FEBS Lett, 1985 Oct 7, 190(1), 157 - 60 Biosynthesis of abnormally glycosylated hepatoma secretory proteins in cell cultures; Alm R et al.; We studied, by electrophoretic techniques, the physiochemical properties of 4 glycoproteins, alpha 1-antitrypsin, alpha 1-antichymotrypsin, alpha 1-acid glycoprotein and transferrin synthesized by three different human hepatoma cell lines . A common feature was the export of glycoproteins with retarded electrophoretic mobility, indicating incomplete sialylation, and a predominance of atypical, highly branched carbohydrate chains . The abnormal glycosylation pattern may be specific for malignant transformation of hepatocytes and possibly related to the intracellular accumulation of some of these proteins in malignant cells. Brain Res, 1985 Oct 7, 344(2), 369 - 72 Effects of norepinephrine on rat neocortical neurons in dissociated cell culture; Rosenberg PA et al.; Intracellular recordings were made from neurons in dissociated cell culture of neocortex during application of norepinephrine (NE) or other adrenergic agonists . In the population of neurons generally studied, greater than 18 micron in diameter, adrenergic agonists from 1 nM to 50 microM produced no change in membrane potential or input resistance 120 cells) . Adrenergic agonists increased synaptic activity impinging on the impaled cell in 25/120 neurons (21%) . In neurons in cocultures of locus coeruleus and cerebral cortex, again the same synaptic response to perfusion with NE was noted in 13/93 neurons (14%) . In addition, direct effects of NE were noted on 6/93 neurons recorded from in cocultures, all close to the explant . In these cells, NE hyperpolarized the membrane in association with a small decrease in input resistance (11%) . These responsive cells may have originated within the explant . A paradigm was used for testing the possibility of a responsive element in the cultures distinct from the impaled soma . 'Hot spots' were found using concentrations of isoproterenol as low as 10 nM. J Neurosci Methods, 1985 Oct-Nov, 15(2), 101 - 12 Glycogen accumulation in rat cerebral cortex in dissociated cell culture; Rosenberg PA et al.; Incorporation of {3H}glucose into {3H}glycogen was demonstrated in individual coverslip cultures of dissociated rat cerebral cortex . The time course of incorporation of {3H} glucose into {3H}glycogen was investigated, and it was found that {3H}glycogen accumulation monotonically increased during at least the first hour of incubation with {3H}glucose . In order to identify which cells accumulate glycogen in these cultures we attempted to demonstrate cytochemically the localization of glycogen . We found, however, that conventional aqueous methods of glycogen cytochemistry did not reliably or consistently stain the cultures . By labelling glycogen using {3H}glucose, we were able to show that the entire {3H}glycogen compartment was extractable by water after ethanol fixation . Therefore we developed a non-aqueous technique which preserves tissue glycogen by exploiting its solubility properties . Using this technique we were able to cytochemically demonstrate glycogen in at least two different cell types in the cultures. J Biol Stand, 1985 Oct, 13(4), 303 - 8 A simple endpoint dilution method for evaluating serum used for cell culture; Jiang SD et al.; A simple endpoint dilution method for evaluating foetal calf serum quality is described . The test uses a series of doubling dilutions of cells on microtitre trays with the test sera added to replicate dilution series . After five to six days of incubation the cells are stained with crystal violet and the end points read macroscopically . The cell growth-promoting property of serum may be expressed as a reciprocal of the cell dilution resulting in an approximately 50% coverage of cells. J Invest Dermatol, 1985 Oct, 85(4), 381 - 4 Dermatofibrosarcoma protuberans: altered collagen metabolism in cell culture; Albini A et al.; Dermatofibrosarcoma protuberans is a low-grade malignant tumor that grows invasively but rarely forms metastases . Its origin is still controversial . We characterized the synthesis of collagen in detail in cells which were obtained from dermatofibrosarcoma protuberans tumors by enzymatic tissue disintegration . Similar to fibroblasts, all tumor cell strains produced considerable amounts of collagen . However, the rate was reduced compared to normal skin fibroblasts . Cells grown from the tumors synthesized type I collagen, but no type II could be detected . After serial passaging the cultures started to produce type III collagen, which is probably due to a slow overgrowth by normal fibroblasts. Br J Haematol, 1985 Oct, 61(2), 293 - 302 In vitro erythropoietin assay based on erythroid colony formation in fetal mouse liver cell culture; Sakata S et al.; Critical studies were made on erythroid colony formation from cultured fetal mouse liver cells in an attempt to develop a simple and sensitive erythropoietin (Epo) assay procedure . The maximum colony formation was observed 24 h after plating of the cells when an evident dose-response relation was found for Epo added . The colony forming ability decreased steadily as the gestational age of the fetus advanced and was gradually lost by postnatal days 10-11 . By morphological and cytochemical criteria almost all the colonies were found to be erythroid . 59Fe-labelling experiments revealed a fairly good correlation between the colony number and 59Fe incorporation into both cells and haem . Dose-response curves for plasma were parallel to the Epo standard curve . Based on these findings we developed a procedure which could measure as little as 0.4 mU of Epo without requiring 59Fe . Using this method, plasma Epo titres were determined in 16 normal and 69 anaemic subjects. Endocrinology, 1985 Oct, 117(4), 1521 - 9 Identification of androgen-binding protein from testis cytosol and sertoli cell culture medium of the cynomolgus monkey, Macaca fascicularis; Keeping HS et al.; The biochemical and immunological properties of monkey androgen-binding protein (mABP) from the cynomolgus monkey, Macaca fascicularis, have been examined in testis cytosol and medium of primary Sertoli cell-enriched cultures . mABP in testis tissue was separated from the serum protein testosterone-estradiol-binding globulin (mTeBG) by affinity chromatography on Concanavalin A-Sepharose (Con A) . mTeBG from serum extracts was completely retained by the lectin and could be displaced with buffer containing alpha-methyl-D-glucoside . In contrast, mABP from testis extracts either did not interact with the Con A and appeared in the void volume or was partially retained by the column and could be eluted with buffer alone . A third component retained by the Con A may represent mTeBG contamination, a form of mABP which binds to Con A, or both . The specific 5 alpha-dihydrotestosterone (DHT)-binding activity in the void volume of the Con A column was designated mABP and was further studied . {3H}DHT binding to mABP was saturable and of limited capacity (0.163 +/- 19 pmol/mg protein) . Scatchard analysis of the data was consistent with a single class of binding sites with an apparent dissociation constant (Kd) at 4 C of 2.6 +/- 0.2 X 10(-9) M . DHT was the most effective competitor of {3H}DHT binding to mABP, followed by 2-methoxyestradiol, testosterone, estradiol, and cyproterone acetate . Concentrated Sertoli cell culture medium subjected to steady state polyacrylamide gel electrophoresis produced a single peak of specifically bound {3H}DHT with a mobility similar to that of other androgen-binding proteins . {35S}Methionine-labeled medium proteins were immunoprecipitated with a rabbit anti-human TeBG antiserum . Two bands, corresponding to mol wt of approximately 46,000 and 48,000, were observed by fluorography, with the lighter component being more intense . After androgen affinity chromatography of radiolabeled medium proteins, these two bands were again observed on sodium dodecyl sulfate-urea-containing polyacrylamide gels . These results demonstrate that 1) mABP may be separated from mTeBG by lectin affinity chromatography, as in humans; 2) hTeBG and mABP are antigenically related; and 3) mABP consists of subunits of different mol wt in unequal ratios. J Neurochem, 1985 Oct, 45(4), 1254 - 61 Acetylcholine synthesis by adult bovine adrenal chromaffin cell cultures; Boksa P; Adrenal chromaffin cells normally synthesize and release catecholamines . In the present study, {3H}acetylcholine synthesis and another characteristic of cholinergic neurons, {3H}choline uptake, were studied in cultures of adult bovine adrenal chromaffin cells . Chromaffin cell cultures took up {3H}choline from the medium and acetylated the {3H}choline to form {3H}acetylcholine . The rate of {3H}acetylcholine synthesis increased after 19 days in culture and continued to increase up to 28 days in culture . {3H}Acetylcholine synthesis could be increased by stimulating the cells with a depolarizing concentration of K+ . The ability for K+ to stimulate synthesis of {3H}acetylcholine developed only after 28 days in culture . {3H}Choline was taken up by the cultures through a single mechanism with a high (to intermediate) affinity for choline . {3H}Choline uptake was enhanced by Na+ omission in day-14 cultures, but was at least partially Na+-dependent in day-29 cultures . Hemicholinium-3 (IC50 less than 10 muM) inhibited {3H}choline uptake into chromaffin cell cultures . It is concluded that bovine adrenal chromaffin cells, maintained in culture, are able to exhibit cholinergic properties and this capacity is retained even by the mature adult cell. Zh Mikrobiol Epidemiol Immunobiol, 1985 Oct, (10), 32 - 5 {Effect of proteolytic enzymes on the enhanced sensitivity of a cell culture of goat embryonic kidney to Newcastle disease virus}; Korsun LL et al.; The influence of proteolytic enzymes on the increase of the sensitivity of cell cultures to Newcastle disease virus was tested in the continuous culture of goat embryo kidney cells . In this investigation the dose of trypsin and the time of incubation for the treatment of the cell culture was selected . The preliminary treatment of the monolayer culture of goat embryo kidney cells with trypsin by incubation at a temperature of 18-20 degrees C for 30 minutes made it possible to increase the sensitivity of the culture to Newcastle disease virus 10-fold. J Histochem Cytochem, 1985 Oct, 33(10), 1087 - 9 Identification of gastrin-producing cells in cell cultures and smears: an avidin-biotin-peroxidase complex (ABC) technique; Gower WR Jr et al.; A method is described that uses the avidin-biotin complex (ABC) immunoperoxidase technique to provide a rapid, sensitive, and specific means to quantitate isolated G cells in cultures and suspensions. Antimicrob Agents Chemother, 1985 Oct, 28(4), 552 - 60 Antiviral activity of 5-ethyl-2'-deoxyuridine against herpes simplex viruses in cell culture, mice, and guinea pigs; Schinazi RF et al.; The susceptibility of 3 laboratory strains and 24 clinical isolates of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) to 5-ethyl-2'-deoxyuridine was determined in plaque reduction assays in Vero cells . The median effective doses were 8.6 and 7.8 microM, respectively . The drug was less potent than acyclovir and other related antiviral drugs, but it had a high therapeutic index against both HSV-1 and HSV-2 . Drug-resistant viruses were readily produced in cell culture . These variants were cross-resistant to acyclovir, 2'-fluoro-5-iodoaracytosine, and 2'-fluoro-5-methylarauracil but were susceptible to vidarabine or phosphonoformate . These findings confirm that the selective antiviral activity of 5-ethyl-2'-deoxyuridine is mediated by the virus-induced thymidine kinase . Oral or intraperitoneal administration of the drug at nontoxic doses was ineffective in protecting mice against intracerebral challenge with virus . Using implanted osmotic minipumps or coadministering the drug with dimethyl sulfoxide failed to decrease the mortality rate . In guinea pigs infected genitally with HSV-2, topical drug treatment was more effective than placebo in reducing lesion severity and other clinical and virological variables . These effects were noted whether the drug treatment was initiated 3 or 24 h after infection (ascertained serologically) . Drug-treated animals had a significantly lower herpes antibody titer than did placebo-treated guinea pigs, suggesting that the drug can also reduce the viral antigen load . In this model, the drug appeared to be as effective as topical phosphonoformate or acyclovir. J Comp Pathol, 1985 Oct, 95(4), 565 - 72 The alterations in pathogenicity and immunogenicity of a Kenya sheep and goat pox virus on serial passage in bovine foetal muscle cell cultures; Davies FG et al.; A Kenyan sheep and goat pox virus was attenuated by serial passage in bovine foetal muscle cell cultures . The pathogenicity of the strain was lost between the 15th and 20th passages . Serum-neutralizing antibody developed after inoculation with passages tested up to the 50th . These passages appeared to protect animals against laboratory challenge by intra-dermal titration . The 18th passage was successfully and extensively used to control the disease under field conditions. J Neurosci, 1985 Oct, 5(10), 2690 - 5 gamma-Aminobutyric acid receptors on chick ciliary ganglion neurons in vivo and in cell culture; McEachern AE et al.; In the chick ciliary ganglion, preganglionic terminals maintain cholinergic synapses on the choroid neurons and both cholinergic and electrical synapses on the ciliary neurons . The preganglionic terminals also contain enkephalin- and substance P-like immunoreactivity, suggesting that transmission through the ganglion is more complicated than is indicated by the known synaptic connections . We report here that embryonic chick ciliary ganglion neurons also have