摘要:

AbstractThe cell shape of monkey epithelial cells was varied from flat to spheroidal by gradually reducing the substrate adhesiveness with poly (HEMA) films of increasing thickness. The decrease in cell spreading is accompanied by a dramatic response in cellular macromolecular metabolism in the nucleus. Within 14 to 16 hr, DNA and RNA syntheses are inhibited by more than 95%, while the level of protein synthesis is reduced by only twofold after 24 hr in spheroidal‐suspension culture. When epithelial cells, spread to various degrees, are infected with SV40 or herpes virus a parallel inhibition of virus replication and cellular macromolecular metabolism occurs. However, VSV can proliferate in the metabolically active cytoplasm of epithelial cells in which nuclear activity is inhibited owing to alterations in cell shape. The results suggest that the metabolic restrictions imposed on epithelial cells, owing to changes in cell spreading, are a dominant phenomenon that cannot be overcome by virus infection. Rather, virus replication, which is dependent on the cellular metabolic machinery, is inhibited in parallel with the inhibition of cellular macromolecular metabolis

ISSN:0021-9541    

DOI:10.1002/jcp.1041140202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractProstaglandin E1(PGE1), one of the components in the hormone‐supplemented, serum‐free medium for Madin Darby Canine Kidney (MDCK) cells (Medium K‐1), is required for both long‐term growth and for dome formation. Variant cells have been isolated from MDCK populations, which lack the PGE1, requirement for long‐term growth in Medium K‐1. These variants will be useful in identifying the molecular events initiated by PGE1which are necessary for the growth response to be observed. The growth and functional properties of five independently isolated PGE1independent clones have been examined. Normal MDCK cells grew at an equivalent rate in Medium K‐1 and in serum‐supplemented medium; the growth rate was lower in Medium K‐1 lacking PGE1. In contrast, PGE1independent clone 1 grew at an equivalent rate in Medium K‐1 minus PGE1, and in serum‐supplemented medium. When PGE1was added to K‐1 minus PGE1, less growth of PGE1independent clone 1 was observed. A similar observation was made with one other PGE1independent clone which was studied. A hormone deletion study indicated that PGE1independent clone 1 still retained growth responses to the other four supplements in Medium K‐1 (insulin, transferrin, T3, and hydrocortisone). The molecular alterations associated with loss of the PGE1requirement for long‐term growth were examined. At confluency, all of the PGE1independent clones studied had higher intracellular cyclic AMP levels following PGE1treatment, as compared with normal MDCK cells. The increased cyclic AMP levels in the variant cells could result from a number of different types of defects, including reduced cyclic adenylic acid (cyclic AMP) efflux, an increased affinity of PGE2for the PGE1receptor, or a defect in cyclic AMP metabolism. However, in all of the variant clones studied a decreased rate of cyclic AMP degradation by cyclic AMP phosphodiesterase was observed. Thus, the increased cyclic AMP levels in the PGE1independent variants may result from alterations which affect cyclic AMP metabolism. The effect of PGE1on dome formation by the variant cells was also examined. The frequency of dome formation by PGE1independent clone 1 was enhanced in a dosage‐dependent manner, like normal MDCK cells. This observation suggests that PGE1affects MDCK cell growth and dome form

ISSN:0021-9541    

DOI:10.1002/jcp.1041140203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractTunicamycin was found to specifically inhibit the incorporation of a number of sugars into L1210 leukemia cell glycoproteins. This inhibition of glyco‐protein biosynthesis led to a cessation of cell growth which was reversible in a dose‐dependent and time‐dependent manner. After removal of the antibiotic from L1210 cell cultures resumption of sugar incorporation preceded that of thymidine incorporation and the recovery of cell growth. The treatment of cells with tunicamycin resulted in a significant increase in the intracellular pool of UDP‐N‐acetylglucosamine which occurred concurrently with alterations in cell ultrastructure including distentions of the endoplasmic reticulum and nuclear membranes. Similar ultrastructural changes and increases in the intracellular pools of UDP‐sugars were observed in L1210 cells exposed to 5 mMD‐glucosamine, which suggested that the antiproliferative effects of tunicamycin may be related to the accumulation in the endoplasmic reticulum of one or more nucleotide sugar precursors of asparagine‐linked glycoprotein biosynthesis. However, the biological effects of tunicamycin could be distinguished from those caused byD‐glucosamine. Exposure of L1210 cells to tunicamycin resulted in specific alterations in the biochemical composition of the plasma membrane and in the inhibition of cellular agglutination by wheat germ agglutinin which were not apparent following exposure to equitoxic concentrations of the aminosugar. These studies, together with those which demonstrated that recovery of the cellular capacity to synthesize glycoproteins was obligatory for the recovery of cellular proliferation in tunicamycin‐treated cells, suggested that inhibition of the synthesis of glycoproteins was the major factor limiting L1210 leukemic

