J Immunol, 1981 Feb, 126(2), 540 - 4 Fate of a common acute lymphoblastic leukemia antigen during modulation by monoclonal antibody; Pesando JM et al.; Modulation of a human common acute lymphoblastic leukemia antigen (CALLA) by specific monoclonal antibody (J5) has been studied with the immune precipitation method to identify radiolabeled antigen . Surfaces of leukemic cells have been labeled using 125I both before and after modulation by J5 antibody for different time intervals . Leukemic cells have also been metabolically labeled with 35S-methionine before modulation . These studies indicate that the 100,000-dalton glycoprotein expressing CALLA (gp 100-CALLA) cannot be detected in cells that were modulated with J5 antibody before surface labeling but that it is easily detectable in cells that were surface labeled before modulation for 10 hr . At later time points, gp 100-CALLA is selectively lost from cells that were surface labeled before modulation . Gp 100-CALLA is not detected in the supernatants from cultures of these modulated cells . We conclude that gp 100-CALLA is rapidly internalized during modulation and that CALLA is degraded . Gp 100-CALLA is not shed into the culture media, nor does it remain on the cell surface in an altered form . Incubation of leukemic cells with antisera to beta2-microglobulin or IgM does not affect the expression of gp 100-CALLA. Infect Immun, 1981 Feb, 31(2), 704 - 11 Effect of herpes simplex virus infection on murine antibody-dependent cellular cytotoxicity and natural killer cytotoxicity; Kohl S et al.; Mice intraperitoneally inoculated with a sublethal dose of herpes simplex virus (HSV) produced immunoglobulin G antibody-dependent cellular cytotoxicity (ADCC) and radioimmunoassay (RIA) antibody as early as 3 days after infection . There was a rise in natural killer cytotoxicity (NKC) to infected and uninfected target cells 1 to 3 days postinfection mediated by nonadherent peritoneal cells (PC) in mice inoculated with HSV, but also with other substances commonly used in tissue culture media . HSV caused the highest and most consistent increase in NKC . PC-NKC, as ADCC, was inhibited by latex and silica, both macrophage inhibitors . PC-ADCC markedly declined 3 to 8 days after HSV inoculation . This was not due to a soluble or cellular suppressor factor, was not reversed by incubation or trypsin treatment of PC, was not associated with a change in PC Fc receptors, adherence, or acridine orange staining characteristics, and could not be induced by inactivated HSV . In vitro inoculation of PC with HSV similarly caused a reduction in the ability of PC to mediate ADCC to HSV-infected target cells . These data demonstrate the complex stimulatory and inhibitory interactions between virus and host defense mechanisms. Chromosoma, 1981, 84(3), 365 - 72 Cell-specific variation of X chromosome replication in the Syrian hamster (Mesocricetus auratus); Cawood AH; The sub-division of S-phase in Syrian hamsters, on the basis of Brd/U/Hoechst 33258/Giemsa banding, has allowed a quantitative comparison of the replication of individual chromosome bands within defined subphases of S . This analysis has shown that in hamsters, as has been reported in humans, there are distinct patterns of early replication in vitro in the early X, the late X in fibroblasts, and the late X in lymphocytes . In addition, it has been possible to show that, although the pattern of replication of the late X in fibroblasts differs from that in lymphocytes, the time in S at which bands first appear on this chromosome is the same in the two cell types . No significant heterogeneity can be ascribed to differences between individuals, adult or embryonic sources, culture media, or time of exposure to BrdU . The absence of any detectable heterogeneity in the replication band frequencies in autosomal heterochromatic arms suggests that the cell-specific variability of the late-replicating X is a feature of facultative rather than constitutive heterochromatin. Histochemistry, 1981, 73(1), 137 - 50 {The histochemistry of human endosalpingeal ciliated cells (author's transl)}; Kugler P; The human endosalpingeal ciliated cells are not a homogeneous cell population . They can be distinguished into mitochondria prominent and normal cells . The morphological appearance of ciliated cells was studied in organ culture using different hormones and hormone combinations in the culture media . Histological (HE, semithin sections) and histochemical methods (PAS, Alcian Blue, SDH, LDH, ATPase, 5'-Nucleotidase, acid and alkaline Phosphatase) were applied . Those mitochondria prominent ciliated cells which are seen in the native endosalpinx can in vitro also be determined mainly under the influence of steroid hormones (hydrocortisone and progesterone) . In hormone free incubation media the apices of ciliated cells are separated . This will happen in hormone containing media, too, but with delay, however . Some results are in agreement with the possible transformation from ciliated cells to secretory cells. Folia Microbiol (Praha), 1981, 26(5), 408 - 12 Production and purification of Penicillium citreoviride toxin and its effect on TPP-dependent liver transketolase; Datta SC et al.; The production isolation and purification of a yellow mycotoxin from Penicillium citreoviride NRRL 2579 in different culture media was described . When injected subcutaneously to albino rats it alters the kinetic pattern of transketolase (EC 2.2.1.1) in liver in vivo in a competitive manner . In vitro, the inhibition is noncompetitive in nature . However, addition of thiamine diphosphate (TPP) to the in vitro system relieved the inhibitory effect . These findings suggested a relationship between citreoviridin-induced beriberi and the probable antithiamine effect of the toxin. J Immunol Methods, 1981, 44(1), 3 - 14 Human lymphocyte transformation induced by mitogens and antigens in a serum-free tissue culture system; Needleman BW et al.; The use of serum to supplement lymphocyte tissue culture media introduces uncontrolled variables; different serum sources, lots and concentrations can produce variability in experimental results, serum can stimulate or inhibit lymphocytes, and components of serum can react with substances whose effects on lymphocytes are being studied . To avoid these problems, we studied the ability of human peripheral mononuclear cells to survive and to respond to stimulation in an entirely synthetic medium, RPMI-1640 supplemented with L-glutamine, gentamicin, HEPES buffer and magnesium . Optimal cell concentration in this serum-free RPMI-1640 was 2.5 x 10(6) cells/ml, whereas optimal cell concentration in serum containing RPMI-1640 was 1 x 10(6) cells/ml . In this serum-free RPMI-1640, 50% of the cellular input was recovered as viable cells after 7 days of culture, which was similar to results in serum containing RPMI-1640 . Mononuclear cell transformation transformation was induced by phytohemagglutinin, concanavalin A, pokeweed mitogen, streptolysin O and candida . Optimal doses of stimulants and the kinetics of the responses were similar in serum-free and serum containing RPMI-1640 . This system can be used to avoid the problems inherent in systems which supplement tissue culture media with serum. Arch Virol, 1981, 67(1), 31 - 43 The detection of influenza A virus antigens in cultured cells by enzyme-linked immunosorbent assay; Watanabe H et al.; An enzyme-linked immunosorbent assay (ELISA) was employed to investigate the expression of influenza A/Hong Kong/68 (H3N3) virus structural proteins on the surface of infected MDCK cells, and to detect viral antigens in culture media and cell extracts . Infected cells were fixed with 0.1 per cent glutaraldehyde before being examined for the presence of cell-surface antigens . Viral antigens were first observed on the surface of cells 4 hours after infection and reached a maximum 10-12 hours after infection, when measured by haemadsorption with chicken erythrocytes and by ELISA and immunofluorescence with hyperimmune antiserum to Hong Kong virus . A good correlation was found between the three assay systems . The presence of individual virion structural proteins on the cell surface was determined by ELISA using specific antibodies purified by differential affinity chromatography . Either or both or the internal matrix and nucleoprotein antigens were expressed from 2 to 6 hours after infection, with maximum expression after 2 hours, and the strain-specific and common antigenic determinants of haemagglutinin were observed on the cell surface from 4 hours after infection, and reached a maximum 8 to 10 hours after infection . Low levels of neuraminidase were detected between 4 and 8 hours after infection . Culture media and cell extracts were titrated by infectivity and haemagglutination assays, and by ELISA . Titres obtained from the culture media showed a close correlation between the three assay methods, with peak titres being attained 24 hours after infection . Viral antigens were first observed in cell extracts by ELISA 4 hours after infection, and infectious virions and haemagglutinin 2 hours later, but whereas maximum titres of infectious virus and haemagglutinin were found 10 hours after infection, the ELISA titre continued to rise until 24 hours after infection, which suggested that virus structural proteins were being accumulated in the cells after most of the progeny virions had been released . The results are discussed in terms of the potential use of ELISA in rapid virus diagnosis. Med Interne, 1981 Jan-Mar, 19(1), 73 - 7 The relationship between air-borne fungal spores and Dermatophagoides pteronyssinus in the house dust; Chirila M et al.; The prevalence of mould genera and their association with the house dust mite (Dermatophagoides pteronyssinus) were investigated in the homes of 192 asthmatic patients and 125 healthy people in a Romanian town with a relatively high degree of humidity . The dominant moulds which developed in the culture media exposed on the bedroom floors were Cladosporium, Penicillium and Aspergillus . Living Dermatophagoides pt . and their eggs were present mostly in the colonies of Cladosporium; in those of Penicillium the house dust mites were dead and disintegrated . It is concluded that allergy to Dermatophagoides pt . must not be connected to house dust as such but to the presence in the dust of certain mould spores. Cancer Chemother Pharmacol, 1981, 6(3), 205 - 10 Methodologic problems in clonogenic assays of spontaneous human tumors; MacKintosh FR et al.; Colony formation in soft agar was used to investigate growth properties and drug sensitivity in 102 tumor specimens from 91 patients . Sufficient colony growth for sensitivity testing with various drugs was obtained in 36 of 67 specimens (54%) with adequate cell yield and pathologically documented malignancy . Room temperature (20-24 degrees C) is superior to both 4 degrees C and 37 degrees C for 12-36 h storage and transport of malignant effusions . By contrast, fine mincing in sterile saline or balanced salt solution, and refrigerated storage (4 degrees C) appear optimal in experiments with three solid tumors . The use of buffered NH4Cl to lyse red blood cells markedly reduced plating efficiencies, and also reduced the percentage of tumors in which drug sensitivities could be tested from 64% to 38% . Several combinations of potential growth factors and culture media have been tested . Insulin enhanced plating efficiency (PE) in all six adenocarcinomas tested . Drug sensitivity of tumors was not affected by varying plating efficiency up to five-fold in two tumors . In eleven cases tumor cells were exposed to combinations of two or more drugs, and results assessed for evidence of drug interactions . In almost all cases, these two-drug combinations produced additive cell killing than either antagonistic or greater-than-additive effects.U Clin Orthop, 1981 Jan-Feb, (154), 234 - 48 Bone-forming and bone-resorbing cell lines derived from bone marrow in tissue culture; Hirano H et al.; The differentiation of connective tissue outgrowths of adult bone marrow in response to the organic matrix of bone was observed in tissue culture by correlated histologic, electron microscopic, biochemical, and radioisotope-labeling methods . On a substratum of bone matrix gelatin, myelogenous cells degenerate and disappear while stromal and perivascular cells proliferate and differentiate into mesenchymal-type, monocytoid, and giant cells . From primary cultures of bone marrow cells on bone matrix, cartilage differentiates in isolated areas but only in small islets . With continuous subculture through 25 generations, the proportions of two functionally different populations of chondrogenetic and matrix-resorbing cells gradually emerge . Up to the time of the ninth generation of subculture, the chondrogenetic population predominates . After the 14th to the 25th generation, clones of matrix-resorbing large monocytoid cells predominate and rapidly digest the matrix substratum . Measurements of 35S uptake demonstrate that control cultures of muscle-derived mesenchymal-type cells produce about twice as much cartilage as marrow-derived mesenchymal-type cells . A decline in the chondrogenetic cell population and corresponding rise in the matrix-resorbing cell population is demonstrable by a progressive increase in the quantity of hydroxyproline-containing peptides in the culture medium . This decline is not attributable to conditions in culture because there was progressive loss of chondrogenetic activity of the eighth and 20th passage even when the cells were transplanted in diffusion chambers back into an isologous host . The problem is how to account for the competence of marrow stromal cells to differentiate into bone without bone matrix in vivo but not in vitro . Mixed cultures of muscle and marrow outgrowths produce only half as much cartilage (measured by 36S-uptake/microgramDNA in the system) as muscle outgrowths alone . These observations suggest that a bone marrow-derived matrix-resorbing cell population, by some unknown mechanism, inhibits proliferation of cartilage-bone precursor cell populations . The nature of the inhibition requires investigation by detailed biochemical analyses of marrow cell culture media and chemical extracts of whole bone marrow tissue. J Cancer Res Clin Oncol, 1981, 102(1), 49 - 55 Human erythroid cell lines derived from a patient with acute erythremia; Okano H et al.; Three continuous human cell lines, designated KMOE, derived from a patient with acute erythremia (Di Guglielmo's disease) are reported . The cell lines are the cultures of (1) bone marrow cells, (2) peripheral blood cells, and (3) cells from a tumor developed into an athymic nude mouse after transplantation of the cultured bone marrow cells . Cells of all three lines show morphology of immature erythroblast and have i(17q) marker chromosome . They are negative for both Philadelphia chromosome and Epstein-Barr virus nuclear antigen . Although all KMOE cells in suspension culture are benzidine-negative, benzidine-positive cells are found within colonies formed in semi-solid culture media . The relative number of colonies with benzidine-positive cells is increased when sodium butyrate is added to the culture. Pharmacol Ther Dent, 1981, 6(1-2), 35 - 43 The effect of dental plaque grown in the presence of xylitol or sucrose on bone resorption in vitro; Tenovuo J et al.; Bone culture was used as an experimental model in studying the ability of dental plaque grown in the presence of xylitol or sucrose to induce bone resorption . Plaque samples were collected in young adults after six or ten days with no oral hygiene and with frequent use of xylitol- or sucrose-sweetened chewing gum . The rate of resorption was assayed by measuring the release of both acid phosphatase and 45Ca from bones into culture media during a three-day incubation period . Sucrose-induced plaque collected after ten days increased both of these indicators of bone resorption, while xylitol plaque decreased or had no effect under identical conditions . Xylitol consumption induced a marked increase in acid phosphatase activity of dental plaque - a phenomenon which would appear to be unrelated to bone resorption . The results suggest that the inflammatory potential of dental plaque may be reduced during xylitol consumption as compared to sucrose consumption. Proc Natl Acad Sci U S A, 1981 Jan, 78(1), 162 - 6 Isolation of functional human coagulation factor V by using a hybridoma antibody; Katzmann JA et al.; Spleen cells obtained from mice immunized with partially purified human coagulation Factor V were fused with NS-1 mouse myeloma cells, and hybrids were selected . Culture media were screened for anti-Factor V activity, and an antibody-positive clone was obtained and passaged as an ascites tumor in mice . The ascitic fluid from the hybridoma-bearing mouse could be diluted 1:10(6) before losing reactivity in an anti-Factor V radioimmunoassay . When immobilized on agarose, the monoclonal antibody quantitatively removed Factor V activity from human plasma . Factor V activity could be eluted with 1.2 M NaCl at pH 6.5 . Homogeneous Factor V was isolated by chromatography of barium citrate-adsorbed, polyethylene glycol 6000 precipitated plasma on the antibody column followed by chromatography on phenyl-Sepharose . The isolated Factor V exhibited a single band upon gel electrophoresis in sodium dodecyl sulfate with an apparent Mr comparable to that of bovine Factor V (330,000) . Upon exposure to thrombin, the activity of Factor V increased 53-fold when measured in Factor V-deficient plasma . This increased activity was associated with discrete proteolytic cleavages of the parent molecule. Acta Physiol Lat Am, 1981, 31(3), 199 - 206 Immunogenicity of Trypanosoma cruzi in different animal species; Rubiolo ER; The immunogenicity of two fractions (1 500 F and 10 000 F) from epimastigotes of Trypanosoma cruzi as well as the supernatant from culture media (SF) were studied using hens, rabbits and opossums . For comparative purposes, sera from individuals with chronic Chagas' disease were also used . A similar, positive response was obtained for the fractions in all the animal species studied using indirect hemagglutination test . Supernatants from culture media were the least immunogenic . By double immunodiffusion test, it was possible to detect a positive response to a different number as well as to different antigens in the three animal species, but there was response to a common antigen by all the different animal species . The common antigen called here major, was present in all the fractions assayed . Human sera from individuals chronically infected showed a variable response . When assayed by double immunodiffusion technique, the major antigen could be detected in just a few samples. Mol Cell Endocrinol, 1981 Jan, 21(1), 43 - 9 Inhibitory effect of gonadotropin releasing hormone upon cultured testicular cells; Hsueh AJ et al.; The direct inhibitory of gonadotropin-releasing hormone (GnRH) upon testicular androgen production was studied in cultured testicular cells . Enzyme-dispersed testicular cells from immature hypophysectomized rats were cultured for 6 days in a serum-free medium with or without various hormones . Culture media were changed every 2 days and media concentration of a androgens was measured by radioimmunoassay . The testis cultures from immature rats secrete predominately 5 alpha-androstane-3 alpha, 17 beta-diol (A-diol) and androsterone but negligible amounts of androstenedione, testosterone and dihydrotestosterone . Incubation of testicular cells with hCG and FSH increased A-diol and androsterone production as compared to control cultures . In contrast, treatment with GnRH or a GnRH agonist (des-Gly10, D-Leu6 (N alpha Me)Leu7, Pro9NHEt-GnRH) inhibited the gonadotropin effect in a dose-dependent manner; 10(-8) M GnRH inhibited A-diol production by approximately 20% on day 4 and 6 of culture whereas 10(-6) M GnRH inhibited A-diol production by greater than 80% . The observed effect of GnRH upon testicular cells in primary culture demonstrates that the hypothalamic peptide directly inhibits testicular steroidogenesis. Connect Tissue Res, 1981, 9(1), 19 - 24 Connective tissue activating macromolecules in macrophage culture medium; Jalkanen M; Fractions isolated by isoelectric focusing (pI values 4.6-5.3) from macrophage culture media stimulated fibroblast proliferation in vitro and increased the synthesis of sulfated glycosaminoglycans (sGAGs) . Cationic fractions from macrophage culture media isolated by the same procedure (pI 10.2-10.4) enhanced secretion of collagen by fibroblasts . Two dimensional electrophoresis of non-fractionated 3H-lysine labeled macrophage media only showed incorporation of label into acidic proteins . The fractions from the pI area which increased DNA and GAG synthesis by fibroblast cultures contained three major labeled proteins with molecular weights of 15 K, 25 K and 135 K. Rheumatol Int, 1981, 1(1), 11 - 6 The production in culture of metalloproteinases and an inhibitor by joint tissues from normal rabbits, and from rabbits with a model arthritis . I . Synovium; Cambray GJ et al.; During 5 days of culture, explants of normal rabbit synovium produced no active collagenase, negligible latent collagenase, but significant levels of free collagenase inhibitor . Synovium from joints exhibiting a proliferative arthritis produced greatly elevated levels of collagenase; the appearance of active enzyme in the medium during the second day of culture was associated with the disappearance of free inhibitor . Enzyme levels in the media correlated well with the arthritic status of joints, when explants were prepared up to 10 weeks after the induction of the model arthritis . Synovium from the contralateral joints of rabbits with unilaterally induced arthritis produced no active collagenase, but approximately one-third as much latent collagenase as found with arthritic joints . Enzymatic activities against gelatin and cartilage proteoglycan substrates were demonstrated in synovial culture media in addition to collagenolytic activity . Gel filtration showed that these activities were not due to a single enzyme, and further characterisation confirmed that the enzymes were metalloproteinases . The results are considered in the light of published data, and the involvement of metalloproteinases and their specific inhibitor in the development of arthritic lesions is discussed. Biochim Biophys Acta, 1980 Dec 15, 633(3), 410 - 21 Effect of RNA and protein synthesis inhibitors on the release of inflammatory mediators by macrophages responding to phorbol myristate acetate; Bonney RJ et al.; The interaction of phorbol myristate acetate with resident populations of mouse peritoneal macrophages causes an increased release of arachidonic acid followed by increased synthesis and secretion of prostaglandin E2 and 6-keto-prostaglandin F1 alpha . In addition, phorbol myristate acetate causes the selective release of lysosomal acid hydrolases from resident and elicited macrophages . These effects of phorbol myristate acetate on macrophages do not cause lactate dehydrogenase to leak into the culture media . The phorbol myristate acetate-induced release of arachidonic acid and increased synthesis and secretion of prostaglandins by macrophages can be inhibited by RNA and protein synthesis inhibitors, whereas the release of lysosomal hydrolases is unaffected . 0.1 microgram/ml actinomycin D blocked the increased prostaglandin production due to this inflammatory agent by more than 80%, and 3 microgram/ml cycloheximide blocked prostaglandin production by 78% . Similar results with these metabolic inhibitors were found with another stimulator of prostaglandin production, zymosan . However, these inhibitors do not interfere with lysosomal hydrolase releases caused by zymosan or phorbol myristate acetate . It appears that one of the results of the interaction of macrophages with inflammatory stimuli is the synthesis of a rapidly turning-over protein which regulates the production of prostaglandins . It is also clear that the secretion of prostaglandins and lysosomal hydrolases are independently regulated. Biochim Biophys Acta, 1980 Dec 5, 620(3), 418 - 28 Cyclooxygenase products of arachidonic acid metabolism by mouse bone in organ culture; Voelkel EF et al.; The products of endogenous and exogenous arachidonic acid metabolism via the cyclooxygenase pathway in mouse bone in organ culture were identified and quantitated by the use of high performance liquid chromatography and radioimmunoassay . Production of prostaglandins E2, F2 alpha, and I2 from endogenous substrate was stimulated by incubation of bone with epidermal growth factor and the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate . Addition of arachidonic acid to the culture medium not only resulted in the accumulation of prostaglandins E2, F2 alpha, and I2 but also thromboxane . Bone metabolized prostaglandins E2 and F2 alpha to their respective 13,14-dihydro-15-keto-derivatives . Prostaglandin I2, measured as 6-keto-prostaglandin F1 alpha was synthesized by bone, and metabolic products of prostaglandin I2 or 6-keto-prostaglandin F1 alpha, either 6,15-diketo-prostaglandin F1 alpha or 13,14-dihydro-6,15-diketo-prostaglandin F1 alpha, were also detected in the culture media . Formation of cyclooxygenase products of endogenous and exogenous arachidonic acid metabolism (both basal and stimulated) and bone resorption were inhibited by indomethacin . Bone as a tissue responded biochemically not only to exogenous prostaglandins and agents that enhance endogenous prostaglandin production but also to exogenous arachidonic acid by biosynthesis of prostaglandins, prostacyclin and thromboxane . Furthermore, bone metabolized these cyclooxygenase products to their more stable metabolites. Early Hum Dev, 1980 Dec, 4(4), 337 - 45 Human amniotic fluid contains fibroblast-pneumonocyte factor; Seybold WD et al.; We have studied 41 samples of human amniotic fluid collected at 27-40 wk gestation . After 'stripping' with Florisil, most fluids studied stimulated {3H} choline incorporation into saturated phosphatidylcholine by human fetal alveolar Type II cells . The activity declined with advancing gestation and was inversely related to cell number following incubation . Following acid-ethanol extraction, activity eluted from Sephadex G75 columns in 1 M acetic acid with a Kd of approximately equal to 0.4-0.6, and was greater at earlier gestations (27-31 wk) than at later periods (32-36 or 37-40 wk) . In contrast, chromatography at neutral pH without prior extraction resulted in retarded elution . The active material was shown to be heat-stable . Thus, the biochemical features of activity recovered from human amniotic fluid are identical to fibroblast-pneumonocyte factor recovered from supernatant culture media of cortisol-treated human fetal lung fibroblasts. Jpn J Exp Med, 1980 Dec, 50(6), 407 - 14 Analyses of some mite antigens found in house dust and in food material; Yago A et al.; Antigen analyses by micro-Ouchterlony, immunoelectrophoresis and passive hemagglutination tests were carried out on extracts from each of the following cultured mites: Dermatophogoides farinae, Dermatophogoides pteronyssinus, Tyrophagus putrescentiae, Aleuroglyphus ovatus and Carpoglyphus lactis; and on an extract from the culture medium for each mite . Antisera were raised against each extract or each medium in rabbits or in guinea pigs . Up to 5 precipitin lines by micro-Ouchterlony test and up to 8 precipitin arcs by immunoelectrophoresis were detected between an extract and its homologous (corresponding) antiserum . Some of the lines or arcs were seen consistently to be common to all species tested . The absorption of common antigens in an extract by heterologous (noncorresponding) antiserum was employed to find the antigens specific