gamma-aminobutyric acid (GABA) receptors and that GABA applied to the ganglion can block transmission elicited by preganglionic stimulation . Studies on the neurons in cell culture indicate that the GABA response is mediated by GABAA receptors: GABA activates a Cl- conductance, and the response can be mimicked by muscimol and blocked by bicuculline or picrotoxin . The GABA receptors are regulated independently from acetylcholine (ACh) receptors on the neurons since the levels of ACh and GABA sensitivity are influenced differently by culture age and by chronic exposure to GABA or elevated K+ concentrations . Application of GABA to intact ciliary ganglia increases the membrane conductance of ganglionic neurons (as in culture), reduces to subthreshold the amplitude of excitatory postsynaptic potentials in the neurons elicited by preganglionic stimulation and completely blocks transmission through the ganglion . A native source of ligand for the receptors in vivo has yet to be identified. Antimicrob Agents Chemother, 1985 Oct, 28(4), 578 - 80 Activity of arildone with or without interferon against acute hemorrhagic conjunctivitis viruses in cell culture; Langford MP et al.; Arildone (WIN 38020; 4-{6-(2-chloro-4-methoxyphenoxy)hexyl}-3,5-heptanedione) inhibited the infectivity of acute hemorrhagic conjunctivitis viruses in tissue culture . Arildone did not inhibit interferon (IFN) production or IFN activity . Treatment of cultures with combinations of arildone and IFN resulted in an additive inhibition of acute hemorrhagic conjunctivitis virus production. Tsitologiia, 1985 Oct, 27(10), 1156 - 63 {Dependence of ion transport across the plasma membrane on cell culture density . II . Active and passive cation transport during the growth of L cell cultures}; Marakhova II et al.; Rubidium and lithium influxes as well as intracellular potassium and sodium contents were investigated in L cells during the culture growth . In sparse culture over the cell densities 0.5-3 X 10(4) cells/cm2 ouabain-sensitive rubidium influx is small and ouabain-resistant lithium influx in high . With the increase in culture density up to 4-5 X 10(4) cells/cm2 the active rubidium influx, mediated by ouabain-sensitive component, is enhanced, and ion "leakage" tested by lithium influx is diminished . Simultaneously with the exponential growth of culture the intracellular potassium content is increased and the intracellular sodium content is decreased resulting in the higher K/Na ratio in cell . During the further transition to dense culture and in stationary state (10-17 X 10(4) cells/cm2) the sodium content and lithium influx do not change significantly, but the potassium content is decreased . The decrease in intracellular potassium is correlated with that in the portion of cells in S-phase from 27-30 to 12% . Thus, in transformed cells the density-dependent alterations in membrane cation transport are observed. Cell Immunol, 1985 Oct 1, 95(1), 124 - 36 Synergistic interaction of a cytokine produced by embryonic fibroblasts and a lymphokine contained in Con A-stimulated spleen cell culture for murine macrophage differentiation: a model experiment using cell line cells, M1; Noro N et al.; The modulating activity of the culture supernatant of Con A-stimulated murine spleen cells for macrophages was investigated by using M-1 cells, which could differentiate into macrophage-like cells (referred to as M+1 hereafter), cocultured in a conditioned medium (CM) containing a differentiation factor (DF) obtained from the secondary culture of murine embryonic fibroblasts . DF induced Ia antigens on M-1 cells at a high rate in parallel with the appearance of Fc-receptor (FcR)-dependent phagocytic activity for erythrocytes coated with an antibody (EA) . In contrast, Con A-sup alone had no modulating effect on M-1 cells . However, the Con A-sup stimulates synergistically M-1 cells with DF . Thus, coculture of M-1 cells with Con A-sup and DF generates M++1 cells which possess higher phagocytic activity than M+1 cells . These cells also exhibited stronger accessory cell activity than M+1 cells when tested for their promoting effect on IL-2 production by Sephadex G-10-passed spleen cells . The accessory cell activity of M++1 cells was further enhanced by culture with lymphocytes in the presence of indomethacin while that of M+1 cells did not change . These findings suggest that M-1 cells probably acquire potentiating, as well as inhibitory activity at the same time when cultured with DF and Con A-sup . The functional maturation caused by Con A-sup seemed to be associated with the expression of a receptor for a lymphokine, termed phagocytosis-augmenting factor (PAF) which is present in the Con A-sup . Such a receptor appeared to be common to macrophage lineage, since PAF in Con A-sup was absorbed out with splenic adherent cells and peritoneal exudate cells (PEC) in addition to M+1 cells, but not with nonadherent splenic lymphocytes or lymphoid cell line cells, such as EL-4 and L-1210 . This fact suggests that PAF is different from interferon-gamma (IFN) which is known to modulate the function of lymphocytes . Inability of PAF to bind Cibacron Blue-Sepharose, unlike IFN, supports this notion . The molecular weight of PAF is approximately 2-3 X 10(4) . Thus, the present studies suggest the requirement of at least two signals for the full maturation of macrophages, a cytokine represented by DF and a lymphokine, by PAF. Biochem Biophys Res Commun, 1985 Sep 16, 131(2), 593 - 9 Partial characterization of intra- and extracellular forms of the glycoprotein hemopexin in liver cell culture and cell-free translation; Katz NR et al.; Primary cultures of rat hepatocytes produce extracellular and intracellular species of hemopexin . We examined the presence of high-mannose oligosaccharides and neuraminic acid residues in these species by comparing their electrophoretic mobility on SDS-PAGE before and after digestion by endoglycosidase H and neuraminidase . The predominant intracellular form was not susceptible to digestion by neuraminidase but was sensitive to endoglycosidase treatment, which digested it to a species with a molecular weight comparable to that of the sole hemopexin species produced by tunicamycin-treated hepatocytes and that produced by in vitro translation . By contrast, both the minor intracellular and the extracellular species of hemopexin were neuraminidase-, yet not endoglycosidase H-sensitive, and may be identical . It can be concluded that the intracellular precursor contains high-mannose type oligosaccharides which are processed to complex type oligosaccharides shortly before secretion of hemopexin. J Microsc, 1985 Sep, 139 ( Pt 3), 321 - 5 A simple re-embedding method for the preparation of testicular cell cultures for light and electron microscopy; Bowen Jones H et al.; A simple and reproducible method is described for the preparation of cell cultures for light and transmission electron microscopy . Mixed testicular cells are grown on collagen layered, PTFE-coated coverslips . Thin sheets of plastic-embedded cells are packed together and re-embedded permitting the examination of sections cut perpendicular to the plane of the culture and, thus, the intercellular relationships at different levels throughout the culture. Prenat Diagn, 1985 Sep-Oct, 5(5), 305 - 12 Reduction of sera requirements in amniotic fluid cell culture; Chang HC et al.; Reduction in serum requirement for culture of primary human amniotic fluid cells can be achieved by the addition of 10 growth-promoting factors to the nutrient medium . This supplemented medium preserves cell types normally found in amniotic fluid cell cultures supplemented with 20-30 per cent fetal bovine serum . The volume of amniotic fluid required to initiate culture can be as little as 1 ml . Amniotic fluid samples contaminated with red blood cells with no visible clot also grow well in the low serum medium . Cell-free amniotic fluid combined with equal parts of supplemented medium is useful in initiating cell culture. Biomaterials, 1985 Sep, 6(5), 346 - 51 Biocompatibility testing of prosthetic implant materials by cell cultures; Pizzoferrato A et al.; The main aspects of biocompatibility are discussed first then the methodology used and the results obtained . The cells used included epithelial cells, lymphocytes, fibroblasts, and macrophages . The most commonly used methods were morphological observation, radioactive tracer uptake, and chemotactic migration analysis . It is concluded that cell cultures are a reliable and sensitive method for initial screening in testing the biocompatibility of the materials used in the construction of prosthetic implants. In Vitro Cell Dev Biol, 1985 Sep, 21(9), 505 - 8 Rat kidney epithelial cell culture for metal toxicity studies; Cherian MG; Evaluation of the potential adverse human health effects of low-level chronic exposure to heavy metals is dependent on the basic knowledge of the cellular and molecular toxicology of these metals . The use of various cell culture systems has greatly facilitated our knowledge of the cellular effects . Inasmuch as most of the acute and chronic toxic effects of metals occur primarily on the renal proximal tubules, the development of a rat kidney epithelial cell culture has provided a unique system to study the uptake and mechanism of toxicity of metals and their intracellular binding ligands . In the presence of D-valine, fibroblast growth was retarded and a primary epithelial monolayer culture was selectively grown from rat kidney cells . A distinct difference in the uptake of chemically similar divalent metals, such as Pb2+, Hg2+, Cd2+, and Zn2+, was observed in these cells . Both Pb2+ and Hg2+ were more avidly taken up by kidney cells than Cd2+ and Zn2+ salts and they also showed increased toxicity . On the other hand, the cellular uptake of Cd from cadmium-metallothionein (CdMT) was much less than from CdCl2, but CdMT was about seven times more toxic than CdCl2 when added to the renal cell culture . The cytotoxicity of CdCl2 was decreased significantly with pretreatment of the cells with CdCl2, although this had no effect on the toxicity of CdMT . The cellular toxicity of CdMT occurred probably during the process of its transport across the plasma membrane whereas that of CdCl2 occurred after it had entered the cell . Thus rat kidney epithelial cells may be a useful tool to study the mechanism of renal toxicity of environmental chemicals and drugs. J Virol, 1985 Sep, 55(3), 760 - 7 ts1, a Paralytogenic mutant of Moloney murine leukemia virus TB, has an enhanced ability to replicate in the central nervous system and primary nerve cell culture; Wong PK et al.; A temperature-sensitive mutant of Moloney murine leukemia virus TB (MoMuLV-TB), ts1, which is defective in intracellular processing of envelope precursor protein (Pr80env), also possesses the ability to induce hind-limb paralysis in infected mice . To investigate whether ts1 has acquired neurotropism and to determine to what extent it can replicate in the central nervous system, we compared viral titers in the spleen, plasma, spinal cord, and brain throughout the course of infection of mice infected with ts1 and parental wild-type (wt) MoMuLV-TB . In both the ts1- and wt-inoculated mice, the concentrations of infectious virus recovered from the plasma and spleen increased rapidly and reached a plateau by 10 days postinfection (p.i.) . In contrast, virus concentrations in the spinal cord and brain of ts1-inoculated mice increased gradually and reached a titer comparable to that in the spleen and exceeding that in the plasma only at 25 to 30 days p.i . At this time, the virus titer was approximately 200X greater in ts1-infected spinal cord tissue and approximately 20X greater in ts1-infected brain tissue than in the same wt-infected tissues . Paralysis became evident at 25 to 30 days p.i . in ts1-inoculated mice, whereas the wt-inoculated mice were normal . In addition, a substantial amount of Pr80env was detected in the spinal cords of ts1-inoculated mice compared with that found in the spinal cords of wt-inoculated mice . The infectious virus isolated from ts1-infected nerve tissue was found to possess the characteristic phenotype of the ts1 virus . Microscopic lesions of ts1-inoculated mice at 30 days p.i . consisted of vacuolar degeneration of motor neurons and spongy change of white matter in the brain stem and spinal cord . Similar but less severe lesions were observed in wt-inoculated mice . With primary cultures of central nervous system tissue we showed that ts1 can infect and replicate in both neuron and glial cells . In contrast, although wt MoMuLV-TB replicated in glial cell-rich culture, viral replication was barely detectable in neuron-rich culture. Endocrinology, 1985 Sep, 117(3), 939 - 46 Catecholestrogen regulation of prolactin synthesis in pituitary cell culture; Shupnik MA et al.; The 2-hydroxycatecholestrogens, 2-hydroxyestradiol {1,3,5-(10)estratriene-2,3,17 beta-triol} (2-OHE2) and 2-hydroxyestrone {2,3-dihydroxy-1,3,5-(10)-estratriene-17-one} (2-OHE1) were tested for their ability to alter PRL production and PRL messenger RNA (mRNA) levels in rat pituitary cell cultures . Treatment of cells with 10(-8) M 2-OHE1 or 2-OHE2 resulted in increased PRL secretion at 24 and 48 h (to 167% and 211% of control, respectively), but not at 4 h . Metabolism studies of radioactive 2-OHE1 and 2-OHE2 in parallel cultures demonstrated that the major metabolite at all times for either compound was the 2-methoxy derivative . After 24 h of treatment, nearly 40% of each compound was the original catecholestrogen, and at no time was there any detectable conversion to estradiol or estrone . Treatment of pituitary cells for 48 h with increasing concentrations of 2-OHE1 or 2-OHE2 resulted in a biphasic PRL dose response . PRL secretion was increased 3.6-fold for 2-OHE2 and 2.4-fold for 2-OHE1 