cultured Trypanosoma cruzi consists mostly in a large network of numerous minicircular molecules (approximately 25 000), with a very low degree of superhelicity . When the intercalating drugs ethidium bromide and 9-hydroxyellipticine were added to the growth medium in concentrations producing trypanocidal effects, the superhelicity of the kDNA was significantly increased . In contrast, the nonintercalating drug berenil had no effect on the superhelicity of kDNA . In the kDNA extracted from trypanosomes resistant to ethidium bromide or 9-hydroxyellipticine, those drugs induce similar effects, although to a lesser extent than in the wild strain.

Cell Tissue Kinet, 1980 Sep, 13(5), 497 - 503
Determination of non-proliferating cells in culture by combining flow cytometry with stathmokinetics; Bohmer RM; Colcemid was added to the growth medium of L-cells in monolayer culture . The proliferating cells continued their progression through the cycle up to metaphase, where they were arrested . At different times after Colcemid addition the cells were trypsinized, suspended immediately in a solution of the DNA-specific fluorescent dye Hoechst 33258 and analysed with a flow cytometer . The histograms were evaluated to give the fraction of cells in the 2c peak as a function of time after Colcemid addition . The flux into the 2c compartment being interrupted, the peak content decreased until all proliferating (G1) cells had entered S-phase . With increasing cell density or with increasing time after serum deprivation an increasing fraction of cells remained in the 2c peak at times greater than the normal G1 duration . The possibility of applying this method to the determination of non-proliferating cells in a population is discussed.

J Gen Virol, 1980 Sep, 50(1), 81 - 8
Metabolic requirements for the maturation of respiratory syncytial virus; Peeples M et al.; The metabolic processes required for maturation of respiratory syncytial (RS) virus was determined by testing with metabolic inhibitors in HeLa cells that had been trypsinized 18 h p.i . Although > 90% of the virus synthesized by that time remained cell-associated, treatment with trypsin inactivated at least 90% of the cell-associated virus . The trypsinized cells, when re-plated in virus growth medium, immediately resumed virus synthesis and this continued exponentially for at least 10 h, during which as little as 1 to 2 p.f.u./cell of new infectious virus could be detected . Treatment of these cells with 6-azauridine revealed that no further synthesis of virus RNA is required for a full yield of infectious virus . Treatment with cycloheximide, 2-deoxy-D-glucose and cytochalasin B suggested that neither protein synthesis nor glycosylation is required for the first 1 to 2 h following the resumption of virus production, but both processes are required for a full yield . These results suggest that with RS virus, the maturation process itself does not require RNA or protein synthesis or glycosylation . Trypsin, in addition to inactivating accumulated virus, released non-infectious particular material . The polypeptides of this material were similar to those of the RS virion except for a deficiency in glycoproteins . The trypsin apparently cleaved the virus glycoproteins on cell surfaces, which resulted in the inactivation of cell-associated virus and the release of a glycoprotein-deficient particle from cell surfaces . The pool of virus glycoproteins is evidently the limiting factor in infectious virus production by cells trypsinized 18 h p.i . The non-glycosylated polypeptides of cell-associated virus are the same as those of free virus.

Mikrobiologiia, 1980 Sep-Oct, 49(5), 746 - 50
{Fatty acids of actinomycete phospholipids}; Koval'chuk LP et al.; The fatty acid composition of phospholipids was studied in different actinomycetes growing in two media in order to detect their biological activity . The total phospholipids of the actinomycetes did not differ qualitatively and their composition was represented by the same series of fatty acids (C13--C19) . The qualitative composition and the quantitative content of fatty acids in phospholipids depended on the composition of the growth medium . When the actinomycetes were cultivated in a complex medium, the proportion between fatty acids (C14:0, C14:1, C16:0, C17:0, C17:1, C18:1, C18:2) changed . The qualitative composition of fatty acids in phospholipids varied among the cultures . However, the content of palmitoleic, palmitic and oleic acids was elevated in all of the cultures . Under the given experimental conditions, the actinomycetes were found to synthesize phospholipids containing fatty acids with a high degree of unsaturation (mainly at the account of C16:1 and C18:1 acids).

J Bacteriol, 1980 Sep, 143(3), 1332 - 44
Synthesis and function of ribonucleic acid polymerase and ribosomes in Escherichia coli B/r after a nutritional shift-up; Shepherd N et al.; The syntheses of stable ribosomal ribonucleic acid (RNA) and transfer RNA in bacteria depend on the concentration and activity of RNA polymerase and on the fraction of active RNA polymerase synthesizing stable RNA . These parameters were measured in Escherichia coli B/r after a nutritional shift-up from succinate-minimal to glucose-amino acids medium and were found to change in complex patterns during a 1- to 2-h period after the shift-up before reaching a final steady-state level characteristic for the postshift growth medium . The combined effect of these changes was an immediate, one-step increase in the exponential rate of stable RNA synthesis and thus of ribosome synthesis . This suggests that the distribution of transcribing RNA polymerase over ribosomal and nonribosomal genes and the polymerase activity are continuously adjusted during postshift growth to some growth-limiting reaction whose rate increases exponentially . It is proposed that this reaction is the production of amino-acylated transfer RNA and that is exponentially increasing rate results in part from a gradually increasing concentration of aminoacyl transfer RNA synthetases after a shift-up . This idea was tested and is supported by a computer simulation of a nutritional shift-up.

Biochim Biophys Acta, 1980 Aug 5, 592(1), 103 - 12
Reciprocal formation of plastocyanin and cytochrome c-553 and the influence of cupric ions on photosynthetic electron transport; Bohner H et al.; The green alga Scenedesmus acutus is able to synthesize plastocyanin and cytochrome c-553 . The concentrations of plastocyanin and cytochrome c-553 vary inversely in response to the cupric-ion concentrations of the growth medium (Bohner, H . and Boger, P . (1978) FEBS Lett . 85, 337-339) . Both proteins form a homogeneous donor pool to the reaction center of Photosystem I . This donor pool can be varied quantitatively and qualitatively by different growth conditions . These variations have no influence on algal growth or photosynthetic electron transport as measured in vivo by oxygen evolution, fluorescence induction and cytochrome f-553 and c-553 redox reactions using Cu2+ concentrations of less than 10 microM in the culture medium . At higher cupric-ion concentrations, which already retard algal growth, specific sites of the photosynthetic electron-transport chain are affected: the oxidizing side of Photosystem II and the reducing side of Photosystem I.

J Clin Microbiol, 1980 Aug, 12(2), 291 - 3
Production of p-hydroxyhydrocinnamic acid from tyrosine by Peptostreptococcus anaerobius; Lambert MA et al.; Peptostreptococcus anaerobius was found to metabolize tyrosine to p-hydroxyhydrocinnamic acid {3-(p-hydroxyphenyl)propionic acid} . This acid was detected in spent growth media by gas-liquid chromatography, and its identity was confirmed by mass spectrometry.

Appl Environ Microbiol, 1980 Aug, 40(2), 274 - 81
Mutation induced by drying of Escherichia coli on a hydrophobic filter membrane; Asada S et al.; Drying of Escherichia coli to a required cellular water level was conducted on a hydrophobic membrane at the corresponding relative humidity . Mutation from an arginine auxotroph to the prototroph was induced by drying to a water activity (aw) of 0.53 and below, but not to an aw of 0.75 and above . The critical aw below which mutation occurred in the course of drying was similar to that for induction of deoxyribonucleic acid (DNA) strand breakage in the bacteria . Some ultraviolet or gamma-irradiation-sensitive strains, e.g., strains of carrying recA, recB, and uvrA recA were more sensitive to drying than the wild-type strains or strains carrying uvrA and polA . The DNA strand breakage of every strain was observed to be to a similar extent after drying to an aw of less than 0.53 . The drying-resistant strains repaired the damaged DNA partially during postdrying incubation in a growth medium but not in phosphate buffer solution, while the drying-sensitive strains could not at all . Significant mutation on drying occurred in the wild-type strains, strains carrying uvrA and polA, but not in strains carrying recA . It is, therefore, concluded that the mutation is caused by errors in rec-dependent repair of the drying-induced breakage in DNA.

Can J Microbiol, 1980 Aug, 26(8), 912 - 20
Influence of S-adenosylmethionine on DAPI-induced fluorescence of polyphosphate in the yeast vacuole; Allan RA et al.; Use of the fluorochrome 4',6-diamidino-2-phenylindole.2 HCl (DAPI) in ultraviolet microscopy revealed fluorescent objects in Brownian motion within the vacuoles of seven species of yeast . The abundance of these bodies increased when cells of Saccharomyces cerevisiae were transferred from growth medium to a glucose-phosphate solution, indicating that they contain polyphosphate . In addition, the effect on vacuolar fluorescence of supplementing a defined growth medium with amino acids provided evidence that they also contained S-adenosylmethionine . These deductions were supported by in vitro studies of the interaction and fluorescence of polyphosphate, S-adenosylmethionine, and DAPI . Vacuolar fluorescence of cells in suspension in flucose-phosphate solution was less after addition of exogenous arginine, lysine, or glutamine but not after addition of alanine, aspartic acid, or methionine . Mithramycin was superior to DAPI as a fluorochrome for ultraviolet demonstration of yeast nuclei since it stained the nuclei much more intensely and did not fluoresce with other material in the cells.

J Bioenerg Biomembr, 1980 Aug, 12(3-4), 249 - 64
Partial characterization of the plasma membrane ATPase from a rho0 petite strain of Saccharomyces cerevisiae; McDonough JP et al.; Crude membrane preparations of a rho0 mutant of Saccharomyces cerevisiae exhibit Mg2+-dependent ATPase activity . Over the optimal pH range, 5.0-6.75, the apparent Vmax of the enzyme equals 590 nmoles of ATP hydrolyzed per minute per milligram protein, with an apparent Km for ATP of 1.3 mM . ATP hydrolysis is insensitive to ouabain, venturicidin, aurovertin, and the protein inhibitor described by Pullman and Monroy; inhibited by oligomycin (at high concentrations) and sodium orthovanadate, and it is sensitive to dicyclohexylcarbodiimide, p-hydroxymercuribenzoate, hydroxylamine, sodium fluoride, and sodium iodoacetate . The pH optimum and the inhibitor pattern distinguish the plasma membrane enzyme from the mitochondrial F1 ATPase still present in these cells (this activity is sensitive to efrapeptin, aurovertin, and the protein inhibitor, but resistant to DCCD) . In addition, the activity of the plasma membrane enzyme and its affinity for ATP are responsive to changes in the composition of the growth medium, with the highest activity observed in cells grown on methyl-alpha-D-glucoside, a sugar which results not only in partial release from catabolite repression but also requires the induction of an active transport system for growth.

Biochem J, 1980 Jul 15, 190(1), 1 - 15
Control of the flux in the arginine pathway of Neurospora crassa . The flux from citrulline to arginine; Flint HJ et al.; The arginine pathway is a complex one, having many branch points and effector interactions . In order to assess the quantitative role of the various mechanisms that influence the flux in the pathway, the system was divided experimentally into two moieties by the introduction of a genetic block abolishing ornithine carbamoyltransferase activity . This normally produces citrulline from ornithine within the mitochondria . The endogenous citrulline supply was replaced by citrulline in the growth medium, and control of the influx rate was achieved by using glycine or histidine as uptake inhibitors . By modulating the influx rate over a large range of values, the importance of such factors as reversibility, saturation, inhibition and induction in affecting the flux and the sizes of intermediate pools between citrulline and arginine was assessed . The role of expansion fluxes as important controls in the exponentially growing system was established.