to D . farinae and D . pteronyssinus . By these tests, at least 3 species-specific antigens for D . farinae and 2 for D . pteronyssinus were found . In passive hemagglutination tests, higher titers were obtained mainly in homologous systems, although weak cross reaction was observed in every system . Culture media did not participate in any reaction combinations used here and did not interfere with the analyses. Prostaglandins, 1980 Dec, 20(6), 1075 - 87 The prostaglandins of articular cartilage . I . Correlates of prostaglandin activity in a chondrocyte culture system; Copeland M et al.; Suspensions of aggregated chondrocytes display active prostaglandin (PG) production . Radioimmunoassay of culture media and thin layer chromatographic analysis suggests that PGE2 is the primary PG synthesized . In order of decreasing concentration, the following PG were tentatively identified; PGE greater than PGI greater than PGA + PGB greater than or equal to PGF1+2 greater than TxB . An inverse logarithmic relationship was identified between PG synthesis and cells cultured at densities of 1.5 to 7.5 x 10(6) cells/ml . Little or no change in the PG distribution profile was seen at these high cell densities . Maximum PG synthesis was attained after 36 hours of incubation with persistence of high synthetic levels up to 48 hours . PGE2 production measured at various post-isolation intervals indicated an initial high rate of synthesis during the first 4 hours which decreased with time up to 24 hours . Cartilage explant organ cultures demonstrated a similar level of PG synthesis suggesting minimal effect of matrix on cellular PG production . Indomethacin (5 microgram/ml) inhibited PG synthesis by 70% within 4 hours and 85% after 24 hours of exposure . Arachidonic acid supplementation (10 microM) stimulated PG synthesis by 300%. J Clin Endocrinol Metab, 1980 Dec, 51(6), 1432 - 6 Pregnancy-specific beta-1-glycoprotein (SP1) in cultured amniotic fluid cells; Heikinheimo M et al.; The synthesis of pregnancy-specific beta-1-glycoprotein (SP1) was studied in amniotic fluid cell cultures using RIA, immunoperoxidase, and immunofluorescence techniques . SP1 was found by RIA in all 11 sonicates and in 21 of 26 culture media . The SP1-immunoreactive material was immunologically similar to maternal serum SP1 . Immunoperoxidase and indirect immunofluorescence staining were positive in large cells identified as epithelial amniotic cells by labeling with antikeratin antibodies . Fibroblast-like cells were occasionally found in cultures, but they did not contain demonstrable amounts of SP1 . The physiological significance of the findings presented remains unclear. Cancer Res, 1980 Dec, 40(12), 4722 - 7 Specific effects of fibronectin-releasing peptides on the extracellular matrices of cultured human fibroblasts; Keski-Oja J et al.; Fibronectin-releasing activity has recently been found in concentrated serum-free culture media of the established human fibrosarcoma cell line, 8387 . We used these M.W . 10,000 fibronectin-releasing polypeptides (10K peptides) to study their effects on the cell-free matrices of cultured diploid human lung fibroblasts . Metabolically labeled cultures of fibroblasts were extracted with sodium deoxycholate and hypotonic buffer to prepare the matrices . The isolated matrices contained fibronectin and procollagen as their major radiolabeled proteins, and some as yet unidentified polypeptides were also detected . the matrices that were attached on the culture dishes were exposed to increasing concentrations of the 10K peptides in serum-free medium, and the changes in the radiolabeled polypeptides were studied . As a result of this treatment, there was a massive release of both fibronectin and procollagen from the matrices . No major cleavages of either released protein could be seen . After digestion of the matrix-associated collagen with collagenase, the 10K peptides released only fibronectin . Collagenase treatment, on the contrary, did not affect matrix-associated fibronectin . On the other hand, a M.W . 66,000 matrix-associated protein was constantly cleaved to a M.W . 62,000 form that remained in the matrix as a result of the incubation of the matrices with the 10K peptides . The 10K peptides did not affect the other radiolabeled polypeptides present in the matrix . the results indicate that the fibronectin-releasing peptide behaves as a specific protease on the matrices of cultured human fibroblasts. Br J Vener Dis, 1980 Dec, 56(6), 390 - 3 Assessment of transport and isolation methods for gonococci; Taylor E et al.; Urethral discharge from men was diluted to give heavy and light inocula and cultured on seven different solid culture media, including two transport/isolation media, or held in three types of semi-solid transport medium for varying periods and then cultured . The amount of growth was quantitated and the performance of the different systems compared . Fresh non-selective media were best, with up to two failures in 254 cultures on each medium . With selective media there were 9-23 failures with heavy inocula and 22-47 failures with dilute inocula . For Transgrow or the Jembec system incubation before holding at ambient temperature was better than holding followed by incubation . Transport media yielded good results if cultures were set up within six hours; only minor losses occurred after 24 hours. Cancer Lett, 1980 Dec, 11(2), 81 - 7 Formation of metastasis by human breast carcinoma cells (MCF-7) in nude mice; Shafie SM et al.; MCF-7 cells, a human breast carcinoma line, forms tumors when injected into athymic nude mice . These tumors are able to metastasize to lungs, liver and spleen . 17 beta-estradiol treatment increases both the growth rate and frequency of metastases . Castration or diabetes prevents metastasis formation, but treatment with estrogen or insulin restores the metastasizing capacity . MCF-7 cells secrete into the culture media collagenases able to lyse types I and IV collagens . Estrogen or insulin addition to the culture enhances collagenase production . Attention is called to the coexistence of enhancement in collagenase production and metastasis formation. Zh Mikrobiol Epidemiol Immunobiol, 1980 Dec, (12), 57 - 60 {Modification of the agarose drop micromethod for studying mouse macrophage migration inhibition factor}; Lebedev VS; A modification of the drop agarose micromethod for the study of the factor inhibiting the migration of mouse macrophages is proposed . The syngeneic system was used instead of type RPMI-1640 complex culture media and calf embryonal serum, commonly used in the work with mouse macrophages . The production of the factor inhibiting the migration of macrophages by sensitized lymphocytes from mice of different strains has been shown to occur in vitro under the influence of pertussis corpuscular antigen . The method of obtaining supernatant containing macrophage migration inhibiting factor, from immune mouse lymphocyte culture is proposed . This supernatant may be used for, the analysis of the above-mentioned factor and other lymphokins. Acta Obstet Gynaecol Jpn, 1980 Dec, 32(12), 1937 - 44 {Establishment of persistent cytomegalovirus infection in primary cultured trophoblastic cells (author's transl)}; Yabuki Y; The primary cultured cells (Ch) derived from chorionic villi were infected with human cytomegalovirus (CMV) . The main results obtained are as follows . 1) The primary cultured trophoblastic cell (Ch) infected with CMV have been maintained in the state of CMV persistent infection for over 6 months . These cells (Ch/CMV) were the CMV-carrier cells in a balance between the cytolysis by CMV-specific cytopathic effect (CPE) and the growth of uninfected cells, showing the CMV persistent infection at the cell population levels . 2) The Ch/CMV cells have persistently released infectious CMV with titers ranging from 1.6 X 10(2) to 1.2 X 10(4) PFU/ml into the culture fluid . CMV-specific antigens were detectable by immunofluorescent antibody staining of the virus releasing cells . 3) These Ch/CMV cells still maintained an ability to secret estrone and estradiol . When these abilities lowered in the successive cultures, the cells encountered the cytolysis by CMV release . 4) Infectious titer of CMV released from Ch/CMV cells reduced by the addition of estrogen in the culture media . In this case, however CMV-specific CPE occurred with a similar pattern as the control one lacked an addition of estrogen . 5) It was suggested that estrogen synthesis in the cells may play an important role in an establishment or a maintenance of CMV persistent infection. Circ Res, 1980 Nov, 47(5), 770 - 5 Trophic effect of norepinephrine on the rat portal vein in organ culture; Abel PW et al.; Rat portal veins were maintained in organ culture to study the development of characteristic denervation changes and a possible trophic effect of the neurotransmitter norepinephrine (NE) . Vessels maintained in organ culture for 2 days showed supersensitivity to NE and Ba2+, a more rapid rate of relaxation from a Ba2+ contracture, and partial depolarization of the myovascular cells . All of these changes except the quicker relaxation from Ba2+ contracture could be prevented by incubating the preparations in a NE-containing medium . This evidence suggests that functional changes in vascular muscle cells are caused by the removal of a tropic influence of NE, but can be prevented by NE replacement . However, the failure of NE in the culture media to prevent the increased rate of relaxation from Ba2+ contracture found after 2 days in organ culture suggests that NE is not the only trophic influence acting on the portal vein . In addition, incubation of veins in a NE-containing medium produced a marked subsensitivity to the contractile effects of NE, but not BA2+, and thus possible desensitization of noradrenergic receptors . The data thus support a trophic role for NE in the rat portal vein. Br J Nutr, 1980 Nov, 44(3), 371 - 80 Preparation of an experimental low-fluoride diet from single-cell organisms for rats and mice; Khalawan SA et al.; 1 . A method for producing a standard low-fluoride diet from a green alga and yeast is described . Chlorella pyrenoidosa was grown in a culture medium prepared with distilled water and analytical grade chemical salts . The spent culture medium from the alga culture was reclaimed and replenished with salts and sucrose for the production of yeast, Saccharomyces cerevisiae . 2 . The single-cell organisms were separated by centrifugation from their culture media and the dried cells were blended with sucrose, maize oil, cellulose and a salt mix to produce diet pellets for rats and mice . 3 . The diet was readily accepted as food by rats and mice and it was found to contain 100-300 micrograms fluoride/kg dry weight . Two generations of rats and four generations of mice were bred on this diet . 4 . The use of hydroxyapatite to reduce the fluoride content of the chemical used in the production of the alga and yeast biomass was investigated . Diet pellets prepared with this biomass contained 45-60 micrograms fluoride/kg dry weight. J Clin Endocrinol Metab, 1980 Nov, 51(5), 978 - 87 Ectopic production of somatostatin-like immuno- and bioactivity by cultured human pulmonary small cell carcinoma; Szabo M et al.; Eleven continuous cultures of human pulmonary small cell carcinoma cells were examined, and eight were shown to secrete quantities of somatostatin-like immunoreactivity (SRIF-LI) ranging from 0.07-27 ng/ml culture medium/4 days, SRIF-LI was also found in a 2-N acetic acid extract of one of three human pulmonary small cell carcinomas obtained at autopsy as well as in the extract of a solid tumor resulting from inoculation of nude, athymic mice with SRIF-LI-producing, cultured small cell carcinoma cells . The SRIF-LI produced by one continuous cell line, DMS 53, was characterized in terms of its immunological, chromatographic, and biological properties . SRIF-LI from DMS 53 culture media and lysed cells was heat stable and exhibited parallel displacement to synthetic SRIF standard in a double antibody RIA . DMS 53 SRIF-LI was quantitatively retained on an immunoaffinity column of sheep anti-SRIF-Sepharose 4B under neutral conditions and could be eluted with 2 N acetic acid . Gel filtration chromatography of immunoaffinity-purified SRIF-LI revealed multiple molecular weight forms, the largest of which had an apparent molecular weight of 10,000-12,000 daltons and may represent a precursor form . This high molecular weight SRIF-LI form was resistant to exposure to denaturing conditions (8 M urea or 4 M urea plus 0.5% mercaptoethanol), suggesting the absence of noncovalent and/or disulfide linkages . A low molecular weight form coeluted with synthetic SRIF . Additional evidence for the identity of this form with the tetradecapeptide was provided by highly specific reverse phase high performance liquid chromatography . The rate of degradation of high molecular weight SRIF-LI by the cultures was markedly reduced in comparison to that of the SRIF monomer, resulting in a preferential accumulation of high molecular weight SRIF-LI in 4-day culture medium . Bioactivity of DMS 53 SRIF-LI was assessed in 4-day primary monolayer cultures of rat adenohypophyseal cells where 10(-10)-10(-9) M synthetic SRIF elicited a linear log-dose suppression of 5 X 10(-4) M synthetic SRIF elicited a linear log-dose suppression of 5 X 10(-4) M dibutyryl cAMP-stimulated rat GH release . Immunoaffinity-purified SRIF-LI from DMS 53 lysed cells and 1-h serum-free incubation medium, which consisted predominantly of monomeric SRIF, was equipotent to synthetic SRIF, SRIF-LI from 4-day culture medium consisted mostly of the high molecular weight form and exhibited a reduced bioassay potency ratio relative to synthetic SRIF of 0.73 (95% confidence limits, 0.99-0.53) . Chromatographically purified high molecular weight SRIF-LI had significant bioactivity with a bioassay to immunoassay ratio of 0.19 (95% confidence limits, 0.33-0.09) . The demonstration of ectopic SRIF, production by human pulmonary small cell carcinoma is consistent with the proposed derivation of this tumor from a cell type in the amine precursor uptake and decarboxylation cell series. Lipids, 1980 Oct, 15(10), 838 - 48 Selectivity in incorporation, utilization and retention of oleic and linoleic acids by human skin fibroblasts; Rosenthal MD; Fetal human fibroblasts were grown in culture medium containing 10% fetal bovine serum supplemented with {1(-14)C}linoleate or {1(-14)C}oleate . At all concentrations of exogenous fatty acids, the incorporation of oleate was greater than that of linoleate . With increased medium fatty acid concentrations, linoleate in triacyglycerol (TAG) could be increased from 13 to 75% of the total incorporated; at each concentration, relatively more linoleate than oleate was in TAG . When the cells were exposed to exogenous oleate/linoleate mixtures, the composition of the mixture determined the extent of incorporation of both fatty acids . When the mixture was primarily linoleate, scarce oleate was used preferentially for phospholipids (PL); no such specificity for scarce linoleate was observed . Addition of exogenous fatty acids resulted in a shift of previously incorporated 14C fatty acids from phospholipid into TAG; retention of oleate in PL was greater than that of linoleate . Incorporation of oleate into phospholipids was also higher than that of linoleate from exogenous fatty acid mixtures which were 80% saturated . It is suggested that normal human fibroblasts have adapted to the low levels of exogenous polyunsaturated fatty acids in culture media by increased use of oleate in phospholipid . Even when the cells are supplemented with linoleate, the preferential use of oleate in phospholipid groups is retained. J Exp Med, 1980 Oct 1, 152(4), 1070 - 84 Dendritic cells are accessory cells for the development of anti-trinitrophenyl cytotoxic T lymphocytes; Nussenzweig MC et al.; This study establishes that dendritic cells (DC) are the critical accessory cells for the development of anti-trinitrophenol (TNP) cytotoxic T lymphocytes (CTL) in vitro . We developed a model in which nylon wool-nonadherent spleen cells were used both as the responding and stimulating cells, the latter having been TNP-modified and x-irradiated . Thy-1-bearing CTL developed in C57BL/6, B6D2F1, and CBA mice only when small numbers of DC were added . Maximal responses in 5-d cultures were achieved with 0.5-1 DC/100 responding T cells . The DC did not have to be TNP modified directly . Anti-Ia and complement inactivated accessory cells, whereas similar treatment of the responders had no effect . DC exposed to ultraviolet radiation were ineffective, but x-irradiated DC were fully active . Culture media from DC, or from DC-nylon wool-passed spleen T cell cocultures that contained abundant CTL, would not substitute for viable DC . Enriched preparations of macrophages (M phi) were obtained from blood, peritoneal cavity, and spleens of BCG-immune and unprimed mice . M phi added at doses of 0.2-4% were weak or inactive as accessory cells . The level of Ia antigens on test M phi populations was quantitated and visualized by binding of a radioiodinated monoclonal anti-I-Ab,d antibody, clone B-21 . M phi that bore substantial amounts of Ia from all organs were weak accessory cells . Addition of M phi to DC-T cell cocultures produced inhibitory effects, usually at a dose of 2% M phi . In contrast, 0.5% Ia-bearing M phi from BCG-immune boosted mice inhibited > 80% of the DC-mediated CTL response . Addition of indomethacin reversed M phi inhibition, and 10(-9) M prostaglandin E2 in turn blocked the indomethacin effect . Indomethacin also restored a low level of accessory cell function in immune-boosted adherent peritoneal cells, but not in preparations of monocytes and spleen M phi . Small numbers of DC were identified in preparations of immune-boosted peritoneal cells and may have accounted for the observed accessory activity . We conclude that the development of anti-TNP CTL is an immune response in which (a) DC are the critical accessory cells; (b) Ia-bearing M phi are weak or inactive; and (c) M phi can inhibit DC-mediated response by an indomethacin-sensitive mechanism. Surg Neurol, 1980 Oct, 14(4), 303 - 9 In vitro secretion of follicle-stimulating hormone by pituitary chromophobe adenomas; Takeuchi J et al.; Of 20 female and 14 male patients with chromophobe adenomas, 4 male patients, aged 38 to 47 years, and one female patient, aged 54, showed above normal basal plasma levels of follicle-stimulating hormone (FSH) . An in vitro tissue culture study that was performed using chromophobe cells obtained from 3 of the 4 male patients and the 1 female patient showed moderate FSH secretion into the culture media . The secretion of FSH by pituitary chromophobe adenomas might be more frequent than hitherto suspected. Invest Ophthalmol Vis Sci, 1980 Oct, 19(10), 1204 - 21 Evidence for a role of the plasminogen activator--plasmin system in corneal ulceration; Berman M et al.; Plasminogen is present in the cornea andcan be activated to plasmin by plasminogen activator . Plasmin is able, in turn, to activate latent collagenase . This system could initiate and perpetuate the collagen degradation of corneal ulceration . This report details evidence for such a system in the cornea . Plasmin has been found to activate latent collagenase from organ cultures of ulcerating rabbit corneas and from fibroblast cultures derived from such corneas . As in the case of activation by trypsin, activation by plasmin results in the conversion of the 40,000 MW latent form to an active species of 23,000 MW . Explants of normal or alkali-burned, ulcerating corneas demonstrated plasminogen-dependent lysis of fibrin clots; frozen sections of such corneas demonstrated that lysis begins in the superficial stroma near the periphery of the cornea . Multiply freeze-thawed ulcerating corneas, but not normal corneas, showed initial lysis, not peripherally but at the ulcer region containing polymorphonuclear leukocytes . The fact that the peripheral lytic pattern existed in corneas that were obtained from eyes prefrozen in liquid nitrogen before excision of the corneas would suggest that plasminogen activator is normally contained in cells in vivo and is not made only in response to tissue injury . There was no correlation between the location of blood vessels or the presence of the corneal endothelium and the plasminogen-dependent lysis . Plasminogen activator from the ulcerating cornea and from fibroblasts was characterized by sodium dodecyl sulfate--gel electrophoresis of its cleavage products of plasminogen . The activator cleaves plasminogen into heavy- and light-chain fragments similar to those produced from plasminogen by urokinase . Plasminogen activator activity was quantitated by a new assay that restricts diffusion of the enzyme to one dimension into a narrow bore tube . The addition of plasminogen daily to cultures of ulcerating corneas resulted in earlier rises of plasminogen activator, collagenase, and collagen degradation fragments in the culture media . Although total plasminogen activator levels were not increased by the addition of plasminogen to culture, levels of both collagenase and solubilized collagen were approximately doubled . It is concluded that the plasminogen activator--plasmin system might play an important role in the destruction of stromal matrix in corneal ulceration. J Anim Sci, 1980 Sep, 51(3), 660 - 7 Influence of culture media on in vitro fertilization of ovine tubal oocytes; Bondioli KR et al.; The ability of Synthetic Oviductal Fluid (SOF) and Minimum Essential Medium (MEM) to support sperm binding, sperm penetration and subsequent cell division of ovine tubal oocytes in vitro was evaluated . These media contained no protein, 3% (w/w) bovine serum albumin (BSA) or 20% (v/v) lamb serum (LS) . Midventral laparotomies were performed on 49 ewes synchronized with progesterone and then superovulated, and tubal oocytes were collected by flushing the oviducts . Oocytes were pooled and randomly added to 2 X 10(7) ejaculated spermatozoa in 1 ml of treatment media that had been preincubated in tubes for 3 hr at 37 C under 5% CO2 in air . After an additional 3 hr of culture in a rotating tissue culture drum, oocytes were further cultured in microdrops of the same medium for 48 hr and observed for sperm penetration, cell division and fragmentation . Oocytes were then stained with aceto-orcein and observed for penetration, multiple nuclei and bound sperm . Percentage of oocytes showing penetration in SOF, SOF+ BSA, SOF+LS, MEM, MEM+BSA and MEM+LS was 14, 3.6, 7.8, 11.1, 8.7 and 12.8, respectively; percentage of oocytes cleaving was 8, 3.6, 5.9, 4.4, 4.3 and 4.3; percentage of oocytes fragmenting was 40, 60, 60.8, 20, 60.9 and 29.8, and the average number of sperm cells bound per oocyte was 82, 177, 117, 41, 56 and 47 . No cell division was observed for control oocytes (no sperm added), but 72.7, 80 and 76.9% fragmented in SOF, SOF+BSA and SOF+LS, respectively . No difference was found among the media in their ability to support sperm penetration and subsequent cell division in vitro . All media tested supported a high incidence of sperm binding and oocyte fragmentation. Atherosclerosis, 1980 Sep, 37(1), 11 - 9 Enhanced synthesis of collagen and total protein by smooth muscle cells from atherosclerotic rabbit aortas in culture; Pietila K et al.; Protein synthesis in smooth muscle cells from normal and atherosclerotic rabbit aortas was studied . Cultures of cells from the third passage were incubated with {3H}proline and the synthesis rates of collagen and total protein measured from the radioactivities incorporated into protein-bound hydroxyproline and proline, respectively . The synthesis ratio of collagen types I and III was studied by separating the radioactive procollagens of the culture media by DEAE column chromatography . The synthesis rates of both collagen and total protein were higher in cells cultured from atherosclerotic rabbit aortas . There was no difference in the synthesis ratios of types I and III collagen between cells from the two origins . In addition to procollagens, a third protein eluted before procollagen I in the DEAE chromatography of medium proteins . Compared with control cells, cells from atherosclerotic aortas incorporated relatively less radioactivity into this protein . No differences were detected in the amino acid compositions of cellular proteins between cells from both origins . It was concluded that cells grown from atherosclerotic rabbit aortas synthesize more collagen and total protein than those from normal aortas, the synthesis of collagens I and III being equally enhanced . The results also suggest that there are proteins which are not involved in the general enhancement of protein synthesis. Endocrinology, 1980 Sep, 107(3), 839 - 44 Glucocorticoid dependence of fetal lung maturation in vitro; Torday JS; Earlier studies of the fetal lung in tissue culture have led to the conclusion that its maturation is autonomous . We have studied the role of the small amount of cortisol supplied by the serum routinely added to culture media . It was observed that 28-day-old fetal rabbit lung cells continued to grow and differentiate in culture medium supplemented with 10% calf serum (2.2 X 10(-8) M cortisol) . However, when these cells were propagated in medium supplemented with calf serum which was charcoal stripped or from adrenalectomized animals, growth and differentiation were prevented (<1 X 10(-12) M cortisol) . The addition of cortixol (1 X 10(-10) to 1 X 10(-5) M) to such media restored the ability of these cells to grow and differentiate in culture . Other classes of steroid hormones failed to restore such activity even with a 10-fold excess of cortisol . We conclude that maturation of the 28-day-old fetal rabbit lung in vitro is dependent upon the presence of cortisol. J Cell Physiol, 1980 Sep, 104(3), 349 - 57 Regulation of prostaglandin synthesis during differentiation of cultured mouse myeloid leukemia cells; Honma Y et al.; Mouse myeloid leukemia cells (Ml) were induced to differentiate into mature macrophages and granulocytes by various inducers . The differentiated Ml cells synthesized and released prosetaglandins, whereas untreated Ml cells did not . When the cells wee prelabelled with {14C}arachidonate, the major prostaglandins released into the culture media were found to be prostaglandin E2, D2, and F2 alpha in an early stage of differentiation, but the mature cells produced predominantly prostaglandin E2 . The synthesis and release of prostaglandins were completely inhibited by indomethacin . Dexamethasone, a potent inducer of differentiation of Ml cells, did not induce production of prostaglandins in resistant Ml cells that could not differentiate even with a high concentration of dexamethasone . These results suggest that production of prostaglandins in Ml cells is closely associated with differentiation of the cells . Homogenates of dexamethasone-treated Ml cells converted arachidonate to prostaglandins, but this conversion was scarcely observed with homogenates of untreated Ml cells . Dexamethasone and the other inducers stimulated the release of arachidonate from phospholipids . Therefore, induction of prostaglandin synthesis during differentiation of Ml cells may result from induction of prostaglandin synthetase activity and stimulation of the release of arachidonate from cellular lipids . Lysozyme activity, which is a typical biochemical marker of macrophages, was induced in Ml cells by prostaglandin E2 or D2 alone, as well as by inducers of differentiation of the cells, but it was not induced by arachidonate or prostaglandin F2 alpha . These results suggest that prostaglandin synthesis is important in differentiation of myeloid leukemia cells. J Biol Chem, 1980 Aug 10, 255(15), 7071 - 4 