between 10(-10) M and 10(-8) M . At concentrations above 5 X 10(-8) M, however, both compounds decreased PRL levels until, at 10(-6) M 2-OHE1 or 2-OHE2, PRL levels were 40-70% of control . Changes in PRL mRNA levels paralleled those of secretion . Treatment of pituitary cells with 10(-8) M of either 17 beta-estradiol (E2), 2-OHE1, or 2-OHE2 resulted in 2- to 5-fold increases in translatable and hybridizable PRL mRNA . The addition of 10(-7) M E2 plus 10(-8) M 2-OHE1 or 2-OHE2 resulted in PRL secretion and PRL mRNA levels equal to those resulting from E2 stimulation alone . The inhibition in PRL secretion and PRL mRNA levels caused by 10(-6) M 2-OHE1 or 2-OHE2 was partially overcome by coincubation of cultures with E2 . Thus, 2-OHE1 and 2-OHE2 at low concentrations (less than 10(-8) M) can act on the pituitary as E2 agonists to increase PRL synthesis but at high concentrations may act as inhibitors of PRL production. Biull Eksp Biol Med, 1985 Sep, 100(9), 330 - 1 {Immunologic typing of splenic lymphocytes from human fetuses--donors of pancreatic islet cell cultures}; Krasavtseva TK et al.; To determine the HLA-phenotype of a potential donor of pancreatic islet cells, use was made of lymphocytes from 18-25-week-old human fetuses . The HLA-phenotype was clearly established in 39 out of 52 cases . In 13 cases, the authors failed to reveal histocompatibility antigens because of low viability of lymphocytes . The relationship was ascertained between the detectability of HLA-antigens in fetal donors and cytophysiological characteristics of islet cell cultures prepared from the pancreas. Cell Immunol, 1985 Sep, 94(2), 587 - 97 Effects of thymic myoid cell culture supernatant on cells from lymphatic tissues; Kamo I et al.; Conditioned media (MCM) of cloned thymic myoid cells (IT45R92, R613Ad, and R615B2) were used to investigate their possible involvement in thymic biological events . Those myoid cells produced in a culture medium biological activities capable of stimulating the growth of thymocytes, spleen cells, and bone marrow cells of mice and rats . Surface markers detected on spleen cells proliferating in MCM were characteristic of monocyte-macrophage lineages (C3R, Fc gamma R, asialo GM1) and T-cell lineages (Thy 1) but not B cells (sIgG) . Chromatographic studies also suggested that the biological activities of MCM could be separated into two different molecular entities, such as a colony-stimulating activity and an interleukin 1-like activity which supported the growth of monocyte-macrophage lineages and T-cell lineages, respectively . These results indicate that thymic myoid cells produce cytokines important for the regulation of intrathymic interleukin cascade by which clonally differentiated thymic lymphocytes may be expanded into a sizable pool. Mol Cell Endocrinol, 1985 Sep, 42(2), 175 - 83 Electrophoretic characterization of active renin from human kidney and inactive renin from a human chorionic cell culture; Auzan C et al.; Enzymatically inactive human renin from chorionic cells in culture is significantly distinct in polyacrylamide gel electrophoresis (pH 8.17, 0 degree C) from active human kidney renin . The inactive renin is larger and more basic than the active renin; their molecular weights derived from gel electrophoretic retardation coefficients relate as 47.5/35.3 kDa, their valences (net protons/molecule) as 2.14/1.85 . In gel electrofocusing conducted in a mixture of simple buffers, both inactive and active renins exhibit 2 components at the steady-state . The molecular size and basicity of inactive renin are consistent with the hypothesis that it may be a precursor (prorenin), although the possibility that it is an inhibitor complex cannot be ruled out. Proc Natl Acad Sci U S A, 1985 Sep, 82(18), 6330 - 4 Transplants of Schwann cell cultures promote axonal regeneration in the adult mammalian brain; Kromer LF et al.; Transplantation of embryonic brain tissue or mature peripheral nerves into the adult mammalian central nervous system promotes axonal regrowth from axotomized central nervous system neurons; however, the cellular origin and molecular nature of the factors promoting axonal growth in vivo are unknown . To further characterize cellular environments that facilitate regeneration of central nervous system axons, we developed a methodology whereby cultured cell preparations can be transplanted into the brain of mature mammals . For this procedure, lesions are produced in the septal-hippocampal system of adult rats, and selected regions from collagen-supported Schwann cell/neuron cultures (consisting of Schwann cells, extracellular matrix, and degenerating neuronal processes and myelin but devoid of neuronal perikarya and fibroblasts) are positioned within the intracephalic cavity so that they bridge the lesion gap (approximately 3 mm) separating the septum and hippocampus . At various time up to 3 weeks after transplantation, specimens were prepared for acetylcholinesterase histochemistry and the immunocytochemical localization of laminin (an extracellular matrix protein) and C-4 (a Schwann cell membrane antigen) . All specimens (from uninjured controls and from animals with either acellular collagen or mature Schwann cell/extracellular matrix transplants) contained laminin immunoreactivity associated with the meninges, choroid plexus, ependyma, and cerebral blood vessels . All animals with transplants showed prominent laminin staining on astrocytic processes along the intracephalic cavity, but only the Schwann cell/extracellular matrix transplants exhibited dense laminin and C-4 immunoreactivity within the cellular portion of the transplants . Regeneration of acetylcholinesterase-positive septal fibers occurred only in animals containing Schwann cell/extracellular matrix transplants . By 6 days after transplantation, acetylcholinesterase-positive fibers were observed both on laminin-positive cellular tissue strands connecting the septum and the Schwann cell/extracellular matrix transplants and on the initial portions of the transplants . By day 14, acetylcholinesterase-positive fibers traversed the entire lesion cavity in intimate association with the laminin- and C-4-positive cellular layer of the transplants and reinnervated the host hippocampus . However, cholinergic fibers were not associated with all laminin-containing processes along the lesion cavity nor did they grow along acellular collagen transplants . These results indicate the presence of factors in transplants of cultured Schwann cells and their associated extracellular matrix that promote rapid regeneration of central nervous system cholinergic axons in vivo. J Cell Physiol, 1985 Sep, 124(3), 411 - 23 Phosphate uptake by primary renal proximal tubule cell cultures grown in hormonally defined medium; Waqar MA et al.; The uptake of labeled inorganic phosphate into primary rabbit kidney proximal tubule cells has been examined . Phosphate was accumulated into the primary proximal tubule cells against a concentration gradient . This accumulation was sensitive to inhibition by metabolic inhibitors . The dependence of phosphate uptake on the extracellular phosphate concentration was examined . Similarities were observed between primary proximal tubule cells and the LLC-PK1 cell line in these regards . These phosphate uptake data were then plotted on a Lineweaver-Burke plot . A nonlinear plot was obtained, which suggested that phosphate uptake occurs by means of a Na+ dependent, carrier mediated process, as well as by another Na+ independent mechanism . The pH dependence of phosphate uptake was also examined . Unlike previous observations with LLC-PK1 cells, optimal phosphate uptake occurred at pH 6.5 . However, this difference between the two cell culture systems may possibly be explained by differences in uptake conditions . The dependence of phosphate uptake on the extracellular NaCl concentration was examined at three different pH values . The rate of phosphate uptake at pH 7.0 was observed to saturate at a lower NaCl concentration than at either pH 6.0 or pH 6.5 . Furthermore, the optimal rate of phosphate uptake at pH 7.0 was observed to be higher than at the other two pH values studied when the NaCl concentration was below 120 mM . However, when the NaCl concentration was raised to 150 mM, optimal phosphate was observed to occur at pH 6.5 rather than at pH 7.0 . These observations may be explained if the pH affects not only the rate of phosphate uptake but also the affinity of the phosphate uptake system for sodium . Phosphate uptake was also observed to be sensitive to several agents, Na2 X SO4 and NaSCN, which affect the membrane potential . As observed with phosphate uptake by LLC-PK1 (and renal brush border membrane vesicles), phosphate uptake was highly sensitive to inhibition by the phosphate analogue arsenate . Novel observations were that the phosphate analogue vanadate and its cellular metabolite vanadyl stimulated the initial rate of phosphate uptake. Biochem Int, 1985 Sep, 11(3), 273 - 80 Monensin inhibits collagenase production in osteoblastic cell cultures and also inhibits both collagenase release and bone resorption in mouse calvaria cultures; Cowen KS et al.; Monensin, a monovalent cation ionophore, inhibited collagenase production in mouse osteoblast-rich bone cell and clonal osteogenic cell cultures . Inhibition of parathyroid hormone-stimulated bone resorption by monensin was also studied in calvaria cultures . Collagenase activity levels in the medium decreased concomitantly with the inhibition of bone resorption by monensin, indicating that monensin inhibited bone resorption by blocking collagenase secretion from osteoblasts in bone explants. Cell Struct Funct, 1985 Sep, 10(3), 271 - 8 Myelin formation in dissociated cell cultures of rat embryonic cerebral hemispheres; Hirano S et al.; Developing cultures of dissociated cerebral hemispheres obtained from 18-day-old embryonic rats synthesized, or activated, a myelin-related enzyme, 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNPase) . This increase in activity coincided with the onset of myelination . The presence of CNPase in oligodendrocytes and myelin was demonstrated using immunocytochemical staining techniques . Myelinated axons and myelin sheaths were clearly made visible by electron microscopy of 28-day-old cultures. Brain Res, 1985 Sep, 354(1), 55 - 65 The possible modulation of the development of rat dorsal root ganglion cells by the presence of 5-HT-containing neurones of the brainstem in dissociated cell culture; Forda S et al.; Reliable methods for coculturing dissociated rat brainstem cells together with dorsal root ganglion (DRG) cells have been established . The cells were characterized using autoradiographic, morphological, immunocytochemical and electrophysiological techniques . Light-level microscopy showed the cocultures to be extensively invaded by serotonin (5-HT)-containing neuronal processes and intense clustering of 5-HT-containing varicosities to occur in the vicinity of large round phase-bright neurones thought to be DRGs . Rather extensive fine ramification of the neuronal processes throughout the culture dish were visualized using scanning electron microscopy . Intracellular recording showed the brainstem neurones to be spontaneously active, electrically excitable and sensitive to a variety of transmitter candidates including serotonin (5-HT) and gamma-aminobutyric acid . Several different responses to 5-HT have been observed . These include a depolarization accompanied by an increase in membrane resistance, a depolarization accompanied by a decrease in membrane resistance and a hyperpolarization accompanied by an increase in membrane resistance . As established by others, DRG cells grown in isolation were always quiescent . The application of 5-HT produced no effect on membrane potential, resistance or excitability . In the brainstem-DRG cocultures 52% of DRG cells exhibited synaptic activity and occasional spontaneous spiking, both of which were abolished in the presence of tetrodotoxin or low-Ca2+/high-Mg2+ solution . Transmission electron microscopy confirmed the presence of synapses on the DRG cells . The spontaneously active DRG cells were also found to be sensitive to the application of 5-HT . Thus it appears that a source of 5-HT nerve terminals may regulate the development and pharmacological sensitivity of primary afferent neurones in culture. Blood, 1985 Sep, 66(3), 714 - 7 Effects of dibutyryl adenosine 3',5'-cyclic monophosphate on erythropoietin production in human renal carcinoma cell cultures; Hagiwara M et al.; A human renal carcinoma from a patient with erythrocytosis, serially transplanted into athymic nude mice, was grown in primary monolayer cell cultures . After reaching confluency, the cultured cells formed multicellular hemicysts (domes), which became more abundant as the cultures approached saturation density . Erythropoietin (Ep) production by this renal carcinoma in culture was only slightly increased at the time of semiconfluency but showed a marked increase after the cultures reached confluency, in parallel with dome formation . Dibutyryl adenosine 3',5'-cyclic monophosphate significantly (P less than .01) stimulated Ep production and dome formation in the semiconfluent and confluent cultures of the renal carcinoma. Probl Endokrinol (Mosk), 1985 Sep-Oct, 31(5), 67 - 70 {Results of transplantation of pancreatic islet cell cultures to patients with diabetes mellitus}; Shumakov VI et al.; Altogether 110 intramuscular pancreatic islet cell (PIC) culture transplantations were performed in diabetes mellitus patients from April 1981 to May 1984 at the Research Institute of Transplantology and Artificial Organs, USSR Ministry of Health: 65 allotransplantations of human fetal PIC and 45 xenotransplantations fo swine fetal PIC . Immunosuppressive therapy was not employed . The paper is concerned with an