Transfusion, 1980 Jul-Aug, 20(4), 462 - 4
The action of anti-A,AB isoantibodies on group A, AB-positive human lymphocytes; Skinnider LF et al.; Human lymphocytes of group A and group AB individuals were exposed to anti-A,B antisera both in growth medium and in fresh autologous plasma . Agglutination developed slowly but was present in all positive specimens by four hours and was negative in the group O controls even after 24 hours . There was no loss of viability even in the presence of the fresh plasma . The lymphocytes in the agglutinates appeared larger and more irregular suggesting stimulation of the lymphocytes but assessment by 3H thymidine uptake showed no evidence of a mitogenic response.

J Cell Physiol, 1980 Jul, 104(1), 47 - 52
Increased neurite development and plasminogen activator expression by exposure of human neuroblastoma cells to plasminogen-deficient growth medium; Becherer PR et al.; Growth of human neuroblastoma strain SK-N-SH in a plasminogen-deficient medium results in about a 40% increase in the number of differentiated cells (cells with a neurite-like process at least 50 micrometers in length) and about a five-fold increase in the amount of plasminogen activator liberated per cell . Plasminogen deficiency has no effect on the growth rate of SK-N-SH cells . These results are consistent with the hypothesis that plasminogen activator is involved in neuroblast development.

J Cell Biol, 1980 Jul, 86(1), 156 - 61
Rapid retraction of neurites by sensory neurons in response to increased concentrations of nerve growth factor; Griffin CG et al.; The phenomenon of growth cone (GC) and neurite retraction resulting from a rapid incrase in concentration of the trophic molecule NGF was studied . Neurite outgrowth from explants of 8-d chick embryo dorsal root ganglia was achieved at very low NGF concentrations with heart conditioned medium during overnight culture . Quickly incrasing the NGF concentration in the growth medium dramatically affected GC and neurite morphology: the majority of GCs and neurites collapsed and retracted towards the cell body over a course of approximately 2-5 min . Retraction was elicited by increasing NGF levels from 0 to 0.05 ng/ml to as little as 0.5 ng/ml but did not occur if the NGF concentration during the initial overnight culture period exceeded 0.8 ng/ml, regardless of how much the concentration was elevated . Similar concentration changes of cytochrome c or insulin did nt result in retraction . Neurites that had been separated from their cell bodies by cutting close to their exit from the explant still retracted when NGF levels were raised . Cytochalasin B reversible inhibits retraction, whereas colchicine allows retraction to occur . Observation of cell-substratum adhesion during retraction revealed that some adhesion points remain during retraction and that they correspond to the ends of NGF leels and that it may involve microfilaments in the neurite cytoskeleton . The NGF concentration changes that elicit neurite retraction suggest that a primary event in retraction may be increased occupancy of a high-affinity NGF receptor on neurites.

J Gen Microbiol, 1980 Jul, 119(1), 17 - 26
Growth medium constituents contaminating mycoplasma preparations and their role in the study of membrane glycoproteins in porcine mycoplasmas; Nicolet J et al.; Several Mycoplasma and Acholeplasma species chosen at random and solubilized with sodium dodecyl sulphate showed a common periodic acid-Schiff positive band with an apparent molecular weight of about 64 000, when examined by polyacrylamide gel electrophoresis . Another more cathodic minor band was detected in M . hyopneumoniae and M . flocculare . The common periodic acid-Schiff positive band appeared when a precipitate of serum constituents of the uninoculated growth medium after incubation was examined . The minor band was identified as a serum glycoprotein contaminating mycoplasmas grown in the presence of swine serum . We draw attention to the compounds as a possible source of error in serological tests or in the lymphocyte stimulation response . After lithium diiodosalicylate solubilization and aqueous phenol extraction, polyacrylamide gel electrophoresis showed a periodic acid-Schiff positive band in membranes from M . hyopneumoniae (molecular weight 75 000) and M . hyorhinis (molecular weight 80 000), suggesting the presence of a membrane glycoprotein . Such a glycoprotein was absent from A . granularum . Since the common periodic acid-Schiff positive band was not extracted by aqueous phenol, this growth medium constituent did not contaminate the preparations of membrane glycoproteins . However, the minor band was present in glycoprotein preparations of M . hyopneumoniae grown in the presence of swine serum.

In Vitro, 1980 Jul, 16(7), 616 - 28
Chemically defined serum-free media for the cultivation of primary cells and their susceptibility to viruses; Weiss SA et al.; Chemically defined media SFRE-199-1 for the growth and SFRE-199-2 for the maintenance of primary baboon kidney (Bak) cell cultures were formulated by supplementing medium M199 with insulin, sodium pyruvate, zinc sulfate, and increasing arginine-HCl, cysteine, cystine, L-glutamine, L-glutamatic acid, glycine, histidine, tyrosine, and glucose to maximally active nontoxic concentrations . For prolonged maintenance of the cells, physiological pH control, and blocking of excessive lactic acid accumulation in the spent medium of the cell cultures, it is necessary to supplement the medium containing Earle's balanced salts with D-(+)galactose . The cells grew and were maintained equally well on glass or polystyrene surfaces . Selenium, when added to growth medium or substituted for insulin and zinc sulfate, did not stimulate cell growth . Electron microscopy showed that numerous dense particles, approximately 250 to 400 A in diameter, with the appearance of glycogen, were found throughout the cytoplasm in the cells grown in SFRE-199-1 and maintained in SFRE-199-2 . Echovirus types 1 to 3, poliovirus types 1 to 3, coxsackievirus types B2, B4, B5, Herpes-virus hominis type 1, simian herpesvirus H . simiae and SA8, and simian adenovirus SV34 when titrated in primary Bak cells and grown and maintained in SFRE-199-1 and 2, respectively, developed titers comparable to those obtained in conventionally grown and maintained cells.

Clin Chem, 1980 Jul, 26(8), 1127 - 32
Identification of fluorescent Pseudomonas species; Shelly DC et al.; Fluorescent pseudomonads may be identified through examination of the fluorescence "profiles" of diffusible pigments released into the growth medium . The profiles are obtained in the form of an emission-excitation matrix, the elements of which correspond to fluorescence intensity as a function of multiple exciting and emitting wavelengths . Use of a rapid scanning fluorometer to acquire these data provides a relatively fast method for identification of the microorganisms after incubation . This procedure can potentially be expanded for the identification of other microorganisms.

Biochim Biophys Acta, 1980 Jun 23, 618(3), 399 - 406
Prostaglandin I2 synthesis and elevation of cyclic AMP levels in 3T3 fibroblasts; Claesson HE; Arachidonic acid and prostaglandin H2 elevate the levels of adenosine 3':5'-monophosphate (cyclic AMP) in Balb/c 3T3 fibroblasts . This effect was inhibited by 15-hydroperoxy-5,8,11,13-eicosatetraenoic acid, an inhibitor of prostaglandin I2 synthase (Claesson, H.-E., Lindgren, J.A . and Hammarstr!om, S . (1977) FEBS Lett . 81, 415-418) . After addition of arachidonic acid to 3T3 cultures, cellular cyclic AMP levels and growth medium concentrations of 6-ketoprostaglandin F1 alpha (degradation product of prostaglandin I2) were quantitatively determined . The stimulatory effect of exogenously-added prostaglandin I2 on cellular cyclic AMP levels was also determined . The results indicate that the endogenous production of prostaglandin I2 is sufficient to explain the stimulatory action of arachidonic acid on cyclic AMP formation in 3T3 fibroblasts.

J Biol Chem, 1980 Jun 10, 255(11), 5448 - 53
A glycoprotein from neurites of differentiated neuroblastoma cells; Littauer UZ et al.; Neurites were prepared by a novel method from differentiating mouse neuroblastoma cells . When electrically differentiated cells were labeled metabolically with L-{3H}fucose or D-{3H}glucosamine, both the neurites and the surface membranes showed the presence of a glycoprotein of apparent Mr = 200,000 . In contrast, the level of this glycoprotein was reduced in the surface membranes from nondifferentiated cells and a radioactive glycoprotein of similar molecular weight was found in the growth medium . The method for the isolation of neurites is of potential usage in distinguishing specific proteins associated with growing neurites.

Biochim Biophys Acta, 1980 Jun 6, 598(3), 606 - 15
Alanine transport by Chinese hamster ovary cells with altered phospholipid acyl chain composition; Ryan J et al.; The Na+-dependent transport of alanine has been examined in Chinese hamster ovary (CHO) cells as a function of the fatty acid composition of their membrane lipids . Significant changes in the fatty acid composition of the CHO cell phospholipids were achieved by supplementation of the growth medium with specific saturated (palmitate) or monoenoic (oleate) free fatty acids . Arrhenius plots of the temperature-dependent uptake of alanine were constructed for cells of altered fatty acid composition . Alanine uptake was characterized by a single discontinuity in the Arrhenius plot . The temperature of this break was observed to be dependent upon the fatty acid composition of the cell phospholipids, ranging from 16 degrees C for cells enriched with oleate to 32 degrees C for cells enriched in palmitate . Calculation of the Km value for the uptake process showed no significant change with temperature or fatty acid supplementation . Correlations are made between the physical state of the membrane lipids and the temperature-dependence for alanine transport . The results are discussed in terms of membrane fatty acid composition, ordered in equilibrium fluid phase transitions and amino acid transport.

Mol Biochem Parasitol, 1980 Jun, 1(3), 143 - 9
Production and secretion of Leishmania braziliensis proteins; Hernandez AG et al.; The kinetics of secretion of proteins by Leishmania braziliensis was followed by incorporation of {3H}leucine into macromolecules produced by the cells which are released into the growth medium . About 10% of the total protein synthesized by actively growing cells is secreted . Cycloheximide (100 microgram/ml) and puromycin (0.5 mM) inhibited the incorporation of labelled leucine by 85 and 99%, respectively . The secreted proteins do not seem to result from cell lysis since, first, the kinetics of production are linear and, secondly, less than 1% of thymidine or uridine incorporated by the cells is found in the medium . Cells grown with {3H}leucine and then transferred to fresh medium show two phases of secretion . During the first six hours, it is slow and reaches a plateau . The release increases about ten-fold during the next six hours . An analysis of the secreted material showed that following precipitation with methanol and sodium acetate, three isotopically labelled peaks were eluted from Sephadex G-120-150 . The first of these, containing 50% of the radioactivity, did not react with anti-leishmanial serum, while the last two did . Since the last two fractions could be labelled with {3H}glucosamine as well as {3H}leucine it is suggested that they are glycoprotein in nature and are similar to the products released by other species of Leishmania.

J Med Genet, 1980 Jun, 17(3), 187 - 90
Alkaline phosphatase activity of normal and cystic fibrosis fibroblasts; Aitken DA et al.; Alkaline phosphatase (ALP) activities were compared in fibroblasts from three cystic fibrosis patients and two normal controls after culturing the cells in normal growth medium and in medium containing Tamm-Horsfall glycoprotein, isoproterenol, and theophylline . No consistent alterations in ALP activities were noted, either between the same cell lines grown under different conditions, or between normal and cystic cell lines . It is concluded that it is not possible to use changes in ALP activity in cultured cells for the prenatal diagnosis of cystic fibrosis.

J Cell Biol, 1980 Jun, 85(3), 777 - 85
Inhibition of DNA chain elongation in a purine-auxotrophic mutant of Chinese hamster; Zannis-Hadjopoulos M et al.; DNA synthesis has been examined in a purine-auxotrophic mutant cell line of Chinese hamster (V79 pur 1) under conditions of purine deprivation . At 6 h after the removal of purines from the growth medium, there is a decrease in semiconservative DNA replication . Alkaline velocity centrifugation of the DNA synthesized during a 1-min pulse under conditions of purine deprivation shows that approximately 50% of the newly replicated DNA is the size of Okazaki pieces . These are not incorporated into bulk DNA during a 1-h chase . If the purine supply is restored to the growth medium, these short DNA pieces are jointed to full-sized DNA within 1 h . DNA fiber autoradiolgraphy reveals a retardation in the rate of DNA replication fork movement but no apparent inhibition of initiation of synthesis on replication units within clusters actively engaged in replication . Our results indicate that purine deprivation specifically inhibits elongation of nascent dna chains.