NH2-terminal sequence analysis of pro-C4, the precursor of the fourth component of guinea pig complement; Goldberger G et al.; The amino acid sequence of the NH2-terminal regions of the intracellular precursor of the fourth component of guinea pig complement (pro-C4) and isolated alpha, beta, and gamma chains of native C4, synthesized by peritoneal macrophages in culture, were determined with a microradiosequencing technique . Radiolabeled pro-C4 was immunoprecipitated from cell lysates and native C4 from culture media which contained one of six 3H-amino-acids or {35S}methionine . The purity of these proteins was established and molecular weights were estimated by electrophoresis in sodium dodecyl sulfate . Seventeen of the first 22 residues of pro-C4 were identified . Fifteen of these 17 were nonpolar . The eight amino acids ascertained within the first 9 NH2-terminal residues of guinea pig pro-C4 were identical with those reported for human C4 beta chain, and identity with residues determined for guinea pig C4 beta chain was established . These data indicate that beta chain is the NH2-terminal segment of pro-C4, that no residues are cleaved from the NH2 terminus during the conversion to native C4, and that this segment of the pro-C4 molecule is conserved phylogenetically. Science, 1980 Aug 8, 209(4457), 701 - 2 Estrogen and the growth of breast cancer: new evidence suggests indirect action; Shafie SM; The growth of the MCF-7 human breast cancer cell line is unresponsive to the presence of estrogen in culture media . Paradoxically, in nude mice, growth of these cells and formation of solid tumors are dependent on estrogen . Tumors fail to develop in ovariectomized mice, but do develop in intact mice and in ovariectomized mice given estrogen . Primary cultures derived from MCF-7 tumors revert to unresponsiveness to estrogen . However, when these cultures are again transplanted into nude mice, estrogen is required for tumor formation . The continuous culture, the solid tumor, and the primary cultures therefrom have similar estrogen-binding capacities and affinities . These results indicate that mammary carcinoma cell growth in vivo is subject to inhibition that can be overcome by estrogen. In Vitro, 1980 Aug, 16(8), 669 - 74 An improved culture medium for mouse blastocysts; Spindle A; Eagle's basal medium, modified to contain essential amino acids at the concentrations optimal for mouse blastocyst hatching, attachment, and outgrowth, supported in vitro development of the mouse blastocyst better than other tissue culture media tested . This medium was improved for growth and differentiation of the inner cell mass by doubling the concentration of amino acids and glucose and by adding uridine (10(-5) M) and beta-mercaptoethanol (10(-5) M) . In this improved medium nearly all blastocysts grown from the two-cell stage hatched and formed trophoblast outgrowths, and 62% developed into two-layer egg cylinders. Appl Environ Microbiol, 1980 Aug, 40(2), 391 - 9 Cholesterol as a limiting factor in the growth of Mycoplasma pneumoniae; Johnson JK et al.; Ultracentrifugation was used to separate three commercial lots of bovine serum fraction (BSF) into components designed to contain lipoproteins . Each BSF lot and component was tested for ability to support the growth of tree strains of Mycoplasma pneumoniae . In general, the level of growth-promoting activity corresponded to the amount of cholesterol present in the BSF or BSF components rather than to the amount or type of lipoprotein Cholesterol was the limiting nutritional factor of BSF with low growth-promoting activity . The addition of cholesterol and bovine serum albumin to BSF with low activity resulted in growth equal to or greater than that observed for BSF with high growth-promoting activity . When cholesterol was added to agar medium containing BSF of low activity, mycoplasma colonies were greater in number, possessed larger mean diameters, and had centers that were more distinct than those observed when this BSF was used alone . Variability in growth-promoting actions of commercial lots of BSF was eliminated by increasing their cholesterol content to an optimum level . An adjustment of the cholesterol and albumin levels of any serum product used in culture media may provide a simple convenient method to improve growth and isolation of mycoplasmas. Am J Anat, 1980 Aug, 158(4), 433 - 444 Maintenance of gonadotrophs in pituitary autografts under the kidney capsules of female rats given sex hormones or LRH; Shino M et al.; Female Sprague-Dawley rats were hypophysectomized and the anterior pituitary gland was immediately placed under the kidney capsule . For 1 week after surgery, groups of pituitary autograft-bearing animals were treated with twice-daily injections of estradiol 17 beta (E), progesterone (P), estradiol 17 beta and progesterone (EP), or luteinizing hormone-releasing hormone (LRH) . Within 2--4 hours following the last injection, the pituitary grafts were removed and placed into organ culture . They were maintained in culture with or without added LRH (10(-7) M) for 1 hour at 37 degrees C . The culture media were then frozen for later radioimmunoassay of FSH and LH . The tissues were kept in culture for an additional 24 hours, at which time they were fixed and prepared for immunocytochemistry or electron microscopy . Results showed that treatment of the animals with E, EP, or LRH enhanced the release of FSH and LH into the culture media, and that the release of these hormones was increased further by acute incubation with LRH . The ultrastructure of the gonadotrophs was well maintained by treating the animals with E or the combination of E and P or with LRH . Graft tissue from animals treated with LRH, which was incubated subsequently for 24 hours with LRH, showed the best maintenance of gonadotroph morphology . This experimental procedure should be useful for obtaining gonadotrophs for use in establishing gonadotroph cell lines. J Biol Chem, 1980 Jul 25, 255(14), 6579 - 83 Secretion of the vitamin K-dependent protein of bone by rat osteosarcoma cells . Evidence for an intracellular precursor; Nishimoto SK et al.; Four clonal cell lines derived from a rat osteosarcoma were tested for the ability to secrete the gamma-carboxyglutamic acid-containing protein of bone (BGP) using a specific radioimmunoassay for this protein . Two cell lines secreted BGP into culture media while the other two did not . Other investigators have shown that these two cell lines are also the only ones with the high parathyroid hormone responsiveness and alkaline phosphatase activity expected for osteoblast cells in culture . Both cell lines also form a mineralized sarcoma when implanted in rats . The BGP in culture media is identical in molecular weight and in electrophoretic mobility with the 5800-dalton BGP purified from rat bone . Thus, BGP is probably secreted by osteosarcoma cells directly and not derived from an extracellular precursor by proteolytic cleavage . There are two immunoreactive components within osteosarcoma cells which secrete BGP . One component is identical in molecular weight and electrophoretic mobility with BGP from rat bone . The other component has a higher molecular mass (approximately 9000 daltons) and about half the electrophoretic mobility of BGP from bone . The presence of both components within these cells raises the possibility that the larger component may be an intracellular precursor which is processed to BGP prior to secretion. J Biol Chem, 1980 Jul 10, 255(13), 6036 - 9 Undersulfated proteoglycans are secreted by cultured chondrocytes in the presence of the ionophore monensin; Tajiri K et al.; The monovalent ionophore, monensin, inhibits secretion of many different proteins from a wide variety of cells . The site of blockage is at the golgi complex . We have exposed chick embryo chondrocytes in suspension culture to monensin, at concentrations ranging from 10(-8) to 10(-6) M . At the higher concentrations, between 10(-7) and 10(-6) M, monensin inhibited secretion of type II procollagen, which accumulated in the chondrocytes . At these concentrations of the ionophore, proteoglycan synthesis was inhibited, as measured by radioactive serine incorporation into core proteins and by radioactive glucosamine or SO4 incorporation into glycosaminoglycans . However, at a monensin concentration of 3 x 10(-8) M, the incorporations of serine and glucosamine were close to normal while SO4 incorporation was at 30% of control values . The ratio of glucosamine to serine in pronase-released glycosaminoglycans from culture media was unaffected by 3 x 10(-8) M monensin but the sulfate to serine ratio decreased to 29% of control values . Examination of the glycosaminoglycans by gel filtration showed a progressive increase in Kav values as sulfation decreased . Undersulfation was demonstrated by radiochromatographic analysis of the digestion products following incubation with chondroitinase ABC . The composite results show that monensin interferes with sulfation of newly synthesized proteoglycans. J Cell Physiol, 1980 Jul, 104(1), 41 - 6 Murine sarcoma growth factors affect the growth and morphology of cultured mouse epithelial cells; Keski-Oja J et al.; A mouse embryo epithelial cell line, MMC-E, was used to study the effects of purified murine sarcoma growth factors (SGFs) on the growth of epithelial cells . Murine SGF was a partially purified preparation from the serum-free culture media of mouse fibroblasts transformed by Moloney murine sarcoma virus . SGFs stimulated DNA-synthesis in resting cells and induced them to grow to higher densities than control cells . With continued exposure to SGFs, MMC-E cells lost their postconfluency inhibition of division and formed small expanding foci . When the SGFs were removed and the cells were subcultured, they regained their normal phenotype, showing that the effects of SGFs are reversible on these epithelial cells . SGFs could also stimulate the MMC-E epithelial cells to grow in soft agar, like the syngeneic fibroblasts (MMC-F) from the same mouse embryo, but slower than the control fibroblastic clone . Microgram quantities of SGFs were needed to stimulate soft agar growth of MMC-E cells . The results indicate that SGF can bring about a phenotypic change in the growth pattern of epithelial cells as well as fibroblastic cells. Biomedicine, 1980 Jul-Aug, 33(4), 103 - 5 Effect of levamisole on granulopoiesis in agar culture; Hellmann A et al.; We studied the effect of levamisole on the proliferation of granulocyte-committed progenitor cells (CFU-c) in the agar system . Media conditioned by pooled human peripheral blood mononuclear cells served as source of colony-stimulating activity (CSA) . Levamisole at concentrations ranging from 0.1 to 10.0 micrograms/ml had no detectable stimulatory or inhibitory effect on granulopoiesis in vitro . Levamisole added to culture media neither enhanced nor reduced the release of mononuclear cells of CSA . We conclude that the cases of granulocytopenia, occasionally severe, associated with levamisole therapy are more likely to be due to individual idiosyncrasy or hypersensitivity then to dose-related myelotoxicity. J Clin Microbiol, 1980 Jul, 12(1), 74 - 8 Monoclonal antibodies specific for dengue virus type 3; Dittmar D et al.; Mouse lymphocyte hybridomas were prepared by polyethylene glycol-mediated fusion of cells from a mouse plasmacytoma line with lymphocytes from a mouse hyperimmunized with dengue virus type 3 (dengue-3) . Media from 50 hybrid colonies were screened; 46 of them showed antibody activity against dengue-3-infected cells as determined by an indirect immunofluorescent antibody technique . Dengue monoclonal antibody obtained after cloning one of these colonies demonstrated activity in hemagglutination inhibition and indirect immunofluorescent antibody assays with dengue-3 antigen, but not type 1, 2, and 4 antigens . In addition, this antibody activity could be removed from culture media only by absorption with dengue-3 antigen. Acta Physiol Scand, 1980 Jul, 109(3), 275 - 81 Effects of purified macrophage RNase on granuloma fibroblasts with reference to silicosis; Aho S et al.; Two alkaline RNases, designated RNase 1 and RNase 2, were isolated from the culture media of silica-treated and non-treated macrophages . The yield of RNase from the medium of silica-treated macrophages was 30% of that from the non-treated control . The effects of these RNases on cultured granuloma fibroblasts and on granulation-tissue nuclei were studied . RNase 1 inhibited thymidine incorporation into fibroblasts except at low concentrations, where it was observed to be stimulatory . RNase 1 also inhibited the protein synthesis of fibroblasts . The incorporation of cytidine into RNA in cultured fibroblasts was not affected by RNase 1, but the incorporation into isolated nuclei was decreased . In pulse chase experiments RNase 1 increased the release of cytidine, but not that of thymidine, from the cells . RNase 2 had no effect on the protein or nucleic acid metabolism of the fibroblasts or on the RNA metabolism of isolated nuclei, perhaps because of impermeability . These experiments confirm that macrophage RNase activity is able to regulate the metabolism of granulation-tissue fibroblasts by increasing RNA degradation . Through this action it also regulates DNA and protein synthesis and other metabolic functions of those cells. J Gen Virol, 1980 Jul, 49(1), 83 - 9 Enzyme-linked immunosorbent assay for coronaviruses HCV 229E and MHV 3; Kraaijeveld CA et al.; The antigenic relationship between human cornonavirus strain 229E (HCV 229E) and mouse hepatitis virus strain 3 (MHV 3) was studied by means of the indirect form of enzyme-linked immunosorbent assay (ELISA) . A cross-reaction was found with hyprimmune rabbit sera between HCV 229E and MHV 3 which may be due to the adherence of bovine serum componeants from tissue culture media, which were present on virus particles even after extensive purification . No cross-reaction was observed with immune sera absorbed with bovine serum, or with HCV 229E grown in tissue culture without serum . This indirect ELISA with HCV 229E may prove to be useful for studies with human sera. J Clin Endocrinol Metab, 1980 Jul, 51(1), 182 - 4 Evidence for a peptide similar to 16K fragment in man . Its relationship to ACTH; Bertagna X et al.; A radioimmunoassay directed toward the NH2-terminal region of mouse pro-ACTH/endorphin (called 16K fragment) was used to examine human samples . Culture media from two corticotropic adenomas and plasmas from 11 patients with various ACTH hypersecretory syndromes gave parallel displacement curves; displacement curves for human samples were not parallel to purified mouse 16K fragment . Following sodium dodecyl sulfate polyacrylamide gel electrophoresis of culture medium from one adenoma, a major peak of 16K fragment immunoreactivity with an apparent molecular weight of ca . 