analysis of a therapeutic effect of the IC cell cull culture transplantation on 30 patients who were followed-up for not less than 1 year after operation . A pronounced and long-term antidiabetic effect was observed in wost of the recepients subjected to the intramuscular human fetal PIC culture transplantation: demand in exogenous insulin decreased, a durable stabilization of a course of disease in patients with labile diabetes mellitus was observed, and in patients with such diabetic complications as polyneuropathy, retinopathy and glomerulosclerosis certain regress of these complications was observed . As to the results of the intramuscular swine fetal PIC culture xenotransplantation the authors note that xenogenic transplantation as compared with IC culture allotransplantation possesses a less noticeable offect on diabetes mellitus complications. Parasite Immunol, 1985 Sep, 7(5), 467 - 78 Giardia muris-induced depression of the primary immune response in spleen and mesenteric lymph node cell cultures to sheep red blood cells; Belosevic M et al.; Primary in vitro plaque-forming cell (PFC) responses to sheep red blood cells (SRBC) were examined for spleen and mesenteric lymph node (MLN) cell populations from susceptible (A/J) and resistant (B10.A) mice during the infection with Giardia muris . Spleen and MLN cells isolated from mice during the acute phase of the infection were less responsive to SRBC in vitro than those from uninfected mice . Depressed anti-SRBC PFC response was detected earlier and was more pronounced in MLN cell cultures when compared to the response of spleen cell cultures . Spleen and MLN cells from donors infected with G . muris for 15 days had the capacity of depressing PFC response to SRBC of cells isolated from uninfected mice . This suppressor activity was localized in the plastic-adherent fraction of spleen cell populations isolated from A/J and B10.A mice . Since G . muris is a gastro-intestinal infection of mice, lower capacity of the MLN cells to respond to an antigenic stimulation in vitro may explain, in part, the proliferation of the trophozoites during the acute phase of the infection. Neuroscience, 1985 Sep, 16(1), 213 - 21 Neurons dissociated from rat myenteric plexus retain differentiated properties when grown in cell culture . III . Synaptic interactions and modulatory effects of neurotransmitter candidates; Willard AL et al.; We have used intracellular recordings to study synaptic interactions between myenteric neurons grown in dissociated cell culture . Intracellular stimulation of individual myenteric neurons caused several types of synaptic effects in nearby neurons: fast excitatory synaptic potentials mediated by nicotinic acetylcholine receptors; slow, non-cholinergic synaptic potentials; dual transmission having both fast cholinergic and slow non-cholinergic components and inhibition of spontaneously occurring fast nicotinic synaptic potentials . Fast nicotinic synaptic potentials were elicited by about 40% of neurons tested and often occurred spontaneously . The fast synaptic potentials were similar to those that have been studied in other autonomic neurons with respect to their estimated reversal potential and their sensitivity to cholinergic antagonists . The amplitudes of the fast synaptic potentials declined if evoked at frequencies greater than 0.5 Hz . Potentiation of the fast synaptic potentials was observed following high-frequency stimulation of presynaptic neurons . Several transmitter candidates modulated fast cholinergic transmission . Substance P and vasoactive intestinal peptide promoted nicotinic transmission by causing increased amplitudes of evoked and spontaneous fast synaptic potentials and an increased frequency of spontaneous synaptic potentials . gamma-Aminobutyrate and {Met}enkephalin both caused decreased amplitudes and frequency of nicotinic synaptic potentials . Serotonin depressed synaptic potentials in some neurons while enhancing them or having no effect in others . Slow, non-cholinergic, synaptic potentials were elicited by about 10% of neurons tested . These synaptic effects lasted 15-300s, caused depolarizations of 3-15 mv and were accompanied by increased neuronal input resistance . The transmitter(s) causing these slow synaptic potentials has not yet been identified.(ABSTRACT TRUNCATED AT 250 WORDS) Res Vet Sci, 1985 Sep, 39(2), 241 - 6 Adherence of Moraxella bovis to cell cultures of bovine origin; Annuar BO et al.; The adherence of five strains of Moraxella bovis to cell cultures was investigated . M bovis adhered to cultures of bovine corneal epithelial and Madin-Darby bovine kidney cells but not to cell types of non-bovine origin . Both piliated and unpiliated strains adhered but piliated strains adhered to a greater extent than unpiliated strains . Antiserum against pili of one strain inhibited adherence of piliated strains but caused only slight inhibition of adherence to the unpiliated strains . Treatment of bacteria with magnesium chloride caused detachment of pili from the bacterial cell and markedly inhibited adherence of piliated strains but caused only slight inhibition of adherence by the unpiliated strains . The results suggested that adhesion of piliated strains to cell cultures was mediated via pili but that adhesins other than pili may be involved in the attachment of unpiliated strains of M bovis to cells. Neuroscience, 1985 Sep, 16(1), 201 - 11 Neurons dissociated from rat myenteric plexus retain differentiated properties when grown in cell culture . II . Electrophysiological properties and responses to neurotransmitter candidates; Willard AL et al.; We have used intracellular recordings to study the electrophysiological and pharmacological properties of neurons that have been grown in cell cultures after having been dissociated from the myenteric plexus of the small intestine of newborn rats . Studies of action potential mechanisms revealed that all of the neurons could generate Na+-dependent action potentials in the presence of Ca2+-channel blockers and that about 70% could generate Ca2+-dependent action potentials when Na+ channels were blocked with tetrodotoxin . No neurons generated long afterhyperpolarizations after single action potentials but about 50% of neurons did so following trains of action potentials . Over 95% of the neurons tested accommodated rapidly to sustained depolarization . The effects of several enteric neurotransmitter candidates were studied by superfusing or pressure-ejecting test solutions while recording neuronal responses . All of the cultured neurons tested had nicotinic responses to acetylcholine . Subsets of neurons responded to muscarinic cholinergic agonists (slow depolarization and increased excitability), serotonin (fast depolarization or slow depolarization and increased excitability), gamma-aminobutyrate (fast depolarization), substance P (slow depolarization, biphasic fast and slow depolarization or increased excitability without a change in membrane potential), vasoactive intestinal peptide (slow depolarization and increased excitability), or {Met}enkephalin (slow hyperpolarization and/or decreased action potential duration) . We conclude that myenteric neurons grown in cell culture retain many of the physiological and pharmacological properties that they have in situ . Such cultures will permit detailed biophysical and pharmacological studies of the mechanisms of action of enteric neurotransmitter candidates. Neuroscience, 1985 Sep, 16(1), 187 - 99 Neurons dissociated from rat myenteric plexus retain differentiated properties when grown in cell culture . I . Morphological properties and immunocytochemical localization of transmitter candidates; Nishi R et al.; We have developed procedures for dissociating neurons from the myenteric plexus of the small intestine of newborn rats and for growing those neurons in cell cultures for up to 3 months . Neurons in these cultures retain many of the differentiated properties of myenteric neurons in vivo . This is the first of a series of 3 papers describing those properties . In this paper, we describe the morphology of cultured neurons that we have observed with light and electron microscopy; we also describe the patterns of straining observed when immunocytochemical techniques were used to localize neurotransmitter candidates in the cultured neurons . Intracellular injections of a fluorescent dye, Lucifer yellow, revealed that many of the cultured neurons had morphologies similar to those of myenteric neurons in vivo . When thin sections of cultures were viewed in an electron microscope, many neurons were observed to have numerous small (40-60 nm), clear synaptic vesicles and/or large (80-150 nm), opaque-cored (p-type) vesicles . Synaptic profiles were most often observed on neuronal somata . Neurons containing immunoreactive serotonin, substance P, somatostatin, enkephalin, bombesin and gastrin/cholecystokinin were observed in about the same proportions as they occur in the intact myenteric plexus . Neurons containing immunoreactive vasoactive intestinal polypeptide were found in higher numbers than reported in vivo . Neurons containing immunoreactive neurotensin, secretin and glutamate decarboxylase were not observed . An antiserum directed against choline acetyltransferase stained 40-50% of the neurons . We conclude that myenteric neurons continue to express much of their normal differentiated properties even when they are removed from the gut, dissociated into a suspension of single cells and grown in culture . Such cultures will be useful for correlating the morphological, biophysical, pharmacological and synaptic properties of individual myenteric neurons and for testing the ability of altered environmental conditions to change those properties. Tsitologiia, 1985 Sep, 27(9), 1011 - 20 {Dependence of ion transport across the plasma membrane on the density of the cell culture . I . Ion flows and the potassium and sodium content in 3 Chinese hamster cell lines (CHO)}; Marakhova II et al.; Cation transport has been investigated in three lines of Chinese ovary cells CHO-K1 during the cell culture growth . With the increase in the cell density potassium and sodium contents decreased from 1.2 to 0.8-0.5 and from 0.5 to 0.15-0.1 mmole/g protein, respectively . The time courses of potassium and sodium changes were different, and the increase in intracellular K/Na ratio from 1.5-2.0 to 5-10 with the increase in cell density was revealed . The rubidium influx was found to decrease during the culture growth mainly due to the decrease in ouabain-inhibitable and (ouabain + furosemide)- non-inhibitable influxes . The changes in cation fluxes and cation contents were observed in transformed cells without contact inhibition of division and were considered as a manifestation of density-dependent alterations of plasma membrane. Proc Natl Acad Sci U S A, 1985 Sep, 82(18), 6325 - 9 Dissociated cell culture of cholinergic neurons from nucleus basalis of Meynert and other basal forebrain nuclei; Nakajima Y et al.; Degeneration of cholinergic neurons from the basal forebrain nuclei is suspected to be the cause of Alzheimer disease . We have developed dissociated cultures of cholinergic neurons from these nuclei (the nucleus basalis of Meynert, the medial septal nucleus, and the diagonal band nuclei) . Brain slices of the forebrains were made by a vibratome, and the basal forebrain nuclei were dissected out, dissociated, and cultured . Choline acetyltransferase immunocytochemistry and acetylcholinesterase cytochemistry revealed large cholinergic cells (average diameter, 20-25 micron) in these cultures . About 75% of large neurons (20 micron or larger in diameter) were cholinergic . Electrophysiological experiments were performed on these large neurons . The neurons usually did not show spontaneous firing, but steady depolarizations produced trains of action potentials, which adapted quickly . The neurons responded with depolarization to the application of L-glutamic acid . Substance P produced depolarization (sometimes hyperpolarization), and during the depolarization membrane resistance was increased. Mol Pharmacol, 1985 Sep, 28(3), 269 - 77 Barbiturates decrease voltage-dependent calcium conductance of mouse neurons in dissociated cell culture; Werz MA et al.; Barbiturates have been shown to reduce presynaptic release of neurotransmitter . It is likely that barbiturates alter transmitter release by decreasing calcium entry since barbiturates decrease calcium influx into synaptosomes and reduce the maximal rate of rise and duration of calcium-dependent action potentials . The mechanisms of barbiturate action on neuronal calcium entry have been studied using mouse dorsal root ganglion neurons in cell culture . Dorsal root ganglion neuron action potentials have a calcium-dependent component which is decreased by the barbiturates, pentobarbital (50-500 microM) and phenobarbital (500-2000 microM) . Calcium-dependent action potential after hyperpolarization was also decreased by barbiturates . Intracellular injection of the potassium channel blocker, cesium, enhanced barbiturate actions . In voltage-clamp studies, barbiturates reduced inward calcium current and calcium chord conductance without altering the leak conductance which is present after all calcium conductance was blocked by application of cadmium ions (100 microM) . Calcium current inactivation was accelerated by barbiturates but unaffected by cadmium . We conclude that barbiturates reduce calcium conductance by enhancing calcium channel inactivation or by producing open channel block of calcium channels. Cancer Res, 1985 Sep, 45(9 Suppl), 4595s - 4597s Evidence for HTLV-III in T-cells from semen of AIDS patients: expression in primary cell culture, long-term mitogen-stimulated cell cultures, and cocultures with a permissive T-cell line; Zagury D et al.; The development of acquired immunodeficiency