Int J Radiat Biol Relat Stud Phys Chem Med, 1980 Jun, 37(6), 591 - 600
Repair of potentially lethal damage in unfed plateau phase cultures of Ehrlich ascites tumour cells . II . Monolayer cultures; Iliakis G; Monolayer cultures of EAT cells when plated immediately after irradiation show a desrease in survival as they "age" in the plateau phase of growth . This decrease, which is manifest as a diminution of the shoulder width of the survival curve down to values approaching zero, is reversible if the cells are kept in their growth medium for some hours after irradiation before trypsinization and plating . Survival curves obtained by this holding procedure are similar in shape to those shown by exponentially growing or early plateau phase cells . We interpret this effect in terms of repair of potentially lethal damage which occurs after immediate plating in young cultures but only declared during plating in cultures which have "aged" in the plateau phase . The kinetics of this repair and the effects caused by the addition of serum after irradiation in the cultures have been studied.

In Vitro, 1980 Jun, 16(6), 507 - 15
Feasibility studies of oncornavirus production in microcarrier cultures; Manousos M et al.; Studies conducted with virus-infected monolayer cell cultures have demonstrated the feasibility of producing several tumor-associated viruses in microcarrier (mc) cultures (Sephadex G50 beads treated with DEAE-chloride) . The efficiency of cell adherence to mc varied with the cell type, the pH of the growth medium, and the stirring force applied to keep the mc in suspension . Most cells attached firmly to the mc and could not be removed easily with routine trypsinization procedures . Techniques using Enzar-T and Pronase were effective in detaching cells from mc in 10 to 15 min while retaining 95% cell viability . After detachment, Ficoll gradients were used for rapid and complete separation of viable cell suspensions from the mc . Retrovirus production in large volumes of mc cultures was investigated with periodic harvesting of growth fluids . Physical, biochemical, and biological properties of the Mason-Pfizer monkey virus and the RD114 virus recovered from the mc cultures were identical to those produced in conventional cultures . The utilization of mc has several applications in research and short-term cultures, but the as-yet-unsolved technical problems met were found to be serious limitations when attempting mass cell culturing on a long-term basis.

Rev Esp Fisiol, 1980 Jun, 36(2), 155 - 61
{Variations in the P . aeruginosa polysaccharide synthesis conditioned by aminosugars (author's transl)}; Jorba G et al.; The production of capsular and extracellular polysaccharides by P . aeruginosa species was conditioned by the presence of hexosamines in the growth media . Glucosamine and most remarkably galactosamine, strongly reduced the production of extracellular polysaccharides . Studies on the dynamics of the polysaccharide production show that these products are used by the cells after the carbon source (glucose of glucosamine) of the growth medium has been exhausted . These results suggest the presence of adaptative hexosaminidases . The concentration of total hexosamines in the polysaccharides changed according to the carbon source (glucose, glucosamine or galactosamine) and so did the molecular composition as demonstrate by IR spectrometry . IR spectra show variations in the intensity of some characteristic bands, as those located at 1,650-1,560 cm-1 (N-acetyl-hexosamines), 1,420 and 1,320 cm-1 (aminosugars) and at 1,380 cm-1 . This last band disappears from polysaccharides after prolonged incubation . Bands between 1,300 and 700 cm-1 varied and even disappeared according to the aminosugar employed and the length of the incubation period.

J Cell Physiol, 1980 Jun, 103(3), 455 - 66
On the effect of inhibitors of mitochondrial macromolecular-synthesizing systems and respiration on the growth of cultured chick embryo cells; Morais R; We have found that chick embryo fibroblasts (DEF) cultivated in the presence of tryptose phosphate broth (TPB) are inherently resistant to the growth inhibitory effect of ethidium bromide (EB) . As demonstrated by cytochrome oxidase activity and oxygen consumption measurements, analyses of reduced-minus-oxidized cytochrome spectra and electron microscopic observations, TPB did not seem to prevent the inhibitory effect of EB on mitochondrial DNA transcription . EB-treated chick cell populations cultivated in the presence of TPB behave essentially the same as populations treated with chloramphenicol (CAM) and grow with mitochondria devoid of a functional respiratory chain . In contrast to CAM-treated CEF populations, however, the respiratory activity of EB-treated cell populations did not reappear when the cells were shifted back to EB-free medium . Attempts to demonstrate that TPB confers resistance to the growth inhibitory effect of carbomycin and mikamycin, inhibitors of the mitochondrial protein-synthesizing system, have failed, the drugs being cytotoxic at doses where protein synthesis on mitoribosomes is not suppressed . On the other hand, the present results demonstrated that chick cell populations proliferate in the presence of the respiratory inhibitors rotenone, antimycin A, amytal and oligomycin whether or not TPB is present in the growth medium.

J Cell Physiol, 1980 Jun, 103(3), 385 - 92
In vitro proliferation and lifespan of bovine aorta endothelial cells: effect of culture conditions and fibroblast growth factor; Duthu GS et al.; The effect of culture conditions on calf dorsal aorta endothelial cells was studied . Population doubling time varied as a function of the cell seeding density, growth Medium, serum supplement, and concentration of fibroblast growth factor (FGF) . The shortest population doubling time was found for cells (population doubling level 0--30) grown in Eagle's Minimal Essential medium (MEM) supplemented with 10% fetal bovine serum (FBS) and 100 ng/ml FGF . The stimulatory effect of FGF on bovine endothelial cell proliferation was dependent on cell inoculation density . FGF significantly increased cell division rate at cell inocula less than 1 x 10(4) cells/cm2 but not at higher densities . The population doubling time and cell size increased as the mass culture population doubling level increased . The replicative lifespans of bovine endothelial cells grown in medium supplemented with 20% FBS were 10--15% greater than parallel cultures supplemented with 10% FBS . Cultures grown in medium supplemented with 10% FBS and 50 ng/ml FGF showed a 50% increase in replicative lifespan compared to cultures grown in medium supplemented with 10% FBS alone . When FGF was used the increase in the number of doublings was a function of the length of time the cells were grown in the presence of FGF . This report extends comparable observations on the in vitro aging of human diploid fibroblasts to bovine endothelial cells.

J Cell Biol, 1980 Jun, 85(3), 910 - 5
Evidence for a pronounced secretion of cyclic AMP by Tetrahymena; Nandini-Kishore SG et al.; The unicellular eukaryote Tetrahymena pyriformis secretes significant amounts of cyclic AMP into its external medium . Cells transferred from growth medium into any of the following three different non-nutrient media: (a) 5 mM phosphate buffer containing 47 mM NaCl and 1 mM MgSO4, (b) 10 mM Tris, or (c) 1.3 mM Tris containing 1 mM citrate and 1 mM Ca(OH)2, released to the outside almost 60--80% of the total cyclic AMP produced during 2--5 h of incubation . Tris-citrate-Ca+2 medium was chosen for further experiments because of its minimal nonspecific interference in the cyclic AMP radioimmunoassay . The identity of the secreted material recognized as cyclic AMP by radioimmunoassay was confirmed by demonstrating its almost complete hydrolysis with commerical beef heart phosphodiesterase . Furthermore, the radioimmunoassay-active material in the concentrated medium co-chromatographed on paper with {3H}cyclic AMP, as judged by assay of the eluted material . After resuspending cells in Tris-citrate-Ca2+ medium, the extracellular concentration of cyclic AMP rose steadily over a 5-h period, reaching a level equvalent to approximately 35--50 pmol cyclic AMP/10(6) cells vs . an internal cyclic AMP quantity at 5 h of 8--10 pmol/10(6) cells . After 5 h, the level of extracellular cyclic AMP reached a plateau . There was no degradation or uptake of external cyclic AMP by the cells during this period.

Proc Natl Acad Sci U S A, 1980 Jun, 77(6), 3445 - 9
Differential expression of type I and type II cyclic AMP-dependent protein kinases during cell cycle and cyclic AMP-induced growth arrest; Haddox MK et al.; The activation state of cyclic AMP-dependent protein kinase(s)(ATP:protein phosphotransferase, EC 2.7.1.37) is transiently increased 2-fold as a function of G1 progression in mitotically synchronized Chinese hamster ovary cells . The cellular content of type I kinase increases concomitantly with the increase in general protein, whereas the activity of type II kinase increases as a function of time in G1 to a maximum at the G1/S border . In contrast, in the presence of dibutyryl-cyclic AMP, there is a decrease of type II kinase and a several-fold increase of type I kinase . In proliferating cells, the ratio of type I to type II was 0.37, while in the dibutyryl-cyclic AMP growth-arrested cells it was 3.96 . The increase in type II kinase during G1 transition and the increase in type I kinase during dibutyryl-cyclic AMP treatment were dependent on protein synthesis . A similar pattern of type I and type II kinase expression during cell cycle progression occurred in Rat-1 fibroblasts and Rat-1 cells transformed by Rous sarcoma virus . The inclusion of dibuityryl-cyclic AMP in the growth media promoted a marked increase in type I holoenzyme, which was inhibited by cycloheximide, and a decrease in type II kinase . Neither AMP nor sodium butyrate had any effect on cellular kinase levels, whereas 8-bromo-cyclic AMP mimicked the action of dibutyryl-cyclic AMP . Estimation of half-lives for the kinase types showed that there was little turnover of either type during normal G1 progression, rapid turnover of both types as cells exited from mitosis, and selective turnover of type II upon addition of dibutyryl-cyclic AMP.

J Biol Chem, 1980 May 25, 255(10), 4808 - 13
Chinese hamster ovary cells resistant to beta-aspartylhydroxamate contain increased levels of asparagine synthetase; Gantt JS et al.; The growth of Chinese hamster ovary cells in a complete medium lacking asparagine is inhibited by beta-aspartylhydroxamate . The inhibition is overcome by the presence of asparagine in the growth medium . beta-Aspartylhydroxamate inhibits the activity of both asparagine synthetase and asparaginyl-tRNA synthetase in vitro . beta-Aspartylhydroxamate-resistant clones of Chinese hamster ovary cells have been isolated and three of these have been characterized . One clone, AH12, is 3-fold more resistant to beta-aspartylhydroxamate than the parental line and has 2 times higher levels of asparagine synthetase activity . Strains AH2 and AH5 are 6- to 7-fold more resistant to beta-aspartylhydroxamate and have 5 times higher levels of asparagine synthetase . The regulation of the expression of asparagine synthetase is altered in all three resistant cell lines . Whereas asparagine synthetase activity varies 2- to 3-fold in response to the asparagine content of the medium or to the extent of aminoacylation of tRNALeu in the parental cells, the activity of asparagine synthetase in the resistant cells is elevated under all growth conditions . No significant changes in the Km for substrates, Ki for beta-aspartylhydroxamate, or thermal stability were found for the asparagine synthetase of the resistant cells . These variants should prove useful in understanding the mechanisms involved in regulating the levels of asparagine synthetase in mammalian cells.

Biochim Biophys Acta, 1980 May 8, 598(1), 16 - 26
Studies on the size, location and turnover of calcium pools accessible to growing Tetrahymena cells; Kusamran K et al.; Tetrahymena pyriformis cells in the logarithmic phase of growth accumulate 2.5--3.75 times as much calcium per unit volume as is present in the growth medium . It appears that most of this calcium is stored in a non-ionic form, with approximately 30% existing in the cilia, near its site of action in effecting ciliary reversal . The exchange of extracellular 45Ca2+ with the major internal pools is extremely rapid, exhibiting a t1/2 of less than 0.5 h . Sites located on the cilia are responsible for 35--50% of Ca2+ influx, with the remainder entering through other positions on the cell surface.