16,000 was detected . A significant correlation (r = 0.963 ; p less than 0.001) was found between immunoreactive 16K fragment and ACTH in the patients' plasmas . These data indicate that a peptide similar to 16K fragment exists in man ; that human and mouse 16K fragment are immunologically distinguishable and that human 16K fragment appears to be secreted concomitantly with ACTH. J Biol Chem, 1980 Jun 25, 255(12), 5681 - 7 Human adenosine deaminase and its binding protein in normal and adenosine deaminase-deficient fibroblast cell strains; Daddona PE et al.; Reduced or absent adenosine deaminase activity occurs in all tissues of some patients with severe combined immunodeficiency disease . In this report we describe (a) a sensitive and specific radioimmunoassay for fibroblast adenosine deaminase and its binding protein, (b) the importance of various tissue culture media on the expression of fibroblast adenosine deaminase, and (c) the levels of immunoreactive adenosine deaminase compared to its binding protein in normal and homozygous adenosine deaminase-deficient fibroblast cell strains. Diabetes, 1980 Jun, 29(6), 497 - 500 Monolayer cultures of adult pancreatic islet cells on osmotically disrupted fibroblasts; Meda P et al.; A simple method that permits the attachment of fully dissociated adult pancreatic islet cells to culture dishes, using osmotically disrupted fibroblasts, is described . Using this system, monolayer cultures consisting of either isolated or clustered adult endocrine islet cells can be readily established in standard culture media. Scand J Clin Lab Invest, 1980 Jun, 40(4), 311 - 8 Liberation of a fibrogenic factor from human blood monocytes, ascites cells, cultured histiocytes and transformed mouse macrophages by treatment with SiO2; Aalto M et al.; Human monocytes and ascites macrophages from cirrhotic patients were isolated in Percoll-gradient and cultured with and without silica . Similar experiments were carried out also with cultured malignant human histiocytes and transformed mouse macrophages . The fibrogenic activity of the culture media was tested by measuring the incorporation of {3H}proline and {3H}thymidine into cultured rat granuloma and human synovial cells . Media from silica-treated monocytes, ascites macrophages and certain histiocyte and mouse macrophage lines caused an increase in the incorporation of both {3H}proline and {3H}thymidine into collagen and DNA, respectively, in both cell systems . Alkaline RNase activities were decreased markedly in the media from silica-treated ascites macrophages but not in the media of the monocytes or histiocytes. Biochim Biophys Acta, 1980 May 28, 618(2), 347 - 58 Production of apolipoproteins E and A-I by rat hepatocytes in primary culture; Dashti N et al.; Hepatocytes were isolated from normal adult rat livers and cultured in a modified HI-WO/BA medium . A nearly confluent monolayer was established at the plating concentration employed . The hepatocytes synthesized ansd secreted albumin at rates similar to those observed in vivo . The cells secreted triacylglycerol in the absence of fatty acid substrate . Under these conditions the most abundant triacylglycerol molecular species contained 53 carbons . Incubation with oleic acid markedly increased triacylglycerol secretion predominantly in the form containing a total carbon number of 57 . Approx . 80% of the secreted cholesterol was in the free form and this was unaffected by oleic acid . Employing monospecific antibodies constant rates of synthesis and secretion of apolipoproteins E and A-I were demonstrated by quantitative electroimmunoassay of the cell culture media . The rates of albumin, apolipoprotein E, and apolipoprotein A-I production were 1480, 170 and 60 microgram/h per g cell protein, respectively. Gastroenterology, 1980 May, 78(5 Pt 1), 925 - 30 Effects of carbachol and atropine on gastrin secretion and synthesis in rat antral organ culture; Harty RF et al.; The purpose of the present study was to examine the effects of the cholinergic agent carbachol on gastrin synthesis and secretion by rat antral mucosa in organ culture . In addition, the effect of atropine on basal and carbachol-stimulated gastrin release was investigated . These in vitro studies demonstrate that carbachol stimulated both gastrin secretion and synthesis in a dose-dependent manner . Maximal stimulation of gastrin synthesis and release occurred at a concentration of 1 X 10(-5) M carbachol . The rate of gastrin release stimulated by carbachol (1 X10(-5) M) was 0.79 ng-hr-1-mg-1 which was significantly greater than the control value of 0.37 ng-hr-1 (P less than 0.001) . Atropine, over the dose range from 10(-7) to 10(-3) M, had no effect on basal (unstimulated) gastrin release . However, carbachol-stimulated gastrin release was inhibited progressively by inclusion of increasing concentrations of atropine in the culture media . The results of these studies indicated that the acetylcholine analogue, carbachol, is capable of directly stimulating the antral gastrin cell to significantly increase the rate of synthesis and release of gastrin . Whereas atropine, under these in vitro conditions, does not alter basal gastrin release, the muscarinic antagonist does competively inhibit carbachol-stimulated gastrin secretion. J Pediatr, 1980 May, 96(5), 837 - 41 X-linked mental retardation, macro-orchidism, and the Xq27 fragile site; Turner G et al.; Twenty-three families with X-linked mental retardation were examined for the presence of a fragil site on the long arm of the X chromosome (Xq27 fra) . Specific culture media were necessary to demonstrate this site . In only seven of the families was the Xq fragile site observed; in these, all of the affected males had both the fragile X and macro-orchidism . Macro-orchidism was not observed in the remaining 16 families . In the families with Xq27 fra segregating the fraes . This correlated with the age of the carrier . The 25 affected males with macro-orchidism and Xq27 fra had some minor clinical features in common: there was an increase in birth weight, high forehead, prognathism, pale irides, big ears, and an increased head circumference in infancy and childhood which did not persist into adult life . The majority of the affected individuals were moderately retarded. Biomedicine, 1980 May, 33(3), 61 - 2 Effect of antiobiotics on elimination of mycoplasma from experimentally infected cells; Negre G et al.; The primary and continuous cell cultures and mouse peritoneal macrophages were experimentally infected by Mycoplasma pneumoniae and M . orale I strains and maintained in culture media containing high concentrations of tetracycline or kanamycin . The viable mycoplasmas were eliminated from cell supernatants but not from cells. Ann Endocrinol (Paris), 1980 May-Jun, 41(3), 157 - 92 {Recent data on somatomedins (insulin-like growth factors) (author's transl)}; Binoux M; Data on somatomedins have resulted from the convergence of two initially distinct lines of research relating to the two major aspects of their biological activities, their effects on cartilage growth and their insulin-like action . Human serum has yielded three factors, SM-A, SM-C and NSILA-S . The latter comprises two very similar polypeptides, IGF I and IGF II which structurally closely resemble proinsulin . A fourth factor, MSA, has been isolated from calf serum and from conditioned medium of a rat liver cell strain . All these chemically and biologically closely related factors have a molecular weight of approximately 7 500 but circulate in a form with much higher molecular weight owing to their binding to specific carrier proteins . Over the past five years, the use of purified somatomedins has led to rapid progress in the elucidation of the physiology of these hormones . In this review, present knowledge of somatomedins is analysed in the following terms: physiochemical properties, circulating forms, action on growth, insulin-like activity, interaction with receptors, immunological characteristics, biosynthesis and hormonal regulation of their blood level . Results obtained in our laboratory from rat liver organ culture are discussed: the action of culture media on cartilage sulphation, the detection of a protein which specifically binds NSILA-S and SM-A and its use in a competitive protein binding assay for somatomedins in biological fluids. Arch Ophthalmol, 1980 May, 98(5), 919 - 23 Collagenase levels in a new model of experimental herpetic interstitial keratitis; Campbell R et al.; A reproducible model of severe herpetic interstitial keratitis was developed by injecting live herpes simplex virus type 2 intrastromally into the corneas of presensitized rabbits . Herpes-infected corneas showed significantly more stromal infiltration and vascularization, iritis, conjunctivitis, and epithelial disease than the control corneas injected with cell supernatant without virus . Levels of total collagenase detected in the culture media of the herpes-infected corneas were high and were similar to those observed previously in alkali-burned rabbit corneas . Unlike alkali-burned corneas, the herpes-infected corneas showed much more of the enzyme in the latent form during the first two days of culture. J Cell Physiol, 1980 Apr, 103(1), 47 - 54 Fibronectin from aged fibroblasts is defective in promoting cellular adhesion; Chandrasekhar S et al.; Late passage fibroblasts show decreased cell-substrate adhesion . We provide evidence that the reduced adhesion is due to a defect in the adhesive glycoprotein fibronectin . Late passage cells become more adhesive in culture media that has been conditioned by the growth of early passage cells . Analysis of fibronectins purifed from early and late passage cell conditioned media indicates that there are striking differences in their abilities to promote cell adhesion . Young cell fibronectin supports the maximal adhesion of both young and old cells . However, old cells require quantitatively more fibronectin . In contrast, old cell fibronectin is less effective in supporting the adhesion of either cell type . In addition, neither cell type achieves a normal morphology in the presence of old cell fibronectin . The results support the conclusion that the fibronectin released by late passage cells i defective and does not support normal cell-substrate interactions. In Vitro, 1980 Apr, 16(4), 330 - 6 Evaluation of growth hormone production from GH1 cells in vitro: effect of culture media and time in culture; Bolton WE et al.; Growth hormone production by a rat pituitary tumor cell line (GH1) was measured during lag, exponential, and plateau phases of growth in different culture media . Growth hormone secretion was low during lag and early exponential phase; it increased late in the exponential phase and continued to increase during the plateau phase . This biphasic pattern of growth hormone production was observed in all media and sera utilized . Both the doubling time and growth hormone production were influenced by the choice of media and sera . In addition, the length of time in culture affected the growth fraction with passage level 40 GH1 cells having a 79% growth fraction, whereas the growth fraction of passage level 100 cells was 95% . Using the population doubling time as a criterion for a choice of medium, F-10 medium supplemented with 20% fetal bovine serum consistently yielded the most rapid doubling time (32 hr), whereas Dulbecco's MEM supplemented with 15% horse serum and 2.5% fetal bovine serum yielded the greatest plateau cell density . Growth hormone secretion and the population doubling times were directly related to culture conditions including length of time in culture, choice of tissue culture media, choice of sera, and the phase of cell growth (lag, exponential or plateau). Cancer Res, 1980 Apr, 40(4), 1084 - 90 Effect of fatty acid modifications of cultured hepatoma cells on susceptibility to complement-mediated cytolysis; Yoo TJ et al.; The fatty acid composition of Morris Hepatoma 7777 cells was modified by exposure to culture media that were supplemented with 0.1 to 0.36 mM oleic or linoleic acid for 5 days . Changes occurred in the fatty acid composition of both the cellular phospholipid and the neutral lipid fractions . Exposure to linoleic acid caused a large increase in the polyunsaturated fatty acid content of the cell lipids, whereas enrichment in monoenoic fatty acids occurred when the cells were exposed to high levels of oleic acid . Cellular phospholipid content decreased, cholesterol content did not change, and triglyceride content increased as a result of fatty acid supplementation . The fatty acid-modified cells showed increased susceptibility to complement-mediated cytolysis as compared with control cells grown in unsupplemented culture media . The extent of the increase in susceptibility to cytolysis depended on the degree of lipid modification and also on the cell number and antibody titer used in the assay . Cells enriched with linoleic acid were the most susceptible, but oleic acid