syndrome or of acquired immunodeficiency syndrome-related complex by transmission of human T-lymphotropic retrovirus III by semen has previously been implicated by epidemiological studies . In vitro investigations were performed on mononuclear cells obtained from the semen of patients with acquired immunodeficiency syndrome to identify human T-lymphotropic retrovirus III or related retrovirus . The presence of human T-lymphotropic retrovirus III was demonstrated (a) in primary cell cultures, by the detection of the Mr 24,000 protein by indirect immunofluorescence assays by Day 6; (b) in activated long-term cell culture by reverse transcriptase activity, by indirect immunofluorescence (Mr 24,000 protein); and (c) in cocultures of T-cells from semen of AIDS patients and H9 cells by reverse transcriptase activity, indirect immunofluorescence, and the presence of virus particles by electron microscopy. Biochem Biophys Res Commun, 1985 Aug 30, 131(1), 167 - 73 Hormonal regulation of benzo{a}pyrene metabolism in human adrenocortical cell cultures; Hornsby PJ et al.; In cultured fetal human adrenocortical cells, metabolism of the carcinogen benzo{a}pyrene was found to be unresponsive to the xenobiotic inducers 3-methylcholanthrene, benz{a}anthracene and 2,3,7,8-tetrachlorodibenzo-p-dioxin . However, exposure of cultures to the hormone adrenocorticotropin (ACTH) for 48 hours stimulated benzo{a}pyrene metabolism 3-fold . The major metabolite was the 7,8-diol . Other compounds which stimulate the production of adrenocortical cell cyclic AMP (forskolin and cholera toxin) as well as monobutyryl cyclic AMP also increased benzo{a}pyrene metabolism . Human adrenocortical cells thus provide an unusual example of hormonal regulation of the metabolism of a carcinogen. Neurosci Lett, 1985 Aug 5, 58(3), 293 - 7 Glutamate neurotoxicity in cortical cell culture is calcium dependent; Choi DW; Brief exposure to glutamate produced widespread neuronal death in mature, but not young, cortical cell cultures . Extracellular sodium replacement or addition of tetrodotoxin produced only minor reduction in this toxic neuronal loss . However, removal of extracellular calcium markedly reduced neuronal loss, and elevation of extracellular calcium accentuated neuronal loss . These observations suggest that the toxicity of glutamate on cortical neurons may depend primarily on the presence of extracellular Ca, probably through a mechanism which is distinct from simple 'excitotoxicity'. Methods Find Exp Clin Pharmacol, 1985 Aug, 7(8), 403 - 8 The effect of 4'-deoxypyridoxine on rat heart cell cultures under oxygen deficiency; Millart H et al.; The purpose of this work was to study the consequences of an intracellular depletion of pyridoxal-phosphate (PLP) in rat heart cells in culture submitted to oxygen deficiency . Glucose (GLc) and 4'-deoxypyridoxine (DOP) effects were separately evaluated after 150 mn of partial oxygen deprivation (N2) with regard to enzyme leakage, beating rates, intracellular concentrations of PLP and the ratio glycogen phosphorylase activity (-5'-AMP/+5'-AMP) . PLP concentrations were higher in the air-GLc group than in the air group . alpha-Hydroxybutyrate dehydrogenase (alpha-HBDH) and creatine kinase (CK) leakages were observed in the N2 group, whereas 1.17 +/- 0.17 mM GLc prevented leakage . Beatings stopped in the N2 group, slightly decreased in the N2-GLc group; the ratio (-5'-AMP/+5'--AMP) increased only in the N2 group . With a 1 mM DOP pretreatment a significant decline of PLP was observed either with or without GLc in the culture medium . The ratio (-5'-AMP/+5'-AMP) was lower in the air-DOP group than in the air group, whereas beatings, alph-HBDH and CK activities were identical to the control group . ImM DOP in the N2-DOP group partially prevented alpha-HBDH and CK leakages into the medium but did not prevent the decrease in contractile activity . Surprisingly, PLP concentration increased in the N2-DOP group/air-DOP group and the increase in ratio (-5'-AMP/+5'-AMP) was higher in the N2-DOP group than in the N2 group when compared to the respective control groups.(ABSTRACT TRUNCATED AT 250 WORDS) Xenobiotica, 1985 Aug-Sep, 15(8-9), 705 - 11 Toxicity monitored with a correlated set of cell-culture assays; Borenfreund E et al.; A set of assays for toxicity has been developed in which cell cultures serve as an alternative to toxicity testing in vivo . One test is the assessment of the highest concentration of toxicant which produces minimal morphological alterations in cell cultures, followed by the determination of the amount of neutral red dye uptake by the cells . A second test is based on 50% inhibition of uptake of {3H}uridine after incubation of the cultures with the toxicant . There is good agreement between these assays in the rank correlation of a broad spectrum of compounds tested, as well as with the data from Draize rabbit eye irritancy tests in vivo. Xenobiotica, 1985 Aug-Sep, 15(8-9), 701 - 4 Photometric recording of cell viability using trypan blue in perfused cell cultures; Walum E et al.; A perfusion system was developed to increase the reliability of cell viability estimations by continuous measurement of the uptake of trypan blue dye . Monolayer cell cultures were perfused with buffer containing toxic substances and trypan blue, and the staining of cells was continuously recorded at 591 nm in a spectrophotometer . Using mercuric chloride and methylmercuric chloride as test substances with C6 rat glioma cells, time- and dose-dependent increases in light absorbance were obtained over a 12h recording period . Methylmercuric chloride at 10(-6) M caused a half-maximal increase in relative absorbance in 4.5 h, whereas the corresponding time for mercuric chloride was 10.5 h. Biol Reprod, 1985 Aug, 33(1), 187 - 200 Isolation and cell culture of the epithelial cells of cauda epididymidis of the bull; Joshi MS; A simple and rapid method has been described to isolate the epithelial cells of cauda epididymidis of adult bull . Perfusion of the lumen of the cauda epididymidis with 1 mg/ml collagenase in calcium- and magnesium-free Hank's balanced salt solution and incubation of the tissue at 37 degrees C for 90 min releases the principal and basal cells into the lumen . Several individual epithelial cells and cell aggregates without the contamination of stromal or smooth muscle cells can be flushed out at the end of incubation . The isolated epithelial cells, suspended in Dulbecco's medium with 10% horse serum, attach to plastic dishes within 3 h after seeding the cells and proliferate to form a monolayer in approximately 8-12 days . The electron microscopic study and immunostaining of the cultured epithelial cells indicate that the cultured cells are principal cells . The basal cells of the intact cauda epididymidis of bull show within their cytoplasm the presence of varying amounts of "lipid-like" material often closely associated with whorls of membrane. Brain Res, 1985 Aug, 353(2), 193 - 202 Gangliosides in postmitotic retina of chick embryo: changes in vivo and in cell cultures; Landa CA et al.; Developmental changes in ganglioside composition of postmitotic neural retina of chick embryo were analyzed by thin-layer chromatography . Gangliosides were identified by comparing their chromatographic mobilities with reference standards . The outstanding changes are decrease in the concentration of GD3L and increase in GD1a and GM1 concentrations . By depleting Muller glia cells from retina tissue of 13- and 16-day embryos (R13, R16) we determined that the bulk of the major gangliosides is associated with the neurons . Analysis of gangliosides in monolayer cultures of R13 and R16 cells highly enriched for Muller cell-derived gliocytes indicated that these cells express the same types of gangliosides as neurons, but in somewhat different concentrations and relative proportions; however, after time in culture these cells showed ganglioside types and changes in ganglioside profile that are not characteristic of normal retina . The latter observation is consistent with other evidence that the phenotype of Muller glia cells becomes altered in monolayer culture . In contrast to cultures of early embryonic retina, in organ cultures of later postmitotic retina, ganglioside composition did not continue to change as in normal development . This suggests that in postmitotic retina, normal developmental progression of ganglioside changes requires systemic and/or other conditions which are missing or altered when this tissue is isolated and cultured in vitro. J Neurosci, 1985 Aug, 5(8), 2028 - 34 Differential synapse formation and neurite outgrowth at two branches of the metacerebral cell of Aplysia in dissociated cell culture; Schacher S; The metacerebral cell (MCC) of Aplysia californica was isolated with its bifurcate axon from the cerebral ganglion and maintained in vitro under three conditions: (a) with no targets, (b) with identified buccal ganglion neurons B1 or B2 placed near the stump of the large diameter cerebral-buccal connective (CBC) branch, and (c) with B1 or B2 placed near the stump of the small diameter posterior lip nerve (PLN) branch . After 5 days in culture, the two branches differed significantly in the formation of chemical connections and in the extent of neurite outgrowth . Chemical connections characteristic of MCC-B1(B2) connections in vivo were observed in more than 90% of the cultures in which the buccal neuron was contacted by neurites emerging from the CBC branch, but in only 20% of the cultures in which the buccal neuron was contacted by neurites extending from the PLN branch . Neurite outgrowth from the CBC stump was always greater than growth from the PLN and was not affected significantly by the presence of a buccal neuron target at either branch . In contrast, neurite outgrowth from the PLN decreased significantly when the target was contacted by neurites from the CBC branch . These results suggest that two branches of a single neuron can differ in their capacities to form chemical connections . In addition, the two branches show differential growth as a result of target interaction at one of the branches . This simple in vitro system may therefore be useful in exploring the ways in which individual neurons control neurite extension from different branches as they seek to form chemical connections with their targets. J Lab Clin Med, 1985 Aug, 106(2), 162 - 74 Studies on the purification of thrombopoietin from kidney cell culture medium; McDonald TP et al.; A thrombocytopoiesis-stimulating factor (TSF) has been purified from human embryonic kidney (HEK) cell culture medium . In the initial purification step, crude HEK cell culture medium was fractionated with saturated ammonium sulfate (step I) . The proteins precipitated by 40% to 60% and 60% to 80% ammonium sulfate saturation increased the percent of sulfur 35 incorporation into platelets of assay mice (P less than 0.01) . The ammonium sulfate-precipitated proteins that contained significant TSF activity were further refined on Sephadex G-75 columns (step II) . The fraction containing the highest specific activity (greatest 35S incorporation into platelets of assay mice per milligram of protein) was further purified by diethylaminoethyl (DEAE)-cellulose column chromatography (step III) . TSF activity was eluted from the columns between 0.3 and 1.0 mol/L NaCl . Additional Sephadex chromatography of post-DEAE-chromatographic preparations further increased the purity of the TSF (step IV) . TSF from this four-step procedure was further processed on a DEAE-high-performance liquid chromatography (HPLC) column (step Va) or size exclusion (SE)-HPLC columns (step Vb) . After HPLC, the activity was localized in a region corresponding to a retention time of 6 to 8 minutes for the DEAE-HPLC, but longer times were found after SE-HPLC . TSF was further purified by additional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and SE-HPLC (step VI) . The final product had significant TSF activity and represented a purification of approximately 500,000-fold . It was also shown that the isoelectric pH of partially purified TSF was 4.7 and the molecular weight of the more highly purified preparation was approximately 32,000 . After extraction by a combination of chromatographic procedures, a single homogeneous product was obtained. Dev Biol, 1985 Aug, 110(2), 392 - 401 Nerve growth factor-independent development of embryonic mouse sympathetic neurons in dissociated cell culture; Coughlin MD et al.; Although ganglia from neonatal mouse sympathetic ganglia require nerve growth factor (NGF) for survival in culture, explanted sympathetic ganglia from early embryonic stages do not require added NGF for survival and growth . To determine whether the change in growth factor requirement is due to changes in the neurons themselves, to variations in neuronal populations, or to changes in nonneuronal cells, we examined the response to growth factors by dissociated sympathetic neurons at various stages of development . Results indicate that neurons from the 14-day gestational (E14) superior cervical ganglion (SCG) do not require NGF for initial survival and neurite extension, but do require the conditioned medium neurite extension factor, CMF . By 2 to 3 days thereafter, whether in vivo or in culture, most neurons have developed a requirement for NGF for survival in culture . During the same period, there is a concomitant increase in responsiveness to NGF alone as a trophic agent . Changes in response to NGF are not due to changes in NGF content of ganglia, to interactions in culture with nonneuronal cells, or to age-related differences in NGF requirements for maximum survival . The changes in growth factor requirements may be related to mechanisms regulating specificity of nerve-target connections. J Neurochem, 1985 Aug, 45(2), 536 - 43 Cholesterol biosynthesis and its regulation