Mikrobiologiia, 1980 May-Jun, 49(3), 473 - 8
{Composition of a nutrient medium for the continuous cultivation of Thiobacillus ferrooxidans}; Denisov GV et al.; Thiobacillus ferrooxidans was found to require the following biogenous elements for constructing its biomass: phosphorus, 14 +/- 2; magnesium, 2 +/- 0.5; potassium, 5 +/- 1; nitrogen, 100 +/- 10 (milligrams per gram of dry biomass) . It was shown that the concentrations of biogenous elements in the growth medium, which did not limit the biosynthesis, did not exceed 5 divided by 10 mg per litre for all of the studied elements . The medium 9K that is commonly used for growing Thiobacillus ferrooxidans has been found to differ significantly from the calculated one with the balanced ratio of biogenous elements.

Mikrobiologiia, 1980 May-Jun, 49(3), 389 - 95
{Respiratory system of Endomyces magnusii growing on media with a high fermented sugar content}; Zviagil'skaia RA et al.; No repression of the synthesis of respiration enzymes was found in the petite--negative yeast Endomyces magnusii under the action of high concentrations of a fermented sugar in the growth medium . Transfer of the cells from 0.3% to 5% sucrose (or glucose) did not result in a decrease in their respiration activity or molar content of cytochromes a+a3 (components of the respiratory chain most sensitive to the glucose repression in other yeast species) . The respiration of intact cells grown on 5% sucrose (glucose) was inhibited by 80--94% with cyanide and by 90--98% with cyanide plus salicyl hydroxamate . These data indicate that mitochondria play a predominant role in the oxidative activity of cells . Mitochondria from cells grown on 5% sucrose (glucose) hardly differ in their metabolic activity, effectiveness of oxidative phosphorylation, coupling, and regulation of the metabolic state from mitochondria isolated from completely derepressed cells (the stationary state from mitochondria isolated from completely derepressed cells (the stationary phase of growth, complete exhaustion of the substrate in the course of growth in a medium containing 0.6% sucrose).

In Vitro, 1980 May, 16(5), 426 - 32
Analysis of growth dynamics and the relationship to the fluctuations in alkaline phosphatase in two strains of cells of HeLa S3 derivation; Brahmacupta P et al.; Studies have been carried out to determine an association between glucocorticoid-induced changes in the pattern of growth and the fluctuations of alkaline phosphatase in two HeLa strains . The results showed that growth arrest in steroid-treated cells did not have the characteristics of density-induced growth inhibition . Alkaline phosphatase increased with increased cell density, the increase being greater than control in steroid-treated cells of the "inducible" strain (HeLa S3G, HeLa65) and less than control in the "suppressible" strain (HeLa S3K, HeLa71) . Increased serum concentration in the growth medium (0 to 20%) caused an increase in alkaline phosphatase in S3G strain and a decrease in the S3K strain.

Cancer Res, 1980 May, 40(5), 1722 - 6
Effects of estrogen and antiestrogen on DNA polymerase in human breast cancer; Edwards DP et al.; We have investigated the effects of estrogens and antiestrogens on cellular DNA-dependent DNA polymerase activity in human breast cancer, using as a model the MCF-7 human breast cancer cell line which contains estrogen receptor . 17 beta-Estradiol had little if any effect on cytosol DNA polymerase activity or growth (total DNA per flask) of MCF-7 cells . Incubation of the cells for 4 to 6 days with the antiestrogen nafoxidine, however, resulted in a dose-dependent reduction in cytosol DNA polymerase activity to one-half that observed in untreated cells . Enzyme activity in antiestrogen-treated cells was restored to levels contained in untreated cells by removing antiestrogen from the growth medium and incubating the cells for an additional 4 days with 17 beta-estradiol . The restoration required estrogenic steroids specifically, and the time course, magnitude, and dose dependence of the response were similar to estrogen-stimulated increases in DNA polymerase activity described in other estrogen target tissues . Estrogen-mediated reversal of antiestrogen suppression of DNA polymerase activity was paralleled by increases in total DNA synthesis.

Pflugers Arch, 1980 May, 385(1), 37 - 43
Role of calcium in the regulation of acetylcholine receptor synthese in cultured muscle cells*; Birnbaum M et al.; Embroyonic muscles differentiated in vitro were used to study the effects of intracellular Ca2+ ({Ca2+1}i) variations on the amount of acetylcholine receptors ({AChR}) in the cell membrane . 2 . Increased Ca2+ concentration in the growth medium ({Ca2+}o) caused a marked elevation of AChR levels, apparently through de novo synthesis . 3 . Agents known to increase {Ca2+}i and its accumulation in the sarcoplasmic reticulum (SR), such as ionophore A23187, sodium dantrolene (DaNa), or high {Mg2+}o all enhanced alpha-bungarotoxin (alpha-BGT) binding after 48 h of treatment . 4 . Electrical stimulation or caffeine, both affectors of SR calcium release, brought about a decrease in {AChR} probably by suppressing its synthesis . 5 . The effects of simultaneous treatment with two AChR-inducing agents, namely, high {Ca2+}o in the presence of tetrodotoxin (TTX) or high {Mg2+}o were not additive, thus suggesting action via a common saturable mediator . 6 . Intermediate AChR levels obtained following simultaneous treatments with opposing effects, e.g., electrical stimulation in the presence of high {Ca2+}o or DaNa, suggest contradictory actions on a common mediator . 7 . All these observations indicate a strong correlation between SR calcium levels and {AChR} on myotubes; while calcium accumulation in the Sr was followed by increased AChR synthesis, calcium release was accompanied by suppression of receptor synthesis.

Mikrobiologiia, 1980 May-Jun, 49(3), 408 - 11
{Exoprotease biosynthesis by an Actinomyces spheroides culture on a medium with limited carbon, nitrogen and sulfur sources}; Al'-Nuri MA et al.; The biosynthesis of exoproteases by Actinomyces spheroides 35 was studied in the conditions of limitation of the growth medium with the source of carbon, nitrogen and sulfur . The absence of these sources from the medium was found to induce the synthesis of exoproteases by the washed mycelium of Actinomyces spheroides . The enzymes were synthesized at a highest rate in the absence of nitrogen and sulfur sources . Protein as a sole source of carbon, nitrogen or sulfur in the medium suppressed rather than induced the biosynthesis of exoproteases by the culture.

Int J Radiat Biol Relat Stud Phys Chem Med, 1980 May, 37(5), 475 - 82
Increase of radiation damage to potassium-ion permeability in E . coli cells with decrease in membrane fluidity; Suzuki S et al.; Membrane lipids of an auxotroph of E . coli requiring unsaturated fatty acid were manipulated by supplementing the growth medium with unsaturated fatty acids of different chain lengths and/or configurations, and the radiation damage to K+-permeability of the resulting modified cells was investigated in relation with factors influencing membrane fluidity, such as temperature and procaine . Radiation had greater effects on membranes supplemented with unsaturated fatty acids of the trans configuration with a longer chain than on those of the cis configuration with a shorter chain . Radiation damage also increased with decrease in temperature . Furthermore, procaine-treated membranes showed increased resistance to radiation . All these results indicate that the damage was affected by the physical character of membrane lipids and that it was greater in membranes with decreased fluidity.

Nature, 1980 May 1, 285(5759), 41 - 3
Migration of capillary endothelial cells is stimulated by tumour-derived factors; Zetter BR; Angiogenesis, the growth of new blood vessels, occurs normally during osteogenesis, luteinisation and the development of the embryo, and in pathological states such as chronic inflammation, certain immune reactions and neoplasia . Furthermore, solid tumours have been reported to secrete a diffusible factor which promotes the directional growth of new capillaries towards a growing tumour . Two events required for the formation of a new capillary in response to an angiogenesis factor in vivo are the migration and subsequent proliferation of capillary endothelial cells . Progress in purifying angiogenesis factors and studying their action has been hindered, however, by the lack of quantitative in vitro assays for capillary cell migration and proliferation . Recently, we have been able to isolate clonal cell lines of bovine capillary endothelial cells that can be maintained in long-term culture using tumour-conditioned growth medium . I now report a quantitative in vitro assay for endothelial cell migration based on the phagokinetic track assay of Albrecht-Buehler . The evidence presented here demonstrates that tumour-derived factors stimulate the migration of capillary endothelial cells whereas the same factors have no effect on the migration of aortic endothelial cells.

J Cell Biol, 1980 Apr, 85(1), 70 - 82
Differential effect of thrombin on the growth of human fibroblasts; Hall WM et al.; The ability of thrombin to alter the growth of human skin fibroblasts was studied under a variety of experimental conditions . In agreement with previous reports, we obtained a moderate level of cell growth in confluent cultures using 0.5-8.0 U/ml of thrombin . In subconfluent cultures, the effect was strikingly different and was found to be dependent upon the time in culture when the enzyme was added . Cultures exposed to thrombin 24 h after subculturing showed growth stimulation several days later . In contrast, thrombin added at the time of cell plating produced a complete block of DNA synthesis and cell growth that lasted for at least 3 d . Cells exposed to thrombin under these conditions were morphologically altered and smaller . These thrombin-induced effects were reversible and could be completely prevented by pretreatment of the enzyme with hirudin before it was added to the culture medium . Growth inhibition and altered morphology were found to be the result of changes generated in the growth medium by thrombin and could be blocked by higher serum concentrations . The results of this study indicate that thrombin's influence on cell growth can be stimulatory or inhibitory and suggest that the state of the cell surface determines the response.

Tsitologiia, 1980 Apr, 22(4), 454 - 61
{Participation of the product of gene dnaG in the slow nonmutagenic repair pathway for gamma-induced single-stranded DNA breaks in Escherichia coli cells}; Khanbekian LM et al.; The survival of cells and yield of DNA single-strand breaks after the completion of DNA repair in growth medium M9 and in the buffer at 30 degrees and 43 degrees C has been investigated in four strains of alpha-irradiated Escherichia coli: PC3 dnaGts, NY73 dnaGts polA1, W3110 pol+, P3478 polA1 . The survival of the dnaGts mutant at 43 degrees in M9 and the buffer is lower and the yield of single-strand breaks is higher than at 30 degrees . In the polA1 mutant, the yield of single-strand breaks in M9 and in the buffer at both the temperatures is significantly higher, and the survival is significantly lower than in pol+- and dnaGts strains . These data indicate that dnaG gene product (primase, rifampicin-resistant RNA polymerase) and DNA polymerase I are involved in different pathways of DNA single-strand break repair . DNA polymerase I is the key enzyme for the fast, growth medium-independent repair of DNA single-strand breaks . Thus, the dnaG gene product is involved in the slow, growth medium-dependent polA-independent DNA single-strand break repair . The yield of breaks in double mutant polA1 dnaGts in M9 and in the buffer does not differ from that in single mutant polA1 . Thus, DNA polymerase I in fast repair of DNA single strand breaks is not changed by the dnaG gene product . It has been demonstrated that the dnaG gene product does not participate in the UV-induced mutagenesis.

In Vitro, 1980 Apr, 16(4), 288 - 96
Primary cultures of fetal hepatocytes from the genetically obese Zucker rat: characterization and total lipogenesis; Goldstein AL et al.; Primary fetal hepatocyte cultures derived from Zucker rats and with expected fa-gene frequencies of 0.0 and 0.75 have been established and can be used to detect early effects of the fa gene on hepatocellular metabolism . Proliferative capacity is similar in both types of culture . Changes of the growth media significantly decrease total lipogenesis in both 0.0 and 0.75 fa-gene culture grown in arginine-free DME medium . Paired incubation experiments demonstrate that total lipogenesis in 0.75 fa gene cultures is significantly less than in 0.0 fa-gene cultures under basal conditions . Stimulation of total lipogenesis by pharmacological doses of insulin and excess substrate (glucose) is significantly less in the 0.75 fa gene than in the 0.0 fa-gene cultures . These data suggest that the development of obesity in the Zucker rat cannot be attributed to elevated hepatic lipogenesis in the fetus.