enrichment also produced increased susceptibility to immune cytolysis . The kinetic study showed that the initial rate of cytolysis was higher in fatty acid-modified cells than in the control cells . There was no difference in the osmotic fragility of the control and fatty acid-modified cells . These results indicate that changes in the lipid composition of a target cell can influence its susceptibility to complement-dependent cytolysis. Appl Environ Microbiol, 1980 Apr, 39(4), 818 - 22 Inhibitory effects of spices on growth and toxin production of toxigenic fungi; Hitokoto H et al.; The inhibitory effects of 29 commercial powdered spices on the growth and toxin production of three species of toxigenic Aspergillus were observed by introducing these materials into culture media for mycotoxin production . Of the 29 samples tested, cloves, star anise seeds, and allspice completely inhibited the fungal growth, whereas most of the others inhibited only the toxin production . Eugenol extracted from cloves and thymol from thyme caused complete inhibition of the growth of both Aspergillus flavus and Aspergillus versicolor at 0.4 mg/ml or less . At a concentration of 2 mg/ml, anethol extracted from star anise seeds inhibited the growth of all the strains. Proc Natl Acad Sci U S A, 1980 Apr, 77(4), 2084 - 7 Control of growth of estrogen-sensitive cells: role for alpha-fetoprotein; Soto AM et al.; The role played by alpha-fetoprotein (AFP) during the perinatal period in rats has not yet been ascertained despite earlier suggestions that this plasma protein affected the multiplication, in an "in animal-in culture" system of tumor cells that are stimulated by estrogen (E) for growth . To test this inference developed from our previous work, AFP was purified by reverse affinity chromatography to homogeneity by electrophoretic and immunochemical criteria . Purified AFP was added at different concentrations to horse serum-supplemented medium which by itself is able to allow maximal exponential growth of a rat pituitary tumor cloned cell line C29RAP . These cells carry estrophilins and their growth is stimulated by E in animals but not in culture . At 3 mg/ml in culture media, AFP prevented growth of C29RAP cells; the effect was dose dependent . F4C1, a rat pituitary cloned cell line that carries estrophilins but shows autonomous behavior when injected into male and female Fisher rats, was not affected in cultured by comparable concentrations of AFP in the culture media . The effect of AFP on the growth of E-sensitive cells in culture was not reversed by E administration . We conclude from these experiments that (i) AFP is a specific inhibitor of the cell multiplication of cells that are E-sensitive for growth (as defined in this presentation), (ii) estrophilins seem not to play a significant role in this inhibition, and (iii) E appears not to be a growth-promoting hormone per se. Nippon Naibunpi Gakkai Zasshi, 1980 Mar 20, 56(3), 262 - 73 {Osmotic pressure and angiotensin II stimulation of arginine vasopressin release from a guinea pig hypothalamo-neurohypophyseal complex in organ culture (author's transl)}; Ishikawa S et al.; An organ culture system of a male guinea pig hypothalamo-neurohypophyseal complex (HNC) was established . On day 5 in culture, (Na+ -K+) ATPase activity was 0.83 +/- 0.11 mM Pi/mg prot/hr (mean +/- SEM): that is 67% of that on day 1 in culture . 3H-thymidine incorporated into DNA in the explants of HNC was 1,205 +/- 185 cpm/microgram DNA . The explants responded to the elevated KCl medium and the hypertonic solution of sodium chloride with a 470 +/- 38% and 298 +/- 31% increase in arginine vasopression (AVP) release, respectively . This response was inhibited by the addition of tetrodotoxin to the culture medium . AVP release from the explants in res-onse to angiotensin II increased significantly in a dose dependent manner . {Sar1, Ile8} angiotensin II, however, attenuated the response of the explants to angiotensin II when administered simultaneously with angiotensin II . These results suggest that angiotensin II and its analogue cause the AVP release from the explants in a competitive manner . The concentrations of AVP in the culture media made hypertonic with sodium chloride, sucrose and mannitol were 298 +/- 31% (p less than 0.01), 251 +/- 36% (p less than 0.01) and 255 +/- 59% (p less than 0.05) of their control values, respectively . The hypertonic solutions of sodium chloride, sucrose and mannitol caused AVP release from the explants in vitro, while the hypertonic solutions of glucose and urea were revealed to be poor osmotic stimuli on AVP release . These results support the concept of osmoreceptors to release AVP from the hypothalamo-neurohypophyseal axis. Mycopathologia, 1980 Mar 17, 70(2), 125 - 8 Production of aflatoxin by Aspergillus flavus (UBMI) in some Nigerian indigenous beverages and foodstuffs; Alozie TC et al.; Sixteen samples of some Nigerian indigenous beverages and foodstuffs were analyzed for their aflatoxin content . All the eight samples of beverages tested were found to be contaminated with aflatoxin . Of the eight samples of foodstuffs tested, all contained aflatoxin except ewedu, dawadawa and shoko yokoto . All the beverages used as culture media for Aspergillus flavus (UBMI) supported the growth of the fungus and aflatoxin elaboration . A . flavus was found to grow luxuriantly on all samples of foodstuffs, except dawadawa . However, growth of the fungus on foodstuffs was not synonymous with aflatoxin production. Biochem J, 1980 Mar 15, 186(3), 971 - 5 Effect of tunicamycin and cycloheximide on the secretion of acid hydrolases from I-cell cultured fibroblasts; Miller AL et al.; I-cell cultures fibroblasts secrete excessive amounts of N-acetyl-beta-D-hexosaminidase and alpha-L-fucosidase into the culture media as compared with normal fibroblasts . Addition of tunicamycin or cyd {14C}leucine (40--50%) into trichloroacetic acid-precipitable material decreased the secretion of these I-cell hydrolases to normal values within 24 h, but had no effect on the secretion of acid hydrolases from normal fibroblasts . These results indicate that I-cell cultured fibroblasts secrete at least two types of acid hydrolases: one is tunicamycin- and cycloheximide-sensitive and constitutes the greater proportion of the secreted hydrolases, and a smaller proportion is insensitive to tunicamycin and cycloheximide, similar t9 the acid hydrolases secreted by normal cultured fibroblasts. Am J Vet Res, 1980 Mar, 41(3), 372 - 6 Inhibition of lymphocyte blastogenesis by sera from cows with lymphoma; Jacobs RM et al.; The sera from cows with lymphoma inhibit the blastogenesis of normal lymphocytes . The inhibitory sera were partially characterized . Twenty of 34 sera from cows with lymphoma caused inhibition, whereas 3 of 25 sera from healthy animals were inhibitory . Sera from 15 cows with various inflammatory conditions did not cause inhibition . Seventy-four cows were tested for responsiveness to phytohemagglutinin, and of this group, the stimulation indices of 25 animals which had antibody to the bovine leukemia virus did not differ from the remainder of the group . The inhibitory substance: (a) was heat stable at 56 C for 30 minutes, (b) was not overcome by the addition of phytohemagglutinin, (c) was active in a manner that was proportional to the concentration of serum in the culture media, (d) was not lymphocytotoxic at a concentration which was profoundly inhibitory to blastogenesis, (e) was found in the first peak eluted from a Sephadex G-200 column, (f) was nonspecifically active because it inhibited the blastogenesis of a panel of lymphocytes from healthy animals, (g) was present in 59% of cows with lymphoma in the terminal stages of disease, (h) was rarely present in healthy cows or cows with inflammatory conditions, and (i) was not associated with bovine leukemia virus infection in the absence of tumor development. Acta Paediatr Scand, 1980 Mar, 69(2), 263 - 7 An arthropathic form of osteogenesis imperfecta; Penttinen R et al.; A 14-year-old girl gradually developed severe osteoporosis and destructive generalized joint disease, resulting in joint stiffness and anchyloses . She also had moderate hydroxyprolinemia and hydroxyprolinuria . Rheumatoid arthritis was highly unlikely . Anamnestic data revealed two long bone fractures . Collagen biosynthesis was studied in fibroblasts cultured from the patient's skin . Chromatograms of 3H-labelled culture media proteins on ion exchange celluloses revealed an increased ratio of type III collagen to type I collagen when compared with the chromatograms of age-matched control fibroblasts . This finding is typical of certain cell strains in osteogenesis imperfecta . The patient might thus express a new variety of osteogenesis imperfecta with chronic arthropathy. Clin Sci (Lond), 1980 Mar, 58(3), 201 - 10 Prostaglandins as mediators of bone resorption in renal and breast tumours; Greaves M et al.; Amounts of prostaglandin E and prostaglandin F have been measured by radioimmunoassay in extracts of renal cortical carcinoma and benign and malignant breast tumours after solvent extraction and column chromatography . 2 . Substantial amounts of prostaglandin E were found in extracts of benign and malignant breast tumours and in renal tumours . Much lower amounts of prostaglandin F were present in all tumour types . 3 . Co-cultivation of tumour explants with mouse calvaria led to significant bone resorption in 10 of 13 renal carcinomas, three of eight malignant breast tumours, and two of nine benign breast tumours . Tumours associated with bone resorption had higher concentrations of prostaglandin E in culture media at the end of incubation than did non-resorbers . 4 . Indomethacin (14 mumol/1) greatly reduced bone resorption in the presence of the tumour, but this was not always complete, particularly with breast tumours . Indomethacin had no effect on prostaglandin-induced bone resorption . Theophylline (2.2 mmol/1) significantly increased prostaglandin E production and resorption by an effect on the tumour . 5 . It is concluded that prostaglandins may be important in mediating the effects of renal cortical carcinoma and possibly breast cancer on bone destruction . A non-prostaglandin mechanism may also contribute to bone destruction by breast carcinoma. Ann Intern Med, 1980 Mar, 92(3), 396 - 405 Pemphigus: current concepts; {The maintenance of insulin secretory response of long-term cultured pancreatic islets to glucose et al.; Tsutou A, Kawaguchi A, Taniguchi H, Yoshioka M, Tamagawa M, Seki M, Murakami K, Kobayashi T, Baba S. The present study was performed to investigate whether long-term cultured rat pancreatic islets possess a postcultural insulin secretory response to hormones and neurotransmitters in spite of their lack of stimulation during the culture period . We also investigated the method of maintaining the insulin secretory response of islets cultured in a physiological concentration of glucose . The tissue culture media were TCM 199 supplemented with 5.5 mM glucose (A medium), 5.5 mM glucose plus 1 mM adenosine (B medium), 16.7 mM glucose (C medium), and 16.7 mM glucose plus 1 mM adenosine (D medium) . Short-term incubation after the culture period of 14 days showed that the islets cultured in B, C and D media maintained the same insulin secretory responsiveness to 8.3 mM glucose and/or 5 microM acetylcholine and also to 1 microM epinephrine as did non-cultured islets . A similar response was found among the islets maintained in B, C and D media . An insulin secretory response to epinephrine and phentolamine was deficient in islets cultured in A medium, whereas it was maintained in those cultured in C medium . The responsiveness of the islets cultured in C medium to the concomitant stimulation by epinephrine and phentolamine was not different from that of the non-cultured islets . It was thus concluded that the addition of adenosine in the culture medium containing the physiological concentration of glucose was as effective in amintaining the insulin secretory ability of the islets as was the culture medium containing a high concentration of glucose, and it was suggested that even the pancreatic islets cultured in these media, though separated from the innervation might preserve acetylcholine and adrenergic receptors similar to freshly isolated islets . Considering the action of adenosine, the necessity of enhancing ATP and C-AMP concentrations in B cells was also suggested in order to maintain the insulin secretory ability of cultured pancreatic islets. Am J Obstet Gynecol, 1980 Feb 15, 136(4), 419 - 25 Progesterone-induced glycogen accumulation in human endometrium during organ culture; Shapiro SS et al.; An organ culture system was used to evaluate the response of human proliferative endometrium to progesterone stimulation . The glycogen content of proliferative tissue was increased as much as thirteenfold during organ culture in media containing progesterone . The steroid-induced increase in tissue glycogen was detectable after 16 hours and reached a maximum at 48 and 72 hours of culture . Progesterone induced a significant increase in glycogen at a media concentration of 1.6 x 10(-8)M and a maximal increase at 3.2 x 10(-7) to 3.2 x 10(-6)M . At higher media concentrations of progesterone (1.6 x 10(-5)M), glycogen levels failed to reach the maximum obtainable . The extent of the response correlated poorly with the initial glycogen content of the tissue and not at all with the initial content of high-affinity progesterone-binding sites in cytosol . Addition of estradiol-17 beta to medium (10(-10)M to 10(-7)) has no effect on the progesterone-induced increase in tissue glycogen . Delaying the addition of progesterone to the cultures for 24 hours resulted in a diminished glycogen response; the reduced response may be related to a rapid decrease in high-affinity progesterone-binding sites as measured in cytosol prepared from tissues