in dissociated cell cultures of fetal rat brain: developmental changes and the role of 3-hydroxy-3-methylglutaryl coenzyme A reductase; Volpe JJ et al.; Primary cultures of cells dissociated from fetal rat brain were utilized to define the developmental changes in cholesterol biosynthesis and the role of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase in the regulation of these changes . Cerebral hemispheres of fetal rats of 15-16 days of gestation were dissociated mechanically into single cells and grown in the surface-adhering system . Cholesterol biosynthesis, studied as the rate of incorporation of {14C}acetate into digitonin-precipitable sterols, was shown to exhibit two distinct increases in synthetic rates, a prominent increase after 6 days in culture and a smaller one after 14 days in culture . Parallel measurements of HMG-CoA reductase activity also demonstrated two discrete increases in enzymatic activity, and the quantitative and temporal aspects of these increases were virtually identical to those for cholesterol synthesis . These data indicate that cholesterol biosynthesis undergoes prominent alterations with maturation and suggest that these alterations are mediated by changes in HMG-CoA reductase activity . The timing of the initial prominent peak in both cholesterol biosynthesis and HMG-CoA reductase activity at 6 days was found to be the same as the timing of the peak in DNA synthesis, determined as the rate of incorporation of {3H}thymidine into DNA . The second, smaller peak in reductase activity and sterol biosynthesis at 14 days occurred at the time of the most rapid rise in activity of the oligodendroglial enzyme, 2':3'-cyclic nucleotide 3'-phosphohydrolase (CNP) . These latter observations suggest an intimate relationship of the sterol biosynthetic pathway with cellular proliferation and with oligodendroglial differentiation in developing mammalian brain. Zentralbl Bakteriol Mikrobiol Hyg {A}, 1985 Aug, 260(1), 51 - 6 Intra-cellular and extra-cellular growth of L . monocytogenes in chick embryo fibroblast cell culture; Basher HA et al.; The growth of L . monocytogenes in isolated chick embryo fibroblast cell culture was studied . Hanks balanced salt solution and Eagles minimal essential medium were shown to support a limited growth of L . monocytogenes . Extra-cellular growth on the maintenance medium occurs for 48 h prior to the establishment of intra-cellular organisms . As the uptake of the parasite by the cell culture takes place, intra-cellular replication begins with subsequent release of the organisms into the surrounding medium . The organism continues to replicate intra-cellularly until all the cell culture is destroyed. Lab Invest, 1985 Aug, 53(2), 122 - 31 Glomerular cell culture; Striker GE et al.; Glomerular cell culture has now become a widely used research technique . At the present time procedures are available to obtain isolated glomeruli from nearly all species . The isolation of individual cells has proven problematical . This is due to the lack of defined markers . Thus, it is not yet possible to determine the presence and relative degree of contamination by other glomerular or even nonglomerular, cell types . The importance of dealing with individual cell types, or defined mixtures, is exemplified by the variable results obtained in the assessment of prostaglandin synthesis within and between species . Several important bits of information have, nevertheless, evolved from glomerular cell culture experiments . The sites of synthesis of basement membrane components, as well as their composition, have been determined . Confirmation of the existence of a bone marrow-derived mesangial cell population and some of their properties has been obtained . The response of mesangial cells to, as well as their production of, various mediators has been shown . Finally, clear evidence for interspecies differences and similarities has been documented . Areas of controversy remain, including whether contractile mesangial cells are phagocytic, the presence of C3b receptors on epithelial cells, the amounts and types of certain extracellular matrix products synthesized by the various cell types, and the best methods for separation and culture of the individual glomerular cell types . There remain many fruitful areas for research . Fundamental questions such as the appropriate basal medium and supplements, the type of substrate, and the means to separate the individual cell types remain as unanswered or partially answered questions . When isolated cells are reliably obtained, the study of biosynthetic products in the resting and stimulated states must be again addressed . At that point, the effect of various and deliberate combinations of the glomerular cell types on the biosynthetic or proliferative responses will require further studies . For instance, although contractility mediated by receptors for angiotensin II has been assumed to be a specific property of mesangial cells, recent work shows that epithelial cells also respond to angiotensin II . In addition, the handling of immune complexes by various cells needs to be further investigated (43) . Similarly, the pharmacologic response of the diverse populations of glomerular cells represents another area of study that has just begun . Finally, these data will provide the backdrop on which the analysis of various induced and genetic diseases can be performed.(ABSTRACT TRUNCATED AT 400 WORDS) Genitourin Med, 1985 Aug, 61(4), 255 - 7 Comparison of direct immunofluorescence and cell culture for detecting Chlamydia trachomatis; Foulkes SJ et al.; Conventional cell culture methods were compared with a direct immunofluorescence test (MicroTrak, Syva UK, Maidenhead, Berkshire) to detect Chlamydia trachomatis in 137 patients (126 women, 11 men) attending a sexually transmitted diseases (STD) clinic . Results obtained by the two tests agreed in 87.6% of cases . Of 34 positive specimens, 17 were detected by culture and fluorescence, 15 by fluorescence only, and two by culture only . The excess of specimens that were negative on culture but positive on fluorescence might be accounted for by delays in culture (up to 18 hours) . The MicroTrak test appears to be of value in peripheral hospitals that have to rely on transporting specimens to larger centres for culture. Proc Natl Acad Sci U S A, 1985 Aug, 82(15), 5005 - 9 A contact-insensitive subpopulation in Syrian hamster cell cultures with a greater susceptibility to chemically induced neoplastic transformation; Nakano S et al.; We previously have identified a subpopulation of contact-insensitive (CS-) cells which lacks density-dependent inhibition of cell division in primary and low-passage cultures of Syrian hamster embryonic (SHE) fibroblastic cells . Further, we have shown that the proportion of these CS- cells declines as a result of the stable phenotypic conversion of the CS- cells to contact-sensitive (CS+) cells . To determine whether these transient CS- cells are more sensitive to carcinogenic/mutagenic perturbation, the susceptibility to neoplastic transformation and somatic mutation induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) was examined in clonally isolated cell cultures containing various proportions of CS- cells (0.02-4%) . The frequencies of morphological transformation, focus formation, and neoplastic transformation showed a positive correlation to the proportion of CS- cells in the treated cultures . In contrast, the frequency of MNNG-induced somatic mutation at the Na+,K+-ATPase locus was similar among cultures varying in their proportion of CS- cells . Thus, there is a transient subpopulation of CS- cells in primary SHE cell cultures that is more susceptible to neoplastic transformation although equally susceptible to induced point mutation . This dissociation between somatic point mutation and neoplastic transformation indicates a fundamental difference in the nature of these two phenomena . A possible relationship between the propensity of CS- cells (versus CS+ cells) to carcinogen-induced neoplastic transformation and the state of differentiation of the CS- cells is discussed. J Clin Endocrinol Metab, 1985 Aug, 61(2), 303 - 5 Increased content of neuropeptide Y in human pheochromocytoma cell cultures; Tischler AS et al.; Cells from three human pheochromocytomas grown in monolayer culture for 21 days contained markedly greater amounts of neuropeptide Y (NPY) than freshly dissociated cells from the same tumors . The content of NPY did not consistently correlate with the extent of neurite outgrowth in the cultures and was not further increased in the presence of nerve growth factor . The content of NPY was decreased in the presence of 10(-5) M dexamethasone in two cultures . The findings suggest that human pheochromocytoma cultures may be useful in studies of cellular and molecular mechanisms regulating NPY production, and that these mechanisms may differ somewhat from those regulating production of other regulatory peptides in cultures of the same tumors. Appl Environ Microbiol, 1985 Aug, 50(2), 523 - 6 Membrane-associated viral complexes observed in stools and cell culture; Williams FP Jr; Viral complexes observed to be membrane associated rather than clumped by antibody were detected in a rotavirus-containing stool specimen by negative-stain electron microscopy . These "viral packets" were also observed in cell culture fluids after repeated passaging and contained up to 100 virions . Other stool specimens have been observed to contain similar packets of parvovirus-like particles . Such complexes must be expected in fecally contaminated water. Otolaryngol Head Neck Surg, 1985 Aug, 93(4), 492 - 9 Interferon sensitivity of fibroblast cell cultures derived from patients with neoplasms of the head and neck; Richtsmeier WJ et al.; Interferon (IFN) is a protein with antiviral activity that has been shown to inhibit the growth of many different types of cells . We have measured the IFN sensitivity of nine cell cultures isolated from patients with squamous cell carcinoma and one with malignant melanoma of the head and neck . Normal-appearing fibroblast cultures isolated from these tissues appear quite sensitive to the antiviral effects of IFN . When the encephalomyocarditis virus yield reduction assay is used, these diploid cells are as sensitive to IFN-alpha as are newborn foreskin fibroblast cultures . A similar antiviral effect is seen with IFN-gamma . These cells are relatively insensitive to the antigrowth effect of both IFN preparations as measured by 3H thymidine incorporation and direct observations of cell growth . This is the same relative sensitivity as fibroblasts derived from normal patients . Since these cells are at least 100 times less sensitive to the antigrowth action of the IFNs, it appears unlikely that the IFNs play a significant role in the control of normal fibroblast growth, in contrast to the sensitivity of malignant cell lines. J Lipid Res, 1985 Aug, 26(8), 950 - 4 Cholesterol metabolism: use of D2O for determination of synthesis rate in cell culture; Esterman AL et al.; Cholesterol synthesis in cell culture in the presence of D2O yields a spectrum of enriched molecules having a relative abundance that indicates random substitution of deuterium for hydrogen . Quantitation of the absolute rate of cholesterol synthesis is obtained by isotope ratio mass spectrometry . Mevinolin and 26-hydroxycholesterol both decrease cholesterol synthesis rate but have a discordant effect on HMG-CoA reductase activity. J Clin Microbiol, 1985 Aug, 22(2), 199 - 204 Isolation and differentiation of herpes simplex virus and Trichomonas vaginalis in cell culture; Gentry GA et al.; During the period January 1982 to January 1985, 2,234 specimens were cultured for isolation of herpes simplex virus (HSV) . HSV was isolated from 23% of these, Trichomonas vaginalis was isolated from 1.6%, and 75.3% were negative . In 0.2% of these, HSV and T . vaginalis were isolated from the same specimen . Cytopathic effects produced by HSV were identified by their sensitivity to arabinosylthymine, whereas those produced by T . vaginalis were identified by their lack of sensitivity to arabinosylthymine and by observation of motility . Cytopathic effects produced by T . vaginalis were reproduced by trophozoites from axenic cultures of T . vaginalis as well as by lysates of T . vaginalis added to serum-free BHK cells. Endocrinology, 1985 Aug, 117(2), 488 - 91 Effects of adenosine analogs on glucagon-stimulated adenosine 3',5'-monophosphate formation in Sertoli cell cultures from immature rats; Eikvar L et al.; In the present study, we have examined the effects of two adenosine analogs, (-)N6-(R)phenyl-isopropyl-adenosine (PIA) and 2-chloro-adenosine, on glucagon- and FSH-stimulated cAMP production in Sertoli cell cultures isolated from immature (19-day-old) rats . Both FSH and glucagon caused a 5- to 10-fold stimulation of cAMP levels in the spent media from Sertoli cell cultures during an 18-h incubation . Addition of 1 microM PIA significantly inhibited both FSH- and glucagon-stimulated cAMP levels . In the presence of a maximal concentration of glucagon (2.5 micrograms/ml), PIA caused a concentration-dependent inhibition of cAMP formation, and the concentration of PIA causing half-maximal inhibition of cAMP formation (IC50) ranged from 0.5-1 nM . When Sertoli cells were incubated with increasing concentrations of glucagon (1.28 ng/ml to 4.00 micrograms/ml) in the absence and presence of either PIA (1.0 microM) or 2-chloro-adenosine (10.0 microM), the responses to glucagon, measured as cAMP formation, were almost completely abolished . 