Environ Health Perspect, 1980 Apr, 35, 139 - 46
Mucus and surfactant synthesis and secretion by cultured hamster respiratory cells; Baseman JB et al.; Procedures for the selective isolation and cultivation in monolayer of respiratory cells have been developed . This technique requires repeated protease treatment and gradient centrifugation of hamster tracheal or lung tissues and permits the establishment of proliferating cultures of epithelial cells with biologic specialization . Mucus synthesis was monitored in cultured tracheal cells by incorporation of 3H-labeled N-acetyl-D-galactosamine and 14C-serine into glycoprotein as determined by trichloroacetic acid precipitation of growth medium followed by acrylamide gel electrophoresis . For comparative purposes tracheal explants and several established cell lines were also examined . Synthesis and secretion of the glycoprotein macromolecule by tracheal cell monolayers appeared to be regulated by vitamin A since its addition to the culture medium significantly increased both the number of cell-associated granules and glycoprotein secretion . Lung-originated cell cultures were grown to confluence and radio-labeled with 3H-choline in serum-free medium for 24 hr to examine surfactant synthesis . Cell monolayers and growth medium were then extracted by the Folch method, and total radioactive phosphatidylcholine as well as disaturated phosphatidylcholine were determined by thin-layer chromatography and alumina gel fractionation of osmium tetroxide-reactive phospholipid, respectively . Data indicate that these cultures have a marked ability to synthesize and secrete surfactant when compared to other established cell lines . In addition, naturally transformed cells that arose during passage and senescence of the primary cultures were analyzed for their biosynthetic capabilities.

J Bacteriol, 1980 Apr, 142(1), 202 - 11
Gene expression in Escherichia coli after amino acid, purine, or pyrimidine exhaustion; Blumenthal RM et al.; Three strains of Escherichia coli B auxotrophic for leucine, guanine, or uracil were analyzed after exhaustion of the respective required nutrient from the growth medium . The pattern of transcription was analyzed by ribonucleic acid-deoxyribonucleic acid filter hybridization to specific deoxyribonucleic acid probes, and the pattern of translation was analyzed by autoradiography after the resolution of proteins on sodium dodecyl sulfate-polyacrylamide gels . The results obtained suggest the following conclusions . (i) Specific regulation of rpoBC transcription occurs at both the promoter (PL10) and the putative attenuator between rplL and rpoB . (ii) The stringent response of ribosomal protein gene expression to amino acid insufficiency is only partially mimicked by purine or pyrimidine insufficiency . (iii) Transcription initiation at PL10 decreases in response to guanine exhaustion, but in contrast increases significantly in response to uracil exhaustion . (iv) The expression of the induced lac operon is severely depressed during any of these exhaustions . These conclusions argue against simple models for regulation of ribonucleic acid polymerase production or promoter choice by the intracellular levels of its substrate nucleotides.

Endocrinology, 1980 Apr, 106(4), 1070 - 3
Microculture system for studying monolayers of functional beta-cells; Dobersen MJ et al.; A method is described for growing monolayers of newborn rat beta-cells in microculture trays . After disruption of the pancreas with collagenase, islets were isolated by Ficoll density gradient centrifugation, trypsinized to obtain individual cells, and plated in 96-well tissue culture trays . The cells were incubated for the first 3 days in growth medium containing 0.1 mM 3-isobutyl-1-methylxanthine to promote monolayer formation . The cultures could be maintained in a functional state, as defined by their responsiveness to known modulators of insulin secretion, for at least 2 weeks . As few as 1 X 10(3) islet cells/well gave results that were reproducible within +/- 10% . It is suggested that the microculture system for islet cells might prove to be a rapid and reproducible screening technique for studying drugs, viruses, or other agents that affect beta-cell function.

Biochim Biophys Acta, 1980 Mar 21, 617(3), 410 - 8
Manipulation of alkylglycerolipid levels in cultured cells . Fatty alcohol versus alkylglycerol supplements; Cabot MC et al.; Ehrlich ascites cells and monolayers of L-M cell fibroblasts were grown in medium containing either long-chain fatty alcohols or alkylglycerols . The cells were then analyzed to determine the contribution of these lipid precursors to the synthesis of complex lipids for the purpose of defining the most efficient system to elevate the levels of ether phospholipids . Label from high specific activity {1-14C}hexadecanol, {1-14C}octadecanol, and {1-14C}octadecenol was incorporated into alkyl linkages (C-18 : 1 greater than C-16 : 0 greater than C-18 : 0); however, similar labeling of acyl groups occurred . Increasing the amount of hexadecanol in the growth medium resulted in a higher percentage of 14C-labeled acyl groups than alkyl linkages at all concentrations of the alcohol supplement . Supplements of rac-{1-14C}hexadecylglycerol to the growth media were assimilated into phospholipids, which significantly increased as a function of the amounts added . Mass determinations of the alkyl ether phospholipid content in L-M cells incubated for 24 h with an alkylglycerol mixture (10.8 microgram/ml) showed an approximate 70% increase over control levels; supplementation had only a slight effect on the alk-1-enyl content . The systems described will be useful for investigating biophysical and biochemical effects of alkyl ether enrichment in membranes.

Mikrobiologiia, 1980 Mar-Apr, 49(2), 335 - 41
{Criteria of microbial succession in soil}; Kozhevin PA et al.; The dynamics of bacterial and fungal incidence in two sharply different soil samples after the addition of water and glucose was studied using direct microscopic techniques and a technique of inoculation in growth media . The ratio of the population density indices established by means of microscopy to those found from the data of inoculation characterizes the stage of microbial succession . The techniques of microscopy and inoculation are equitable, furnish different information, and must be used in parallel while examining various problems pertinent to the dynamics of the soil microflora incidence.

Am J Physiol, 1980 Mar, 238(3), R165 - 70
Reconstruction of the gill from single-cell suspensions of the eel, Anguilla japonica; Naito N et al.; Minute gill arches, about 2 mm in length and with 17-20 pairs of filaments, were reconstructed from single-cell suspensions of the adult eel gill . Mixed populations of the dissociated cells were cultured with growth medium (pH 7.2-7.4) either in a Rose chamber or in a roller tube at 20 degrees C . Single cells began to aggregate within 1 h into small cell clusters, which increase in size with selective adhesion of free cells . Melanin cells were frequently observed in the aggregates . Cell aggregates containing melanin cells transformed progressively into a gill barlike structure, and then, filamentlike structures grew from the bar . Secondary lammelalike structures were observed on the filaments . With growth of the gill bar, the filaments increased in both number and length, and after about 1-2 wk minute gill arches were reconstructed with original histological properties and circulatory system.

Mikrobiologiia, 1980 Mar-Apr, 49(2), 258 - 64
{Patterns of the directed biosynthesis of trichothecin and fibrinolytic enzymes in Trichothecium roseum}; Maksimov VN et al.; The production of the antibiotic trichothecin and of fibrinolytic enzymes was studied in the culture of the fungus Trichothecium roseum . Correlations were established between the parameters for the biosynthesis of trichothecin and fibrinolytic enzymes and the growth rate of the fungus . Thease as well as certain ratios found between potassium nitrate and ammonium chloride in the growth medium made it possible to use the function of desirability and to synthesize preferentially either trichothecin or fibrinolytic enzymes in the culture . An approach has been suggested for passing from the measurable parameters of the biosynthetic process to conventional values of desirability.

Cancer Res, 1980 Mar, 40(3), 786 - 95
Induction of two transformation-sensitive membrane polypeptides in normal rat kidney cells by iron deprivation; Fernandez-Pol JA; Rapid depletion of iron from the growth medium induces the synthesis of two membrane proteins with a subunit molecular weight of 160,000 (160K) and 130,000 (130K) in cultured normal rat kidney cells . When iron is maintained at normal levels in the growth medium of normal rat kidney cells, the synthesis of the 160K and 130K proteins is suppressed . We found that 160K and 130K are the underglycosylated forms of two membrane glycoproteins, 163K and 132K, respectively . We estimated the apparent turnover rate of the underglycosylated proteins and showed that it is slower than that of the fully glycosylated forms . Upon removal of iron from the growth medium pf simian virus 40-transformed normal rat kidney cells and Kirsten sarcoma virus-transformed normal rat kidney cells, 160K and 130K did not increase to levels comparable to those of normal rat kidney cells . Two different clones of simian virus 40-transformed normal rat kidney cells subjected to iron deprivation showed greatly reduced levels of 160K, and in one of these clones, 130K was absent . Kirsten sarcoma virus-transformed normal rat kidney cells also showed a defective response to iron deprivation manifested by reduced levels of both 160K and 130K . Additional studies indicate that these glycoproteins are membrane-associated procollagen molecules . The alteration in the coordinated induction of 160K and 130K in transformed cells suggests that these membrane-associated proteins may have an important role in transformation.

J Gen Virol, 1980 Mar, 47(1), 193 - 7
Characterization of altered BHK cells resistant to HVJ (Sendai virus) infection; Kimura Y et al.; An altered baby hamster kidney cell culture which resists the c.p.e . of HVJ (haemagglutinating virus of Japan - the Sendai strain of parainfluenza I virus) has been obtained and characterized . These cells, designated BHK-R, were originally obtained by prolonged cultivation of cells surviving HVJ infection; they have been subcultured in the presence of HVJ . No infectious virus was recovered from BHK-R cells and no evidence for the presence of HVJ antigens in the cells was demonstrated by immunofluorescent staining . When BHK-R cells were inoculated with HVJ the growth of challenge virus was suppressed and no obvious cytopathic changes were detected, while these cells normally supported the replication of mumps, influenza, Newcastle disease, vesicular stomatitis or Sindbis viruses . BHK-R cells became susceptible to HVJ infection after serial subculture in growth medium free of HVJ . It was suggested that sialic acid residues present in the surface of BHK-R cell membranes and responsible for adsorption of HVJ were split off by the action of neuraminidase of virus particles, resulting in inhibition of the attachment of challenge virus of HVJ.

Brain Res, 1980 Feb 3, 183(1), 161 - 80
Long-term survival and development of dissociated parasympathetic neurons in culture; Tuttle JB et al.; Ciliary ganglion neurons from chick embryo were grown in cell culture following enzymatic and mechanical dissociation . The culture conditions necessary for long-term (greater than 21 days) survival of the isolated neurons were investigated . Neurons could be cultured with and without non-neural cells by adjusting the culture substrate . Chick embryo extract was found to be essential in the growth medium, and the inclusion of horse serum had an additional beneficial effect . Also, non-neural cells increased the survival of these neurons in culture . Thus, there appear to be several factors involved in the survival of these neurons in culture . Assay of choline acetyltransferase activity revealed a 100-fold increase in activity over the first two weeks in vitro, and the developmental pattern of the enzyme activity was initially similar to that seen in vivo . The cultured neurons also retained many of the electrophysiological properties of ganglionic neurons in vivo . These included normal resting transmembrane potential, action potential amplitude and an afterpotential . The passive membrane properties of the cultured neurons (membrane resistance, capacitance and time constant) differed somewhat from those of neurons in ganglia . These results suggest that if proper culture conditions are provided, these parasympathetic neurons adapt well to cell culture and develop many of the properties of normal ciliary ganglion neurons in vivo.