ISSN:0021-9541    

DOI:10.1002/jcp.1041140204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractJB6 mouse epidermal cells have been selected for resistance to the tumor‐promoting phorbol diester TPA for (1) the plateau density mitogenic (M) response, and (2) the promotion of tumor cell phenotype (P) response. The purpose of this study was to determine the relationship of hexose uptake to the two TPA‐dependent processes. Monolayers of JB6 mouse epidermal cells showing one of four different phenotypes (M + P+, M + P−, M− P+, M− P−) were exposed to 60 nM [3H(G)]2 deoxy‐D‐glucose (2DG) with or without TPA (10 ng/ml) stimulation. The TPA mitogen‐sensitive (M + P +/−) cells, when in logarithmic growth, had a lower basal 2DG uptake rate than TPA mitogen‐resistant (M− P +/−) cells. At plateau density, however, only the M+P+ cells had a significantly lower basal rate. The M + (TPA mitogen‐sensitive) cells (with low basal rates), when preincubated with TPA, exhibited a two to threefold increase in 2DG uptake, while the M− (TPA mitogen‐resistant) lines, which already showed elevated rates, remained unchanged. There was also a positive association between TPA mitogen sensitivity and slower growth rate. These results suggest that low hexose sugar uptake is related to TPA mitogen sensitivity, but not to promotion sensitivity. Hence the cell's ability to increase its uptake rate may be required for the cells to respond

ISSN:0021-9541    

DOI:10.1002/jcp.1041140205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractThe synthesis of the prostaglandins (PG), prostacyclin (PGI2), PGE2, and thromboxane A2(TXA2), has been investigated in actively growing and contact‐inhibited bovine aortic endothelial cell cultures. Cells were stimulated to synthesize prostaglandins by exposure to exogenous arachidonic acid or to the endoperoxide PGH2and by the liberation of endogenous arachidonic acid from cellular lipids with melittin or ionophore A23187. Increased capacity of the cells to synthesize PGI2and PGE2was observed as a function of time in culture, regardless of the type of stimulation. TXA2production increased with time only upon stimulation of the cells with ionophore A23187. This increased PG synthetic capacity was independent of cell density since it was mainly observed in confluent, nondividing endothelial cell cultures. The fact that increased PGI2production in confluent cells was also observed with PGH2, a direct stimulator of PGI2synthetase, implies that this process is independent of the arachidonate concentration within the cells or in the culture medium. This increased capacity is likely to reflect an increased activity of the PG synthetase system associated with the formation of a contact inhibited endothelial cell monolayer. A similar time‐dependent increase in the PGI2production capacity was also observed during growth of cultured bovine corneal endothelial ce

ISSN:0021-9541    

DOI:10.1002/jcp.1041140206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractCultured pig kidney cells designated LLC‐PK1, previously shown to acquire Na+‐dependent concentrative transport of hexoses as the cells become growth arrested, also show Na+‐dependent concentrative uptake of the amino acid analogs α‐aminoisobutyric acid (AIB) and (methyl) meAIB. This A system‐like transport is most active in sparse, growing cultures and becomes stepped down at confluence. The cell/medium equilibrium distribution ratio of the lipophilic cation tetraphenylphosphonium ion (TPP+) decreases in parallel fashion, suggesting that a decrease in membrane potential may be a major factor in the stepdown. Differentiation inducers (hexamethylene bisacetamide) and phosphodiesterase inhibitors (theophylline, methylisobutyl xanthine) accelerate the stepdown, but even in the presence of these compounds addition of the tumor promoter 12‐0‐tetradecanoylphorbol‐13‐acetate (TPA) results in the maintenance of a high level of AIB and meAIB uptake. In all these respects the changes in A system‐like amino acid transport are the reciprocal of those seen for concentrative hexose transport, although the driving force appears to be the same for both systems. The TPA analogs phorbol and 4‐0‐methyl TPA which are inactive in tumor promotion are inactive in this system as well. In confluent, already stepped‐down cultures, addition of TPA leads to a rapid (2–6 hour) stimulation of AIB and meAIB uptake. The enhancement is sensitive to cycloheximide and actinomycin D. The ouabain‐sensitive fraction of meAIB uptake is not markedly changed in the TPA‐enhanced uptake, nor is the TPP+distribution ratio elevated in TPA‐treated cells, making it unlikely that the TPA effect is through an al