1-Methyl-3-isobutylxanthine (MIX), a well known inhibitor of cAMP phosphodiesterase activity, is also an inhibitor of adenosine binding to receptors on the cell membrane . When Sertoli cells stimulated with glucagon (2.5 micrograms/ml) were incubated in the absence and presence of MIX (0.1 mM) and increasing concentrations of PIA (0.025-10,000 nM), the presence of MIX reduced the inhibitory activity of PIA by almost 2 orders of magnitude (IC50 without MIX, 0.5 nM; IC50 with MIX, 20 nM) . Thus, the present study shows that adenosine analogs inhibit agonist-stimulated cAMP formation in cultured Sertoli cells, and that MIX reduces this effect . This indicates that cultured Sertoli cells from immature rats contain A1-receptors for adenosine mediating inhibitory effects on adenylate cyclase. Exp Cell Res, 1985 Aug, 159(2), 313 - 22 Morphological and functional effects of extracellular matrix on pancreatic islet cell cultures; Thivolet CH et al.; Extracellular matrix (ECM) has been reported to enhance epithelial cell attachment and proliferation as well as to induce differentiation in vitro . In an attempt to determine the benefits of culturing pancreatic islet cells on ECM, we studied the morphological and functional patterns of rat islet cells and an insulin-secreting tumor cell line . ECM enhanced islet cell attachment and proliferation when compared to plastic, as suggested by a higher specific activity of DNA synthesis and a higher mitotic index . Cells on ECM were heterogeneous in size and insulin content . They showed extended areas of confluence . Cultures on plastic demonstrated an organisation in clusters and low mitotic activity . However, ECM did not allow for reconstitution of an islet-like structure . When compared to plastic, an initial decrease in basal and stimulated insulin secretion per million cells was observed on ECM, but B-cell activity was restored after 6 days of culture . Glucagon and somatostatin secretion were similar on both substrates . These data suggest that ECM enhances markedly islet cells attachment and proliferation, as well as long-term culture maintenance. Brain Res, 1985 Jul 22, 339(1), 1 - 7 Effects of the synthetic glucocorticoid betamethasone on the survival of neurons in fetal rabbit brain cell culture; Kari B et al.; Dissociated fetal rabbit brain cells were grown on petri dishes coated with collagen . Culture medium consisted of Dulbecco's Modified Eagles Medium plus 10% serum . The mitotic inhibitor 1-beta-D-arabinofuranosylcytosine was added at 6 days for a 2 day period to inhibit over-growth by glial cells and fibroblasts . In some cases cultures were chronically exposed to 0.5, 1.0 or 2.0 microM betamethasone . Examination of cultures by phase microscopy and acetylcholinesterase (AChE) staining demonstrated that cultures incubated with 1.0 and 2.0 microM betamethasone contained 2-2.5 times as many neurons as compared to control cultures . Furthermore, there was an increase in the specific activities of both AChE and choline acetyltransferase (ChAT) which were proportional to the increase in neuronal cell numbers obtained from phase microscopy and AChE staining . These results suggested that betamethasone enhanced survival of cholinergic neurons . Cultures were also examined for neuron specific gamma-aminobutyric acid (GABA) uptake . Again GABA uptake was approximately 2-2.5 times as great in cultures incubated with 1-2 microM betamethasone when compared to controls . Thus, the increase in GABA uptake paralleled the increase in neurons observed by phase microscopy and AChE staining, suggesting that the survival effect of betamethasone was not specific to cholinergic neurons . While betamethasone treated cultures always contained greater numbers of neurons the percentage of neurons lost from all cultures after 2 weeks was the same.(ABSTRACT TRUNCATED AT 250 WORDS) Biochem Biophys Res Commun, 1985 Jul 16, 130(1), 440 - 6 Extracellular matrix (ECM) synthesis in muscle cell cultures: quantitative and qualitative studies during myogenesis; Rao JS et al.; When the synthesis of extracellular matrix components was examined in G8-1 murine skeletal muscle cells as a function of differentiation, non-collagen and to an even greater extent collagen synthesis was increased . Specifically, collagen types I, III, IV, laminin and fibronectin were identified by SDS-PAGE . Immunoprecipitation, with specific antibodies revealed that both the cell layer and medium of differentiated multinucleated myotubes contained increased levels of type IV collagen and laminin, decreased levels of type III collagen and fibronectin and equivalent levels of type I collagen compared to mononuclear myoblasts. Experientia, 1985 Jul 15, 41(7), 930 - 1 Inhibition of virus multiplication and alteration of cyclic AMP level in cell cultures by flavonoids; Mucsi I et al.; The inhibitory effect of four flavonoid compounds on virus multiplication and their influence on the intracellular cyclic AMP (cAMP) level were studied in cell cultures . Quercetin and quercitrin reduced the yields of Human (alpha) herpesvirus 1 (HSV-1) and Suid (alpha) herpesvirus 1 (pseudorabies virus), but hesperidin and rutin had no effect . Further, quercetin and quercitrin elevated the intracellular level of cAMP, whereas hesperidin and rutin did not alter the cAMP level . Both antiviral activity and cAMP-enhancing effect were dependent on the concentrations of the flavonoids, and these effects turned out to be parallel . This study suggests that a relation exists between the antiviral effect and the cAMP-enhancing activity of flavonoids. Avian Dis, 1985 Jul-Sep, 29(3), 768 - 77 Field vaccination against hemorrhagic enteritis of turkeys by a cell-culture live-virus vaccine; Fadly AM et al.; A cell-culture-propagated (CC) live-virus hemorrhagic enteritis (HE) vaccine was evaluated for efficacy and safety in two field trials conducted in North Carolina (NC) and Minnesota (MN) . At 4 or 5 1/2 weeks of age, 9,839 poults in NC and 15,857 poults in MN were vaccinated with a CC HE vaccine administered via the drinking water . A comparable number of poults were maintained as unvaccinated controls . Vaccinated and unvaccinated poults were compared for seroconversion, response to laboratory challenge with a virulent HE virus at 3 weeks postvaccination, livability, percentage graded A, and average weight at marketing . In both trials, vaccination with the CC HE vaccine resulted in immunity against HE as indicated by seroconversion and by resistance to HE lesions following laboratory challenge with virulent HE virus . Compared with unvaccinated groups, vaccinated groups had a significantly higher percentage of turkeys graded A in the NC trial and in two of three flocks in the MN trial (P less than 0.005) . Further, in the NC trial, livability was significantly higher (P less than 0.005) in vaccinated turkeys than in unvaccinated turkeys . These data indicate that the CC HE vaccine is efficacious and safe to use in the field. Tsitol Genet, 1985 Jul-Aug, 19(4), 250 - 4 {Mitotic cycle of a HeLa cell culture exposed to the maximum permissible concentrations of trace elements}; Strochkova LS et al.; It is shown that administration of certain trace elements in maximum allowable concentrations induces changes in metabolism and functionation of cells in the culture . Zinc, nickel, cobalt, cadmium and fluorine are stated to inhibit mitotic activity of HeLa cells by the end of 24 hours of their action . Parallel with this they promote a decrease in H3-thymidine incorporation into DNA . Besides these substances delay various stages of the mitotic cycle for cells. Toxicol Appl Pharmacol, 1985 Jul, 79(3), 490 - 501 Studies on the toxicity of some glycol ethers and alkoxyacetic acids in primary testicular cell cultures; Gray TJ et al.; Primary mixed cultures of Sertoli and germ cells were prepared from testes of immature rats and their response to the known testicular toxicants ethylene glycol monomethyl ether (EGM) and ethylene glycol monoethyl ether (EGE) was studied . Neither EGM nor EGE produced any morphological evidence of toxicity when added to the culture medium at up to 50 mM for 72 hr . In contrast, their metabolites methoxyacetic acid (MAA) and ethoxyacetic acid (EAA) at 2 to 10 mM for 24 to 72 hr caused degeneration of the pachytene and dividing spermatocytes, the target cells of the parent ethers in vivo . As in vivo, earlier spermatocytes, spermatogonia, and Sertoli cells appeared unaffected . EAA was less potent than MAA whereas n-propoxy- and n-butoxyacetic acid, and methoxyacetylglycine, a further metabolite of MAA, produced no morphological changes under these conditions . The same order of toxicity was observed in concurrent studies with the four acids in rats . In culture, the severity of the morphological changes was paralleled by decreases in the activity of carnitine acetyltransferase and lactate dehydrogenase-X in the attached germ cell fraction . Analysis of culture medium provided no evidence for the conversion of EGM to MAA or other metabolites or for the further metabolism of MAA . The close correspondence between the testicular toxicity of alkoxyacetic acids in culture and in vivo suggests a similar mode of action in both cases and points to the potential value of these cultures for mechanistic studies and for screening purposes . The results also emphasize the role of metabolism in the testicular toxicity of glycol ethers and indicate that MAA is an active metabolite of EGM. In Vitro Cell Dev Biol, 1985 Jul, 21(7), 368 - 72 Human smooth muscle cell cultures of the stomach . Morphologic and biochemical studies; Ionasescu R et al.; Smooth muscle cell cultures were prepared from stomach explants obtained surgically from 10 patients with duodenal ulcer . The cultured cells grew in either overlapping layers in "hills and valleys" or in parallel arrays . The ultrastructure studies showed plasmalemmal vesicles, bundles of myofilaments associated with dense bodies, and gap junctions . The synthesis of contractile proteins illustrated the preponderance of actin on myosin and tropomyosin . The synthesis of contractile proteins in stomach smooth muscle cell cultures is significantly higher than in skin fibroblast cultures, i.e . 20 X higher for myosin, 10 X higher for actin, and 30 X higher for tropomyosin. Am J Trop Med Hyg, 1985 Jul, 34(4), 774 - 80 Continuous propagation of Ehrlichia sennetsu in murine macrophage cell cultures; Cole AI et al.; Ehrlichia sennetsu, the etiologic agent of human sennetsu rickettsiosis was successfully propagated in a continuous cell culture using murine cell lines P388D1 and Raw 264 . Pleomorphic cytoplasmic inclusion bodies similar to Ehrlichia canis morulae were observed 3-4 days after second post-inoculation split . In the Raw 264 cell line E . sennetsu was not seen until the third passage . Relatively heavier infection was observed in P388D1 than in Raw cell line . The latter reached a maximum of 15% infection whereas P388D1 cell line attained saturation . Structural details of the organism were confirmed by electron microscopy . A unique rippled cell mass surrounding the plasma membrane was observed . Supernatants of cultures were shown to contain infectious organisms . The advantages of propagating E . sennetsu in continuous cell lines are discussed with respect to future physiochemical and immunochemical studies of this organism. Cancer Res, 1985 Jul, 45(7), 3322 - 31 Changes in stem cell populations of rat tracheal epithelial cell cultures at an early stage in neoplastic progression; Thomassen D et al.; The development of transformed colonies and concomitant changes in proliferative and nonproliferative cell compartments were studied in rat tracheal epithelial (RTE) cell cultures following exposure to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) . Primary RTE cells were plated onto 3T3 feeder layers and treated with MNNG (0.25 micrograms/ml) or solvent . Seven days later, the feeder cells were removed to select for enhanced growth variants, which are the transformants of the RTE cell system, usually scored 5 weeks after carcinogen exposure . Most of the RTE cell colonies, which originally formed during the first 7 days of culture, disappeared within 2 weeks after feeder cell removal in control and MNNG-treated cultures . In control cultures, about 3% of the original colonies persisted, while in MNNG-treated cultures, a larger percentage (approximately 9%) of the colonies persisted . These percentages remained constant from 3 to 7 weeks . Based on colony size, cell density, and cell morphology, the persistent colonies were classified into transformed colonies (large colony size, high cell density, high nuclear:cytoplasmic ratio) and untransformed colonies (small size, low cell density, low nuclear:cytoplasmic ratio) . In the MNNG-treated cultures, about 50% of all persistent colonies showed transformed morphology . Their frequency remained unchanged between 3 and 7 weeks of culture . In contrast, only 10 to 15% of the persistent colonies in control cultures showed transformed morphology at 3 weeks, but that proportion increased steadily between 3 and 7 weeks . These data suggest that, in control cultures, transformed colonies developed spontaneously as a function of time within untransformed colonies . Autoradiographic studies with {3H}thymidine showed that labeling indices in the early "normal" RTE cell colonies between Days 4 and 7 of culture were very high, ranging between 75 and 90% . In contrast, the labeling indices of persistent colonies, both those without and those with transformed morphology, were low, i.e., between 18 and 25%, indicating that a major proportion of cells was either noncycling or cycling very slowly . The relative compartment sizes of cells with stem cell characteristics and of cells with characteristics of transformed stem cells were estimated before and after transformed colonies appeared.