cultured in the absence of progesterone . High-affinity progesterone-binding sites in endometrial cytosol were found to decrease rapidly during the first 24 hours of culture . The addition of cycloheximide or actinomycin D to the culture media inhibited the increase in tissue glycogen caused by progesterone . These results demonstrate that progesterone can induce an in vitro response in human proliferative endometrium similar to that seen in vivo . The response of the endometrium is reproducible and allows for comparisons between grouped data obtained by using tissues from several different donors. Cell Biol Int Rep, 1980 Feb, 4(2), 185 - 93 Spermine oxidation products are selectively toxic to fibroblasts in cultures of normal human prostatic epithelium; Webber MM et al.; Contamination with fibroblasts has been a major problem in the isolation and establishment of pure cultures of prostatic epithelium, when developing in vitro cell models for studies on carcinogenesis . Addition of spermine to culture media containing fetal bovine serum results in oxidation of spermine to unstable aminoaldehydes which decompose releasing stable acrolein . This oxidation is catalyzed by amine oxidase present in ruminant serum . Prostatic epithelial cells are resistant to these oxidation products at the levels of spermine used . However, these oxidation products are selectively toxic to normal prostatic fibroblasts, which can be eliminated from mixed cultures, and pure cultures of prostatic epithelial cells can thus be established. J Clin Invest, 1980 Feb, 65(2), 268 - 76 Alveolar macrophage-derived chemotactic factor: kinetics of in vitro production and partial characterization; Merrill WW et al.; Alveolar macrophages are the initial phagocytic cells that encounter foreign material and particulates deposited in the terminal airways . We have examined a mechanism by which these cells, after phagocytic challenge, may control or amplify the inflammatory response in lung parenchyma . Normal human alveolar macrophages (AM) were studied from eight subjects . With in vitro culture, AM produced and released two substances into culture media which have potent chemoattractant activity for blood polymorphonuclear granulocytes (PMN) and negligible activity for mononuclear cells . Release of these factors is maximally stimulated by aggregated human immunoglobulin (Ig)G or zymosan particles; however, simple adhesion of the macrophages to plastic surfaces is also sufficient to stimulate release of these chemotactic substances . The larger substance (10,000 daltons) is immunologically distinct from C5a and interacts with a different PMN membrane receptor than that known to exist for formyl-methionyl-leucyl-phenylalanine . Its chemotactic activity is sensitive to the enzymatic effect of trypsin . Although producing a single elution peak on gelfiltration chromatography, electrofocusing in polyacrylamide gels yielded five peaks of radioactivity . Chemotactic activity was localized to a fraction with a pI = 5.0 . The smaller molecular weight substance has been less well characterized . Thus, the human AM can produce at least two factors which attract PMN and this capability may augment the local inflammatory response in the lung. J Clin Endocrinol Metab, 1980 Feb, 50(2), 234 - 9 Ectopic production of pregnancy-specific beta 1-glycoprotein by a nontrophoblastic tumor in vitro; Azer PC et al.; To demonstrate the ectopic production of pregnancy-specific beta 1-glycoprotein (PSbetaG) by a nontrophoblastic tumor, the in vitro secretion of this glycoprotein was evaluated in an hCG-producing ovarian cystadenocarcinoma cell line maintained in long term cell culture . Parallelism was demonstrated between the immunoreactive material present in the tissue culture media and highly purified PSbetaG measured by a sensitive and specific RIA . The immunoreactive material was shown to cochromatograph with purified PSbetaG present in normal term pregnancy serum on a Sephadex G-150 column . The tumor PSbetaG was adsorbed to concanavalin A-Sepharose and eluted with alpha-D-methyl-glucoside in a manner similar to purified PSbetaG . The indirect immunoperoxidase method with anti-PSbetaG sera localized PSbetaG in secretory vesicles and in the endoplasmic reticulum of the tumor cells by light and electron microscopy . The addition of sodium butyrate to the tissue culture media stimulated PSbetaG production in a fashion quantitatively similar to that seen with hCG . The number of PSbetaG-staining intracellular vesicles also increased after exposure to butyrate . These studies provide direct evidence for the ectopic production of PSbetaG by a nontrophoblastic tumor cell line. Cancer Res, 1980 Feb, 40(2), 325 - 8 Procollagens as markers for the cell of origin of human bone tumors; Stern R et al.; Cells derived from osteogenic sarcomas and from Ewing's sarcomas, two malignant bone tumors, were examined for the types of collagens they elaborated into the tissue culture media . Type I procollagen was the predominant species from all osteogenic sarcoma cell lines, a finding consistent with bone cell origin . The Ewing's sarcoma cells contained a prominent peak of type III procollagen and resembled the profile of vascular smooth muscle cells . Fibroblasts derived from skin biopsies taken from amputation specimens synthesized both type I and type III procollagens at the expected ratio of approximately 3:1 . The examination of matrix proteins may provide a general classification scheme for human sarcomas and permit distinction of one tumor from another, as well as from normal fibroblasts. J Bacteriol, 1980 Feb, 141(2), 687 - 93 Cyanophycin granule polypeptide formation and degradation in the cyanobacterium Aphanocapsa 6308; Allen MM et al.; The effect of a number of conditions on the amount of cyanophycin granule polypeptide {multi-L-arginyl poly(L-aspartic acid)} formed in the unicellular cyanobacterium Aphanocapsa 6308 was determined . Light, CO2, sulfur, and phosphorus starvation as well as the addition of arginine to culture media increased the amount of cyanophycin granule polypeptide in cells when compared with that in cells grown under conditions optimal for growth . Nitrogen limitation and reduction of growth temperature to 30 degrees C decreased the amount of cyanophycin granule polypeptide on a dry-weight basis . Shift-up and shift-down experiments suggest cyanophycin granule polypeptide may be a reserve nitrogen polymer in Aphanocapsa 6308. J Cell Physiol, 1980 Feb, 102(2), 267 - 77 Sarcoma growth factor (SGF): specific binding to epidermal growth factor (EGF) membrane receptors; De Larco JE et al.; Cells transformed by murine sarcoma viruses (MSV) produce and release into their tissue culture media several polypeptide growth stimulating factors . One of these has been partially purified using Bio-Gel P-60 column chromatography followed by DEAE-cellulose chromatography . This growth factor was assigned the name sarcoma growth factor (SGF), and is here shown to require the epidermal growth factor (EGF) receptor in order to function as a growth factor . DEAE-cellulose chromatography yielded a product that was several-fold purer than the material present in the Bio-Gel P-60 column pool II . The biologically active material from the DEAE-cellulose column, when labeled with 125I, showed specific binding to EGF membrane receptors . The specific binding could be prevented with the addition of either unlabeled EGF or SGF . Both radiolabeled SGF and EGF will bind to live or fixed cells . We were able to bind 125I-SGF as well as 125I-EGF to fixed cells and elute the bound material from fixed receptors . The eluted SGF showed a greater than 25-fold increase in specific binding . The biological activities of EGF and SGF could be bound to and eluted from fixed receptors . The eluted SGF showed a greater than 25-fold increase in specific binding . The biological activities of EGF and SGF could be bound to and eluted from fixed cells . A 3T3 clone lacking EGF receptors was unable to respond to either EGF or SGF, whereas it responded well to serum and several other purified growth factors . The SGF isolated using DEAE-cellulose chromatography was unable to compete in a radioimmune assay using 125I-EGF and antibody to purified mouse submaxillary gland EGF; it also was not precipitated by anti-EGF antibody . From these studies it appears that the SGF produced and released by these MSV-transformed cells combines with and requires the EGF receptor in order to exert its biological effects . The peptide, however, is antigenically distinct from mouse submaxillary gland EGF. Jpn J Physiol, 1980, 30(5), 767 - 74 Enhancement of erythroid colony formation in vitro by spleen extract from irradiated rats; Kasai S et al.; The effect of spleen extract from irradiated rats on CFU-e and BFU-e colony formation of rat bone marrow cells was investigated by using modified plasma clot culture media . In the presence of erythropoietin (Ep), CFU-e colony formation peaked at 48 hr of culture, and the Ep-induced increase of CFU-e colonies was dose-dependent . The addition of spleen extract enhanced the colony formation more than two-fold in the Ep-containing culture . BFU-e colony formation was also enhanced by the addition of spleen extract . These results indicate that spleen extract from irradiated rats contains factor(s) which stimulates the proliferation of erythroid progenitors. Zentralbl Gynakol, 1980, 102(11), 592 - 5 {Screening method for diagnosis of bacterial infections of the vagina (author's transl)}; Spitzbart H et al.; Reported is the use of TTC-containing culture media . They were used by the authors in screening tests to determine bacterial vaginal flora . The method seems to be promising in terms of better diagnostic and therapeutic results. Folia Parasitol (Praha), 1980, 27(4), 349 - 52 Effects of some sugars, amino acids, glycerol and alpha-glycerophosphate on oxygen uptake of some digenetic trematodes; Nizami WA et al.; Effect of various substrates on the oxygen uptake of some digenetic trematodes -- Srivastavaia indica, Gastrothylax crumenifer, Gigantocotyle explanatum from the buffalo, Bubalus bubalis and Isoparorchis hypselobagri from the fish Wallago attu was studied . In all the four species, the effect of various substrates except maltose was found to be stimulatory . The inhibitory or stimulatory effect of these substrates on oxygen uptake reflects the importance of these compounds in the energy metabolism . In spite of the fact that these trematodes were subjected to similar concentration of substrates, the percentage change in oxygen uptake is different in each of the trematodes, the differential utilization is probably related to the different ecophysiological conditions of the habitat of these trematodes . Such studies can be useful in devising better in vitro culture media for these trematodes. Adv Exp Med Biol, 1980, 128, 635 - 43 Effect of uremic sera on parathyroid hormone (PTH) mediated release of calcium from normal rat embryonal bone maintained in tissue culture; Letteri JM et al.; 1 . 45Ca released from embryonal fetal rat bone into a tissue culture system containing uremic serum was lower than 45Ca measured in culture media containing normal sera . 2 . With stimulation of bone calcium mobilization by the addition of PTH, 1,25(OH)2D3, 24,25(OH)2D3 and 25 (OH)2D3, the 45Ca released into media containing uremic serum was significantly lower than measured in cultures containing normal serum . 3 . Addition of PTH in combination with 24,25(OH)2D3 or with 24,25(OH)2D3 and 1,25(OH)2D3 to bone cultures containing uremic serum increased the 45Ca released into the media when compared to cultures containing uremic serum and PTH. Acta Anat (Basel), 1980, 106(1), 129 - 40 Differentiative ability of the tibial periosteum for the embryonic chick; Scott-Savage P et al.; The claim that the hypertrophic cartilage of a transversely divided 9-day chick tibia is able to induce the differentiation of osteoblasts when placed against the fibrous periosteum of a second, intact, 9-day tibia in vitro, is refuted . Tibia from 7- and 9-day-old chick embryos were transversely sectioned though either the diaphysis or epiphysis and these diaphyseal and epiphyseal grafts were then placed in close association with the fibrous periostea of intact, host tibiae of the same or different ages, at either the diaphyseal or epiphyseal level . All of the sixteen possible combinations of paired tibiae were established in organ culture for 7 days on three types of culture media and two types of atmospheres . In response to the close contact established when diaphyseal grafts were used, the fibrous layer of the periosteum underwent hyperplasia to form fibroblasts at the contact site . Contact of the graft with the periosteum as not sufficient to allow osteoprogenitor cells to accumulate, to differentiate into osteoblasts, or to deposit osteoid . We concluded that the outer, fibrous layer of the periosteum is fibroblastic and not osteblastic. Med Interne, 1980 Jan-Mar, 18(1), 75 - 82 HBs antigen and blood glucose concentration; Lenkei R et al.; HBsAg presence was studied by counterelectrophoresis (CEP) in 76 patients with liver cirrhosis and in 431 patients with diabetes mellitus . A striking correlation was found between high blood glucose values and HBsAg absence . Thus, HBsAg-negative cirrhotics (53%) had significantly higher levels of glycemia than the HBsAg-positive patients of the same age group, i.e . 95.75 +/- 6.36 ng/100 ml compared to 78.30 +/- 10.2/100 ml . This absence of HBsAg was also observed in all diabetics but one . As the incidence of HBsAg (CEP) was found to be of 3.63% in 253,460 subjects from different areas of Romania and 6.84% in 14,690 subjects with various non-hepatic diseases included, the chance of finding the 0.2% HBsAg incidence observed in the diabetics would be less than 0.0002 and 0.0001, respectively . The serum HBsAg absence in cirrhotics with high glycemia and in diabetics