Antimicrob Agents Chemother, 1980 Feb, 17(2), 170 - 8
Isoniazid interaction with tyrosine as a possible mode of action of the drug in mycobacteria; Herman RP et al.; The antitubercular drug isoniazid (INH) was hown by radio-chromatographic studies to react with tyrosine in growth medium . Exogenous tyrosine added to the growth medium interfered with the inhibitory action of INH on Mycobacterium phlei . These observations were confirmed by difference spectra studies which showed that tyrosine would react with INH as long as the tyrosine phenolic hydroxyl group was not blocked . These results led to the hypothesis that INH could exert its influence by interfering with tyrosine residues in mycobacterial proteins . N-acetylimidazole, a tyrosine-acetylating agent, mimicked the action of INH on the reduced nicotinamide adenine dinucleotide oxidase and dehydrogenase activity in electron transport particles from wild-type and INH-resistant M . phlei . Pyrazinamide, a drug structurally related to INH, also mimicked its effect on electron transport particles . To confirm that INH could react with tyrosine in proteins, purified enzymes with known tyrosine positions were tested . Bovine carboxypeptidase A with tyrosine at the active site was inhibited by INH and N-acetylimidazole, whereas the controls, yeast alcohol dehydrogenase and ribonuclease A, were not . It is therefore proposed that tyrosine residues in proteins may serve as the target for INH action in mycobacteria.

Arch Microbiol, 1980 Feb, 124(2-3), 249 - 54
Requirement for calcium ions in mycobacteriophage I3 DNA injection and propagation; Nagaraja V et al.; Ca2+ ions are absolutely necessary for the propagation of mycobacteriophage I3 in synthetic medium . These ions are required for successful infection of the host and during the entire span of the intracellular development of the phage . A direct assay of the phage DNA injection using 32{P} labelled phage, shows that Ca2+ ions are necessary for the injection process . The injection itself is a slow process and takes 15 min to complete at 37 degrees C . The bacteria infected in presence of Ca2+ tend to abort if the ions are subsequently withdrawn from the growth medium . The effect of calcium withdrawal is maximally felt during the early part of the latent period; however, later supplementation of Ca2+ ions salvage phage production and the mature phage progeny appear after a delayed interval, proportional to the time of addition of Ca2+.

Invest Ophthalmol Vis Sci, 1980 Feb, 19(2), 192 - 202
Procollagen and collagen produced by normal bovine corneal stroma fibroblasts in cell culture; Church RL; Procollagen and collagen were isolated from the culture medium of normal bovine corneal stromal fibroblasts . DEAE-cellulose chromatography was used to separate the collagen molecules from the different procollagens present . One collagen and four procollagen peaks were isolated and biochemically characterized . All the procollagen fractions and the collagen fraction yielded, after limited pepsin or chymotrypsin digestion followed by CNBr digestion, molecules that correspond to (alpha 1)2 alpha 2 exclusively . Thus only type I collagen is found in the growth medium of of bovine corneal stromal fibroblast cultures . Each of the individual procollagen peaks contained pro-alpha chains having molecular weights of 120,000, 150,000, 165,000, 180,000, and 190,000 daltons, according to their elution position on DEAE-cellulose . The presence of type I procollagen molecules having pro-alpha chains of 165,000, 180,000, and 190,000 daltons has not previously been reported and probably represents higher-molecular-weight precursor intermediates . The amino acid compositions of the different procollagen fractions are unique, and each contains relatively large amounts of cysteine and tryptophan . Carbohydrate analysis, cyanogen bromide peptide analysis, electron microscopy of SLS-crystallities, and SDS-polyacrylamide gel electrophoresis were used to further characterize the procollagen and collagen molecules.

Appl Environ Microbiol, 1980 Feb, 39(2), 372 - 5
Applications of fluorophore-containing microbial growth media; Ramsey WS et al.; Media containing the fluorogenic compound 8-anilino-1-naphthalene sulfonic acid may be used to discriminate between gram-positive and gram-negative bacteria and to differentiate between various species of bacteria . Fluorescent light emitted from colonies of gram-negative bacteria on 8-anilino-1-naphthalene sulfonic acid-containing agar was visually more intense than that on gram-positive bacteria . The emitted light from the gram-negative bacteria differed in wave-lengths from that of light emitted by colonies of gram-positive bacteria . The fluorescent intensity of colonies on complete 8-anilino-1-napthalene sulfonic acid agar supplemented with 1% of single substrates varied depending on the bacterial species, thus allowing the development of profiles used to identify 12 different species.

J Embryol Exp Morphol, 1980 Feb, 55, 17 - 31
Translocation of neural crest cells within a hydrated collagen lattice; Davis EM; Chick neural tubes were cultured either on planar substrata of collagen-coated Falcon plastic in growth medium with serum or within a hydrated collagen lattice (HCL) in growth medium either with or without serum . Using time-lapse cinemicrography, neural crest cells were observed emigrating from neural tubes over the collagen substrata . Once separated from the neural tube, they seldom reunite with it . Though the average rate at which the neural crest cells translocate was the same in the different culture conditions, approximately 1.0 micron/min, distinct differences in morphology and mode of translocation were observed . Neural crest cells on collagen-coated culture dishes have a flattened fibroblastic morphology and mode of translocation; in an HCl with serum, they have a bipolar shape and translocate by advancing a long, narrow leading protrusion and by periodically retracting the attenuated trailing portion of the cell; and in a serum-free HCL, they have a unipolar shape and translocate by advancing a long, narrow, branched leading protrusion and by periodically transferring the cytoplasm of the large, rounded trailing cell body forward, past a bulbous structure, and into the leading protrusion.

Arch Microbiol, 1980 Feb, 124(2-3), 285 - 7
Effect of growth temperature upon heat sensitivity in Saccharomyces cerevisiae; Walton EF et al.; The resistance of exponentially growing yeast cells to killing by exposure to 52 degrees C increase markedly as the growth temperature was increased . Identical killing curves were obtained for cells suspended in growth medium or in 0.9% saline . Cells resistant to killing at 52 degrees C were quite sensitive to killing at slightly higher temperatures . These results suggest a primary role for membrane damage in the mechanism of heat killing.

J Gen Virol, 1980 Feb, 46(2), 363 - 72
Aminopeptidase activity associated with purified murine leukaemia viruses; Yoshinaka Y et al.; Rauscher leukaemia virus (RLV), extensively purified by rate and density gradient centrifugation procedures, exhibited aminopeptidase (AP) activity . The amount of virion activity was about 0.005 times the specific activity of purified hog kidney aminopeptidase M (EC 3.4.11.2) . The activity was also found in purified Moloney and Gross leukaemia viruses, but not in comparable gradient fractions of uninfected JLSV-9 cells . Furthermore replacement of serum by bovine serum albumin in the growth medium of virus producing cells did not affect the AP activity . These results indicate that murine leukaemia viruses (MuLV) possess a tightly bound AP activity which is present as a minor component of the virion preparation and is probably not due to host cell membrane or serum contamination . Characterization of the pH optimum and substrate specificity show the MuLV aminopeptidase activity is similar to but different from both hog kidney, leucine aminopeptidase (EC 3.4.11.1) and aminopeptidase M (EC 3.4.11.2).

J Hyg (Lond), 1980 Feb, 84(1), 29 - 36
Studies on the effect of growth medium composition on the antigenicity of Mycoplasma bovis; Thorns CJ et al.; Serum proteins adsorbed from the culture medium were detected in Mycoplasma bovis antigens, the number and type of proteins depending on the serum used in the medium . The alpha-globulins cross-reacted with alpha-globulins from different types of sera but the gamma-globulins did not . The removal of non-specific medium antibodies by absorption showed that they affected the gel diffusion and growth precipitation tests, producing cross-reactions between M . bovis and M . bovigenitalium, but that the complement fixation, tube agglutination, and growth inhibition tests were not similarly affected . The presence of serum proteins in the antigens changed their specific reactivity in all the tests . The production of antibodies to serum proteins was increased by the use of an adjuvant, but it appeared that production of specific antibodies to the mycoplasmas was not.

Int J Radiat Biol Relat Stud Phys Chem Med, 1980 Feb, 37(2), 119 - 33
Induction and repair of double- and single-strand DNA breaks in bacteriophage lambda superinfecting Escherichia coli; Boye E et al.; Induction and repair of double- and single-strand DNA breaks have been measured after decays of 125I and 3H incorporated into the DNA and after external irradiation with 4 MeV electrons . For the decay experiments, cells of wild type Escherichia coli K-12 were superinfected with bacteriophage lambda DNA labelled with 5'-(125I)iodo-2'-deoxyuridine or with (methyl-3H)thymidine and frozen in liquid nitrogen . Aliquots were thawed at intervals and lysed at neutral pH, and the phage DNA was assayed for double- and single-strand breakage by neutral sucrose gradient centrifugation . The gradients used allowed measurements of both kinds of breaks in the same gradient . Decays of 125I induced 0.39 single-strand breaks per double-strand break . No repair of either break type could be detected . Each 3H disintegration caused 0.20 single-strand breaks and very few double-strand breaks . The single-strand breaks were rapidly rejoined after the cells were thawed . For irradiation with 4 MeV electrons, cells of wild type E . coli K-12 were superinfected with phage lambda and suspended in growth medium . Irradiation induced 42 single-strand breaks per double-strand break . The rates of break induction were 6.75 x 10(-14) (double-strand breaks) and 2.82 x 10(-12) (single-strand breaks) per rad and per dalton . The single-strand breaks were rapidly repaired upon incubation whereas the double-strand breaks seemed to remain unrepaired . It is concluded that double-strand breaks in superinfecting bacteriophage lambda DNA are repaired to a very small extent, if at all.

Biochim Biophys Acta, 1980 Jan 25, 595(2), 189 - 99
Control of membrane polar lipid composition in Acholeplasma laidlawii a by the extent of saturated fatty acid synthesis; Christiansson A et al.; The low level of endogenous fatty acid synthesis in Acholeplasma laidlawii A strain EF22 was found to be caused by a deficiency of pantetheine in the lipid-depleted growth medium . By supplementing the oleic acid-containing medium with increasing concentrations of pantethein, saturated fatty acid synthesis was stimulated (having an apparent Km of 5 microM for pantetheine) and the incorporation of endogenously synthesized fatty acids in membrane lipids increased markedly . Furthermore, carotenoid biosynthesis was stimulated . Exogenous palmitic acid was found to inhibit partially the endogenous fatty acid synthesis . A gradual stimulation of fatty acid synthesis was accompanied by a linear increase in the molar proportion between the two dominating membrane glucolipids, monoglucosyldiacylglycerol and diglucosyldiacylglycerol . The total amount of charged membrane lipids decreased upon increasing the degree of fatty acid saturation . These regulations are discussed in terms of membrane stability, and influence of membrane molecular ordering and surface charge density on lipid polar head group synthesis.

Artery, 1980, 6(6), 437 - 57
Oral contraceptive steroids and atherosclerosis: lipogenesis in human arterial smooth muscle cells and dermal fibroblasts in presence of lipoprotein-deficient serum from oral contraceptive users; Subbaiah PV et al.; PIP: Oral contraceptives users and control women sera were studied to determine the degree of incorporation of tritiated acetate into lipids by cultured human arterial smooth muscle cells and dermal fibroblasts . When the cells were exposed in vitro to oral contraceptive users serum, they showed a significant increase in synthesis of cholesterol above controls in both fibroblasts (increase of 25-26%) and smooth muscle cells (plus 28-42%) but no significant concomitant change in lecithin synthesis; this exposure also resulted in increased cholesterol/lecithin ratios in newly synthesized lipids much higher than controls in both fibroblasts (plus 50%) and smooth muscles (plus 30%) . However, when steroid ingredients of the oral contraceptives were added in vitro to the growth medium containing control sera, no stimulation of lipogenesis by cells occurred . Tritiated mevalonate incorporation into cholesterol was no different with oral contraceptive user or control serum, indicating that the stimulation noted in cholesterol synthesis by oral contraceptive users serum was caused by an increased 3-hydroxy-3-methyl glutaryl-CoA reductase activity . This study suggests that increased sterol synthesis in arterial smooth muscle cells of oral contraceptive users may be a pathogenetic factor that affects the risk of atherosclerosis .