(ABSTRACT TRUNCATED AT 400 WORDS) Pediatr Neurol, 1985 Jul-Aug, 1(4), 232 - 7 Differential neurochemical effects of chronic exposure of cerebral cortical cell culture to valproic acid, diazepam, or ethosuximide; Sher PK et al.; We have assessed the relative neurochemical effects of valproic acid, ethosuximide, and diazepam on dissociated cultures of mouse cerebral cortex . Cultures were exposed chronically (11 days) to each antiepileptic drug and assayed for number of neurons, total protein, tetanus toxin fixation, high-affinity uptake of gamma-aminobutyric acid and beta-alanine, choline acetyltransferase activity, and specific and clonazepam-displaceable benzodiazepine binding . Ethosuximide-exposed cultures did not evidence neuronal toxicity; exposure to valproic acid and diazepam resulted in modest neuronal toxicity . However, exposure to each of these drugs resulted in a marked reduction in benzodiazepine binding . This effect may relate to a common mechanism of action of drugs used to treat absence seizures. Endocrinologie, 1985 Jul-Sep, 23(3), 155 - 67 Reactivity of cold thyroid nodule under hormonal influences: study on cell cultures; Ghinea E et al.; The authors have studied the reactivity of cold thyroid nodule cells (CN) by comparison with the reactivity of normal thyroid cells (Ty), hot nodules cells (N), Graves' disease cells (G) and cells from the area proximal to cold nodules (TyC), under the influence of increased amounts of T3, TSH, STH, insulin, estradiol and KI present in the culture medium . The experiment lasted for 9-10 days . The study was carried out on monolayer cultures of cells obtained by tripsinization and on organocultures . Proteic synthesis, cytochemical and cytoenzymatic activities in the culture under the influence of the above substances were examined comparatively . After interruption of the treatment, the monolayer cultures were stained with Giemsa and examined by light microscopy . Proteic synthesis and release of Tg, T3 and T4 into the culture medium, were also examined . The organocultures were used in evaluating 125I uptake from the medium under the influence of the treatment . The results showed variation in relation to the tissue and hormone dose applied . It was especially found that TSH and estradiol cause enlargement of CN cell nuclei and an increased number of mitoses and polyploidia which might be premises for possible malignization. Prostaglandins, 1985 Jul, 30(1), 109 - 18 Hypoxia enhances prostaglandin synthesis in renal mesangial cell cultures; Jelkmann W et al.; In view of recent findings which suggest that renal prostaglandins mediate the effect of hypoxia on erythropoietin production, we have studied whether hypoxia is a stimulus for in vitro prostaglandin synthesis . Studies were carried out in rat renal mesangial cell cultures which produce erythropoietin in an oxygen-dependent manner . Production rates of PGE2 and in specified samples also of 6-keto-PGF1 alpha, as a measure of PGI2, and PGF2 alpha were determined by radioimmunoassay after incubation at either 20% O2 (normoxic) or 2% O2 (hypoxic) in gas permeable dishes for 24 hrs . Considerable variation in PGE2 production was noted among independent cell lines . PGE2 production appeared to be inversely correlated to the cellular density of the cultures . In addition, PGE2 production was enhanced in hypoxic cell cultures . The mean increase was 50 to 60% . PGF2 alpha and 6-keto-PGF1 alpha increased by about the same rate . These results indicate that hypoxia is a stimulus for in vitro prostaglandin production. Cell Biochem Funct, 1985 Jul, 3(3), 179 - 84 Human bone cell cultures: a new model for studying the mechanism of action of calcitonin; Fano G et al.; An investigation on cell cultures obtained from temporal human bone fragments showed that they provide a suitable model for studying the mechanism involved in calcitonin action on bone cells . Furthermore they demonstrated: a transitory increase in 45Ca uptake that returned to control values ten minutes after the hormone was added; a relation between 45Ca uptake and increased cAMP concentrations when these were measured at the same time intervals; a reproduction of the salmon calcitonin (sCT) effect after incubation of the cultures with either db-cAMP or db-cGMP and inhibition of 45Ca uptake and parallel decrease in cAMP levels with propanol . These results suggest that in human bone cell cultures, sCT acts as a temporary promoter of 45Ca uptake, probably by activating an adenylate-cyclase system through a beta-receptor. Avian Dis, 1985 Jul-Sep, 29(3), 672 - 80 Cell-culture virus-neutralization test and enzyme-linked immunosorbent assay for evaluation of immunity in chickens against fowlpox; Buscaglia C et al.; Two serological tests--the virus-neutralization (VN) test in chicken embryo fibroblasts (CEF) using a cell-culture-adapted virus, and the enzyme-linked immunosorbent assay (ELISA)--were used for evaluating the immune response in chickens against fowlpox virus . The VN test was conducted in 96-well tissue-culture plates using a fowlpox virus that was adapted to induce cytopathic effects (CPE) in CEF in 48 hr . The ELISA was carried out with an antigen prepared by precipitation of a cell-culture-propagated virus suspension with ammonium sulfate and concentration by centrifugation . A 0.1 M acetate buffer, pH 5, was used as the sensitizing solution for maximum specific binding of the antigen to the microplate plastic well . No antibodies were detected by the VN test in 228 serum samples taken from chickens at irregular intervals between 1 and 39 weeks of age, even though the birds were vaccinated against fowlpox at 13 weeks of age . However, in sera collected 4 weeks after a sample of laying hens was challenged with fowlpox virus, VN titers of 1/10 to 1/40 were detectable . On the other hand, significant antibody reactions were detected by the ELISA on sera from chickens during the growing period, following vaccination and challenge . Although no maternal antibodies were found at 1 week of age, a continuous increase in the mean ELISA titers to fowlpox was demonstrated during the entire experimental period . This study showed that the ELISA was considerably more sensitive and practical than the VN test. Mol Cell Endocrinol, 1985 Jul, 41(2-3), 129 - 36 Electrofocusing fractionation of follicle stimulating hormone in pituitary cell culture extracts from male and female rats; Foulds LM et al.; Three-day pituitary cell cultures from adult male and female rats were incubated for 4 h in the presence of 10 nM LHRH and the molecular heterogeneity of FSH was assessed in the media of LHRH-stimulated cells and in cell extracts from unstimulated cells using an electrofocusing technique . The pI distribution of FSH showed a high degree of similarity between cell media and cell extracts of each sex although differences were observed between sexes . Pituitary cell cultures from male rats were also incubated in the presence of 10(-8) M testosterone and 10(-8) M estradiol and the pI distribution of FSH from media after LHRH stimulation was determined . No significant differences in the pI profiles were observed . Incubation with charcoal-treated bovine follicular fluid (an inhibin source) resulted in a significant reduction in recovered FSH activity in the pH region 3.61-3.92 although this decrease did not markedly alter the pI profile of FSH . Close similarities were observed in the pI distribution of FSH of pituitary cell culture extracts and pituitary gland extracts from intact animals of both sexes, however, differences in pI distribution were noted in pituitary extracts in the male but not the female following gonadectomy . It is concluded that (1) stored FSH is released from the pituitary without major modification to its structure as assessed from its pI profile, (2) sex differences in the pI profile of FSH in pituitary extracts are retained in culture and following LHRH-stimulated release, (3) the pI distribution of FSH is not affected by testosterone or estradiol and only minimally by inhibin in short-term cultures.(ABSTRACT TRUNCATED AT 250 WORDS) Acta Virol, 1985 Jul, 29(4), 312 - 7 Ecology of the murine alphaherpesvirus and its isolation from lungs of rodents in cell culture; Mistrikova J et al.; Together 381 sera of murine rodents (Apodemus flavicollis, Clethrionomys glareolus, Microtus arvalis and Mus musculus) trapped in different localities of Czechoslovakia were examined for the presence of antibodies to a murine alphaherpesvirus (MHV) and to murine cytomegalovirus (CMV) . Positivity of rodent sera to the two MHV and one murine CMV strains varied from none to 12.5% and from none up to 11.3%, respectively, depending on the locality under study . From the lungs of A . flavicollis showing serum antibodies to MHV, a further murine herpes virus strain was isolated in rabbit embryo fibroblasts (REF) . The latter MHV reached a titre of 10(10) TCID50/ml in the 13th passage, causing typical cytopathic effect from the 2nd passage on. Biosci Rep, 1985 Jul, 5(7), 601 - 8 Developmental study on the regulation of neurotransmitter-sensitive adenylate cyclase systems in primary cerebral cell cultures from embryonic mice; Shanker G et al.; An ontogenetic study of the effect of various neurohormones and other activators on adenylate cyclase systems was carried out using cultures of cells from 15-d-old embryonic mouse brain . Dopamine stimulated the enzyme activity at earlier culture ages (i.e . 4 and 10 d) but had little stimulatory effect at later ages (i.e . 20 and 33 d) . Further, this stimulation at the earlier ages was blocked by the dopaminergic blocker, fluphenazine, but not by alpha and beta-adrenergic antagonists . In contrast to dopamine, isoproterenol (a beta-adrenergic agonist) had little stimulatory effect at earlier ages, but its ability to stimulate cyclase activity increased with age . This increase in all age groups was blocked by propranolol (a beta-adrenergic antagonist) . Epinephrine-sensitive enzyme activity showed a steady increase with age, which could be blocked with propranolol except in 4-d-old cultures, where it was blocked instead by fluphenazine . Because the cultures are relatively enriched in neurons at earlier ages and in glia in later ages, the results suggest a predominantly neuronal localization for the dopamine sensitive adenylate cyclases and a glial localization of the isoproterenol and epinephrine sensitive adenylate cyclases . Histamine, serotonin, calcium/calmodulin and chloroadenosine were either only slightly or not at all stimulatory. Am J Pathol, 1985 Jul, 120(1), 67 - 78 Characterization of a primary bile ductular cell culture from the livers of rats during extrahepatic cholestasis; Sirica AE et al.; The establishment of novel bile ductular cell cultures was accomplished with the use of explants of a hyperplastic bile ductular tissue preparation obtained from rat livers at 10 to 15 weeks after bile duct ligation or a bile ductular cell fraction isolated from this tissue preparation by a procedure involving Percoll density gradient centrifugation . Observations made on these primary explant and monolayer bile ductular cell cultures were limited to the first 3 days of culture where the morphologic features of the bile ductular epithelium remained fairly well preserved, while fibroblast contamination was found to be very low . These cultured cells also retained over this period a high specific activity for the bile ductular cell marker enzyme gamma-glutamyl transpeptidase, as well as possessed measurable but decreasing specific activities for leucine aminopeptidase and alkaline phosphatase . Karyotypic analysis of the cultured monolayer cells further showed them to be diploid . In addition, preliminary transplantation studies demonstrated the presence of well-differentiated bile ductular-like structures following inoculation of the freshly isolated bile ductular cell fraction into the interscapular fat pads of recipient rats. J Biomed Mater Res, 1985 Jul-Aug, 19(6), 653 - 61 A modification of the cell culture agar diffusion test using fluoresceindiacetate staining; Schmalz G et al.; In order to reduce the uncertainties involved in the morphologic evaluation of the cell culture agar diffusion test, the originally recommended neutral red stain was replaced by fluoresceindiacetate (FDA) . This stain proved to be nontoxic in the concentrations used in this study (0.002%) . Toxicity tests with phenol, formalin, and methylmethacrylate monomer revealed results corresponding to literature evaluations by other methods . Since FDA makes cell metabolism visible and easily accessible to automated evaluation techniques, it presents certain advantages over the morphologic evaluation using the neutral red stain. Diagn Microbiol Infect Dis, 1985 Jul, 3(4), 353 - 8 Immunoenzymatic staining of viral and chlamydial antigens in cell culture; Richman DD et al.; Methods are described for the conjugation of antibodies with biotin and for the use of these reagents in an immunoperoxidase staining procedure for infected cell cultures . This technique provides a simple, rapid, and specific approach to the identification and characterization of a number of viral and chlamydial isolates in the diagnostic laboratory. J Cardiovasc Pharmacol, 1985 Jul-Aug, 7(4), 799 - 804 Comparative metabolic effects of halothane and enflurane in rat heart cell culture; Albrecht RF et al.; The effects of halothane and enflurane on the oxygen consumption rates and substrate utilization by beating and nonbeating rat heart myocytes in cell culture were compared . Halothane, on an equal dose and equal MAC (minimum alveolar concentration producing immobilization of 50% of subjects) basis, was significantly more effective than enflurane in reducing total myocyte oxygen consumption and contractile rate . The greater effect of halothane on oxygen consumption was not due entirely to its effect on myocyte contractile rate, since quiescent (nonbeating) cells and cells rendered nonbeating by large doses of halothane also showed greater reductions in oxygen consumption than with large doses of enflurane . Both halothane and e