Physiol Chem Phys, 1980, 12(1), 63 - 7
Effect of proline on synthesis of collagen by cells in culture; Baich A et al.; Chick embryo cells, human epithelial cells, and mouse cells in culture were tested for response to elevated levels of proline in the growth medium . In none of the cell lines tested was the amount of collagen formed stimulated by addition of excess proline.

Horm Res, 1980, 12(6), 324 - 32
Progesterone and estrogen receptors in GH3 cells; Roos W et al.; GH3 cells are shown to contain cytosolic progesterone receptor in a mean concentration of 270 fmol/mg protein with a dissociation constant (K(D)) for promegestone of 3 times 10(-9) M (4 degrees C) . Estrogen receptor (K(D) = 1.8 times 10(-10) M) is demonstrated in cytosol (121 fmol/mg protein) as well as in 0.4 M KCl extracts (89 fmol/mg protein) of crude nuclear fractions . No progesterone receptor was detectable in the nuclear fraction . Both receptors are characterized by isoelectric focussing and by competition experiments . Addition of 10(-8) M estradiol to the growth medium increases progesterone receptor levels up to five, suggesting that the progesterone receptor is under estrogenic control.

Arch Virol, 1980, 64(4), 359 - 74
The association of calf serum with the contamination of BHK21 clone 13 suspension cells by a parvovirus serologically related to the minute virus of mice (MVM); Nettleton PF et al.; An investigation of persistent cell deaths of BHK21 suspension cells during subculturing resulted in the isolation of a viral agent . The agent was isolated from samples of dead cells, cell growth media and 2 batches of calf serum . It was established that the agent was associated with the use of certain batches of calf serum . The isolated virus was found to replicate effectively only in rapidly growing BHK cell cultures . In monolayers it caused the formation of large intranuclear inclusion bodies . The isolate was a strong haemagglutinin; it was stable to ether, chloroform, pH 3 and heating at 56 degrees C . It was shown to be a DNA-virus and by electron-microscopy it was evident as unenveloped spherical, small particles (21 nm in diameter) . In sucrose density gradients the virus sedimented as 2 peaks at approximately 114S and 85-92S, and by caesium chloride equilibrium centrifugation, two main peaks at densities of 1.39 and 1.31 g/ml were evident with a minor peak at 1.35 . The capsid of the complete virion consisted of 3 polypeptides . The agent has therefore been provisionally designated a member of the Parvoviridae family, genus parvovirus . Serologically it was found to be related to MVM . The isolated parvovirus was inactivated by 0.05 per cent peracetic acid and 0.05 per cent acetylethyleneimine.

Biochim Biophys Acta, 1980, 613(1), 191 - 202
Purification and characterization of a nicotinamide deamidase released into the growth medium of neuroblastoma in vitro; Wintzerith M et al.; Nicotinamide deamidase (nicotinamide amidohydrolase, EC 3.5.1.19) has been demonstrated in the conditioned growth medium of the M1 clonal cell line of mouse C1300 neuroblastoma . The enzyme has been purified 1200-1500-fold by Sephadex G25, hydroxyapatite, DEAE-cellulose, Sephadex G200 and NAD-Sepharose column chromatographies . The purified protein was characterized by polyacrylamide gel electrophoresis under non-denaturing and denaturing conditions . The apparent molecular weight has been estimated to be 230,000, and the subunits had respective molecular weights of 65,000 and 50,000 . Histidine was the only NH2-terminal amino acid found . The enzyme is a glycoprotein; mannose and N-acetyl-glucosamine have been identified . The effects of various ions on its activity have been investigated . The enzyme has a Km for nicotinamide in the order of 10(-6) M, a pH optimum of 7.2 and a pHi of 5.4 . It is inhibited by heating and by sulfhydryl reagents . The existence of a nicotinamide deamidase with a high affinity for nicotinamide favors the operation of the Preiss-Handler pathway in M1 cells cultured in vitro . We found an induction of nicotinamide deamidase and a cellular increase of NAD with a higher nicotinamide supply and a repression of the released enzyme with supplying NAD in the nutrition medium of M1 cell cultures.

In Vitro, 1980 Jan, 16(1), 31 - 7
A tissue model for the study of cell proliferation in vitro; Norrby K et al.; A procedure for the cultivation of mesentery is described, in which the culture is fully representative of the tissue of origin . The intact mesenteric membrane -- exposed to a minimum of trauma -- was spread out over a hole in a filter paper strip in fluid medium and was cultivated free-hanging . Specimens from rats and guinea pigs were used . The organ culture model appears especially apt for cytochemical and proliferation studies . Proliferation variables based on Feulgen DNA analysis in individual, morphologically defined cells and on mitotic counting and radiochemical analysis were estimated . The tissue was fully viable in chemically defined growth medium and showed an almost unaltered light microscopical appearance after up to 52 hr in culture.

Microbiol Immunol, 1980, 24(1), 31 - 7
Interferon production in human diploid cell strains derived from embryonic lungs; Machida H et al.; Twenty-three strains of human diploid cells derived from embryonic lungs were tested for production of interferon by "superinduction." Strain HAIN-55 produced a relatively high level of interferon . The optimal concentration of cycloheximide for superinduction was essentially equal to that reported with foreskin fibroblasts . On the other hand, actinomycin D at a concentration of 4 to 16 microgram/ml enhanced the production of interferon more strikingly than at a concentration of 1 microgram/ml, which was usually employed for superinduction in the foreskin fibroblasts . Inhibition of interferon production was observed when fetal bovine serum was added to the medium during treatment with metabolic inhibitors for superinduction . Minimal essential medium was superior to Eagle's basal medium as growth medium for interferon production, and serum added after removal of metabolic inhibitors could be replaced by bovine serum albumin . The yield of interferon produced under the best conditions in this study, with strain HAIN-55, was more than 10,000 reference units/ml.

J Bacteriol, 1980 Jan, 141(1), 350 - 8
Amphotericin B-induced changes in K+ content, viability, and ultrastructure of yeast-phase Histoplasma capsulatum; Arnold WN et al.; Yeast-phase cells of Histoplasma capsulatum were challenged with amphotericin B, and membrane perturbation was monitored by K+ efflux . Suspensions of washed cells readily absorbed about 1.12 microgram of amphotericin B per mg (dry weight) and further nonspecific sites were also apparent . The dose-response curve for initial rate of K+ efflux was sigmoidal within the range 0.1 to 1.0 microgram of amphotericin B per ml . A fungistatic concentration of amphotericin B (0.3 microgram/ml) evoked an efflux of 85 to 90% K+ from the cells within 15 min, but cell viability decreased only 13% (yeast phase) or 33% (transformed to mycelial units) . Ultrastructural changes in treated cells were detected within 5 min, and the hallmark was expansion of vacuoles during the 1-h monitoring period . In contradistinction to a previous report, the appearance of the protoplasmic membrane was not altered by fungistatic concentration . When treated cells were returned to a fresh growth medium, there was a pronounced lag (20 h) . During this apparent recovery phase, the large vacuoles fragmented and returned to normal size . It is proposed that vacuoles of H . capsulatum act as a spatial buffer of considerable survival value to stressed cells.

Biochim Biophys Acta, 1980, 606(1), 105 - 12
Loss of nuclear photoreactivating enzyme following ultraviolet irradiation of Chlamydomonas; Small GD; UVS1 is a mutant of Chlamydomonas reinhardi defective in the dark repair of pyrimidine dimers in nuclear DNA . All of the pyrimidine dimers in nuclear DNA can be repaired upon exposure to photoreactivating light immediately after irradiation . However, none of the dimers in nuclear DNA are repaired by photoreactivation if the irradiated cells are incubated in the dark for 24 h in growth medium . Pyrimidine dimers in chloroplast DNA that are unrepaired during the 24 h post-irradiation incubation can be repaired by photoreactivation . Treatment with methyl methanesulfonate to give a similar survival as the fluence of ultraviolet light did not lead to the inactivation of nuclear photoreactivating enzyme after 24 h in the dark . Assay for photoreactivating enzyme in cell-free extracts showed that about 80% of the photoreactivating enzyme activity disappears after incubating ultraviolet-irradiated cells in the dark for 24 h.

Artery, 1980, 7(1), 16 - 31
Inhibition of collagen and non-collagen protein synthesis in cultured aortic smooth muscle cells by hyperlipoproteinemic serum; Holderbaum D et al.; The response of rabbit arterial tissue to prolonged extreme hyperlipoproteinemia is the development of a state of enhanced in vitro protein synthesis where severe atherosclerosis is present . In order to examine this stimulation, arterial cells were grown in cell culture and exposed to hyperlipemic serum as a part of the growth medium . Rather than observing an overall increase in protein synthesis as seen in atherosclerotic tissue, the cells exposed to hyperlipemic serum produced less collagen and non-collagen protein per cell than counterparts exposed to normolipemic homologous serum . We conclude that exposure to hyperlipemic serum for six days in vitro causes enhanced cellular proliferation, as seen by others, and a general perturbation of protein synthesis which may be related to cell population density.

Zentralbl Bakteriol A, 1980, 248(3), 392 - 401
A new anaerobic blood culture medium: laboratory evaluation; Dankert J et al.; A new medium, Diagnostic Anaerobic Growth Medium (DAG) was evolved for the culture of anaerobic bacteria from blood . Redox potential measurements of DAG, in comparison with four other commonly used media (supplemented peptone broth; thioglycollate broth; chopped meat glucose broth; tryptone soya broth) showed that Ecal of DAG medium to be lower than that of the other media tested . The new medium recovered more quickly after addition of blood, to reach "blank" values within 4 h . Growth curves of 21 anaerobic bacteria in DAG medium showed that they all multiplied satisfactorily.

Radiat Environ Biophys, 1980, 18(4), 289 - 300
60 Hz electric field parameters associated with the perturbation of a eukaryotic cell system; Miller MW et al.; Roots of Pisum sativum were exposed for seven days to 60 Hz electric fields ranging from 70--430 V/m in an aqueous medium whose conductivity was approximately 0.07 mho/m . (Corresponding current densities in the exposure medium associated with these field strengths ranged from 0.5--3.0 mA/cm2) . Control and exposed roots were grown concomitantly in the same tank whose growth medium was continuously circulated . Temperature in the exposure medium was held at a constant 19 degrees C . All experiments were conducted "double blind." Root growth rates were determined daily . No perturbations in root growth were observed with electric fields of 150 V/m; there was a slight effect at 360 V/m, and a pronounced decrease in growth rate occurred at 430 V/m . Root conductivities are comparable to that of the growth medium . Under conditions in which growth inhibition occurs, it is estimated that induced 60 Hz cell membrane potentials would be of the order of 3--8 mV.

Mol Gen Genet, 1980, 180(3), 597 - 603
Relation between liquid-holding recovery, DNA repair, and mitotic recombination in the rad3 mutant of Saccharomyces cerevisiae after treatment with diepoxybutane (DEB); Zuk J et al.; The rad3 mutant is characterized by a high level of liquid-holding recovery after DEB treatment . The recovery is abolished when the treated cells are postincubated in growth medium, but the effect can be cancelled by suppression of DNA and protein synthesis by specific inhibitors . Alkaline sucrose gradient sedimentation revealed that DEB induces single strand breaks in DNA which are not repaired during post-treatment incubation in growth medium or during LH . Effective repair takes place only when LH is followed by incubation in growth medium . Split-dose treatment applied to test the possible inducibility of repair by LH did not confirm this presumption . In a diploid homozygous for rad3 mutation, DEB induces mitotic inter- and intragenic recombination with very high frequency . Liquid-holding recovery (LHR) was found to be accompanied by an increase in molecular weight of DNA and by a sharp decrease in the frequency of mitotic recombination . The data suggest that recombination events are not involved in LHR pathway.

Mutat Res, 1980 Jan, 69(1), 43 - 50
Co-mutagenic effects of propidium on petite induction by ethidium in Saccharomyces cerevisiae; Fukunaga M et al.; Propidium, whose structure is closely related to ethidium bromide, induced a low level of petites in yeast, but only at high concentrations with long incubation time, and only in growth medium . When added to growing cells, propidium also caused a large increase in petite induction by ethidium even at submutagenic concentrations of ethidium . Incorporation of adenine into DNA was inhibited by propidium in mitochondria but not in nuclei . Propidium by itself had no effects on fragmentation of pre-existing DNA, but enhanced mitochondrial DNA degradation provoked by ethidium . The proportion of suppressive clones occurring among the petites from ethidium treatment was reduced by the presence of propidium . All of these results indicated that propidium treatment led to degradation of the mitochondrial DNA in petites induced by ethidium but not in native (intact) mitochondrial DNA, nor in spontaneous petite colonies . The results are discussed in terms of possible mechanisms of modulation of petite induction.

Mutat Res, 1980 Jan, 69(1), 19 - 41
The role of dimer excision in liquid-holding recovery of UV-irradiated haploid yeast; Ferguson LR et al.; We have directly tested the theory that liquid-holding recovery is due to an increase in the efficiency of excision-repair during holding in non-growth conditions, by assaying the dimers present in UV-irradiated cells held in saline, in growth medium or first in saline then in growth medium . We observed no differences in the amount of excision in any conditions . By assaying the kinetics of excision and comparing that with the timing of DNA synthesis, we have tested the theory that holding in growth medium allows more repair by extending the time available for it . We found that the observations were more consistent with the onset of DNA synthesis being dependent on the amount of repair rather than the converse . We have analysed the role of repair in liquid-holding recovery in a series of split-dose experiments . As Parry and Parry found, yeast cells which have been irradiated and held in non-growth conditions were much more resistant to further UV-irradiation . The increase in resistance was proportional both to the degree of fractionation of the dose and to the size of the first dose . No effect was observed if this was below 30 J . m-2 . We found that the cells were able to excise more of the dimers induced if the UV dose was fractionated . We have shown that part of this increase in efficiency of excision is due to the relief of "dimer interference" . "Dimer interference" is the name given to the inhibition of excision of a dimer by the presence of a neighbouring dimer . Most of the increase in efficiency, however, was due to the induction of more efficient excision repair per se, that is the excision of a greater fraction of the dimers present than could be excised in uninduced cells . Among the incidental observations we have made which are new and likely to be of interest are (1) that stationary phase cells showed a lag in the onset of excision, but log phase cells did not; (2) that excision was nevertheless constitutive in that it occurred in the presence of concentrations of cycloheximide inhibitory to protein synthesis and (3) that caffeine affected but did not inhibit dimer excision.

J Bacteriol, 1980 Jan, 141(1), 235 - 45
Regulation of phosphoglycerate dehydrogenase levels and effect on serine synthesis in Escherichia coli K-12; McKitrick JC et al.; The level of phosphoglycerate dehydrogenase, the first enzyme in the biosynthetic pathway to serine and glycine, has been studied in Escherichia coli grown under different conditions . The enzyme level was not reduced by growth in a medium which contained the end products of the pathway, nor was it elevated when the growth rates was limited by the supply of serine . Elevation of phosphoglycerate dehydrogenase did not occur when charging of tRNA ser was inhibited by serine hydroxamate . However, phosphoglycerate dehydrogenase levels were subject to regulation . Elevated levels of enzyme activity were observed in merodiploids containing multiple copies of the serA gene, and lowered enzyme levels were found in cells grown on carbon sources other than glucose or when certain amino acids were present in the growth medium . The combined effect of these nutritional changes (carbon source and amino acids) reduced the level of phosphoglycerate dehydrogenase to 10 to 12% of that found in wild-type cells and to about 5% of the level in the merodiploids . By using antibody prepared against purified phosphoglycerate dehydrogenase we established that the decrease in enzyme activity reflected decreased amounts of enzyme protein . Constant intracellular concentrations of 3-phosphoglycerate and serine were found in cells with marked differences in phosphoglycerate dehydrogenase activity, indicating that end product inhibition of phosphoglycerate dehydrogenase activity, rather than the amount of the biosynthetic enzymes, is the major factor in regulating the intracellular concentration of serine.

Antonie Van Leeuwenhoek, 1980, 46(2), 161 - 7
Effects of certain cadmium species on pure and litter populations of microorganisms; Lighthart B; Cadmium inhibition of microorganisms was found to be bacterial and chemical species dependent . E . coli inhibition was a function of the cadmium-ion concentration irregardless of the presence of citrate, a chelator for cadmium that it could not metabolize . Whereas with a Pseudomonas sp . able to metabolize citrate, cadmium inhibition was a function of both the cadmium ion and the presence of citrate . With no citrate, inhibition of this organism occurred only at relatively high cadmium-ion concentrations (above 10(-4) M); when citrate was added to the same cadmium-containing growth medium, inhibition was observed at a 1000 times lower cadmium-ion concentration (i.e., 10(-7) M) . This observation is contrary to the classical understanding where a chelate reduces the toxic form of a metal allowing increased growth of the organism . The species of cadmium also differentially inhibited the Douglas fir letter respiration and nitrogen-fixing community activities.

Rev Argent Microbiol, 1980 Jan-Apr, 12(1), 23 - 8
{Species of the genus Beijerinckia Derx isolated from a soil of Corrientes (Argentina)}; Amor Asuncion JM et al.; Four species of bacteria were isolated from one soil of Corrientes (Argentina) . They were first classified as Beijerinckia genus based on their cultural, morphological and physiological characteristics . The basic criteria were: growth media without combined nitrogen and fixation of high amounts of gaseous nitrogen (as determined by the acetylene-ethylene assay) . Further studies permitted the classification of these microorganisms in the following species: Beijerinckia indica, Beijerinckia fluminensis, Beijerinkia mobilis and Beijerinckia derxii.

Z Allg Mikrobiol, 1980, 20(4), 271 - 81
Factors which influence CaC12 dependent transfection of lambda DNA in Escherichia coli K12 recipients; Pefeifer M et al.; This paper presents further parameters influencing the competence, the process of DNA uptake and the efficiency of plating of CaC12-treated E . coli D12 strains . We have found that the process of DNA uptake depends not only on the treatment of bacteria with a certain CaCl2-concentration but is also influenced considerably by a shift-down of the CaCl2-concentration in the reaction mixture . The pH of the growth media and of the reaction mixture plays an important role in maintaining of optimal transfection . The efficiency of plating is influenced by the thickness of the top layer and the concentration of bacteria on the plate . Without genetic variation of the strains, by only varying the mentioned factors we could improve the efficiency of CaCl2 transfection at about two orders of magnitude to a maximum of 6 X 105 pfu/microgram DNA.

J Bacteriol, 1980 Jan, 141(1), 227 - 34
Control of arginine metabolism in Neurospora: flux through the biosynthetic pathway; Goodman I et al.; The flux into the arginine biosynthetic pathway of Neurospora crassa was investigated using a mutant strain lacking the ornithine-degrading enzyme ornithine aminotransferase (EC 2.6.1.13) . Flux was measured by the increase in the sum of the radioactivity (derived from {14C}glutamic acid) in the ornithine pool, the arginine pool, and arginine incorporated into proteins . Complete cessation of flux occurred immediately upon the addition of arginine to the growth medium . This response occurred prior to expansion of the arginine pool . After short-term exposure to arginine (80 min), flux resumed quickly upon exhaustion of arginine from the medium . This took place despite the presence of an expanded arginine pool . Initiation of flux required approximately 80 min when the mycelia were grown in arginine-supplemented medium for several generations before exhaustion of the exogenous arginine . The arginine pool of such mycelia was similar to that found in mycelia exposed to exogenous arginine for only 80 min . The results are consistent with rapid onset and release of feedback inhibiton of arginine biosynthesis in response to brief exposure to exogenous arginine . The insensitivity of flux to the size of the arginine pool is consistent with a role for compartmentation in this regulatory process . The lag in initiation of flux after long-term growth in the presence of exogenous arginine suggests the existence of an additional regulatory mechanism(s) . Several possibilities are discussed.

J Supramol Struct, 1980, 13(2), 175 - 82
Release of adenosine by C1300 neuroblastoma cells in tissue culture; Green RD; Previous work in our laboratory led us to postulate that N2a cells release adenosine into growth medium, where it acts at the extracellular adenosine receptors to modulate the sensitivity of the cells to the cyclic AMP-elevating effect of adenosine {Green, RD, J Pharmacol Exp Ther 201:610, 1977} . We have now devised a high-performance liquid chromatographic (HPLC) procedure capable of quantitating the concentrations of adenosine in cells and tissue culture media . Growth media of N2a cells and a variant of N2a cells deficient in hypoxanthine-guanine phosphoribosyltransferase (HGPRT-) contain 10-20 nM adenosine, while that of a variant deficient in adenosine kinase (AK-) is elevated severalfold . It appears that the concentration of adenosine in growth media is determined by both the rate at which it is released by cells into the medium and the rate at which it is metabolized by adenosine deaminase present in the serum in the growth medium . Both N2a and AK- cells release considerable amounts of adenosine into serum-free medium (SFM) over a short period . Adenosine release is greater from AK- cells and is accelerated by erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA), a potent adenosine deaminase inhibitor . This accelerated release is retarded by dipyridamole and homocysteine . Surprisingly, dipyridamole and 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (Ro 20 1724), a potent phosphodiesterase inhibitor, stimulate basal adenosine release from N2a but not from AK- cells . It remains to be determined if this is due to an effect of these compounds on adenosine kinase . These results give further support for the hypothesis that adenosine in growth medium modulates the sensitivity of the cells to the cyclic AMP-elevating affect of adenosine, and furthermore they suggest that adenosine in growth media may tonically stimulate adenylate cyclase and affect processes controlled by the cyclic AMP:cyclic AMP-dependent protein kinase system.

Adv Biochem Psychopharmacol, 1980, 22, 291 - 304
Enkephalin inhibition of adenylate cyclase activity in neuroblastoma N18TG2 cells: effect of sulfatide incorporation; Law PY et al.; Potentiation of the morphine inhibition of adenylate cyclase activity in the neuroblastoma N18TG2 cell line after sulfatide incorporation has been observed previously and a similar sulfatide effect on enkephalin inhibition was investigated . Conditions for the sulfatide incorporation were defined . Though N18TG2 cells possess a high affinity for 3H-D-Ala2-Met5 enkephalinamide (14 nM), the met5-enkephalin was relatively inactive in inhibiting the intracellular cAMP level . The IC50 value for met5-enkephalin to inhibit the prostaglandin E1 (PGE1) stimulated cAMP product was 65 nM in N18TG2 as compared to the IC50 value of 3 nM in the neuroblastoma x glioma NG108-15 hybrid . Upon exposure of the N18TG2 cells to 22 microM of sulfatide in the growth medium for 6 hours, both the potency and efficacy of the met5-enkephalin in sulfatide treated cells were found to be 3.4 nM . The degree of potentiation was dependent on the quantity of CS added to the growth medium and the age of the culture . Furthermore, the sulfatide effect appeared to be at the adenylate cyclase complex . Basal and PGE1-stimulated adenylate cyclase activity in the cellular membrane was similarly affected by enkephalin after sulfatide incorporation . There was a lipid specificity, since in the group of lipids tested only CS and PC exhibited the potentiation effect.

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