ISSN:0021-9541    

DOI:10.1002/jcp.1041140207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractThe possibilities that the growth‐promoting effect of the extracellular matrix (ECM) produced by cultured bovine corneal endothelial (BCE) cells could be due to: (1) adsorbed cellular factors released during the cell lysis process leading to the denudation of the ECM; (2) adsorbed serum or plasma factors: or (3) adsorbed exogenous growth factors have been examined. Exposure of confluent BCE cultures to 2 M urea in medium supplemented with 0.5% calf serum denudes the ECM without cell lysis. The ECM prepared by this procedure supports cell growth just as well as ECM prepared by denudation involving cell lysis. Thus, it is unlikely that the growth‐promoting properties of ECM are due to adsorbed cellular factors. When the ECM produced by BCE cells grown in defined medium supplemented with high‐density lipoprotein, transferrin, and insulin was compared to the ECMs produced by cells grown in the presence of serum‐ or plasma‐supplemented medium, all were found to be equally potent in stimulating cell growth. It is therefore unlikely that the growth‐promoting ability of the ECM is due to adsorbed plasma or serum components. When fibroblast growth factor (FGF)‐coated and ECM‐coated plastic dishes were submitted to a heat treatment (70°C, 30 min) which results in the inactivation of FGF, the growth‐supporting ability of FGF‐coated dishes was lost, while the comparable ability of ECM‐coated dishes was not affected significantly. This observation tends to demonstrate that the active factor present in the ECM is not FGF. Nor is it platelet derived growth factor (PDGF), since treatment known to destroy the activity of PDGF, such as exposure to dithiothreitol (0.1 M, 30 min, 22°C) or to β‐mercaptoethanol (10%) in the presence or absence of 6 M urea for 30 min at 227°C, does not affect the growth‐promoting activity of ECM. It is therefore unlikely that the growth‐promoting effect of ECM is due to cellular growth‐promoting agents or to plasma or se

ISSN:0021-9541    

DOI:10.1002/jcp.1041140208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractPrevious studies have shown that cytosolic glycerol‐3‐phosphate dehydrogenase (GPDH; EC 1.1.1.8) can be induced by glucocorticoids in mammalian brain, mammary gland, and thymus, but it was thought that no induction occurred in liver. We report here that GPDH is induced by glucocorticoids in several lines of hepatoma cells and in rat hepatocytes cultured in vitro. When rat hepatoma cells of clone FU5AH were exposed to 3 μM hydrocortisone (HC) for 3 days, GPDH specific activity increased greater than sixfold over control. The rate and extent of induction were similar in exponentially growing and stationary‐phase cultures of cells. Four other hepatoma cell lines were inducible to a lesser extent, and three lines were not inducible. GPDH was also induced by glucocorticoids in cultures of hepatocytes isolated from livers of 6‐day‐old rats. The enzyme was induced threeto fourfold by the synthetic glucocorticoid, dexamethasone, in the presence of 1 nM insulin, but the induction was not observed in the absence

ISSN:0021-9541    

DOI:10.1002/jcp.1041140209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractThe effect of 12‐O‐tetradecanoylphorbol 13‐acetate (TPA) on human hematopoietic cells has been investigated. It was found that 1–10 ng/ml of TPA totally abrogated erythroid and granulocytic colony growth and, simultaneously in the presence of PHA, stimulated T‐lymphocyte colony formation. TPA concentrations insufficient to inhibit myeloid colony growth also failed to stimulate lymphocyte colony formation. Optimal culture conditions for the growth of these colonies required the presence of TPA, PHA, and leukocyte‐conditioned medium in the cultures. Cells within the colonies were 80–90% E‐rosette positive and by monoclonal antibody characterization contained 45–66% OKT3‐positive cells. Colony‐forming cells were found in both E‐rosette‐positive and‐negative fractions. Although by cell surface marker characterization the cells within the colonies had properties of T‐cells, the exact relationship of cells forming colonies under these conditions to those detected in other T‐cell colony

ISSN:0021-9541    

DOI:10.1002/jcp.1041140210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY

摘要:

AbstractA serum‐free medium for serial culture of baby hamster kidney cell line 21 (BHK‐21) as container‐adherent cells was developed. The medium is a 1:1 (v/v) mixture of Dulbecco's modified Eagle's medium and Ham's F‐12 medium supplemented with fibroblast growth factor, fibronectin, insulin, oleic acid (preincubated with fatty‐acid‐free bovine serum albumin as carrier), and transferrin. The fibronectin was required for cell adherence, the other factors for optimal cell multiplication. When cell input was greater than about 1,900 cells/cm2, this serum‐free medium supported cell multiplication at a rate approximately equal to the rate in medium with 10% serum. At lower cell input, growth in the serum‐free medium was poor unless it was supplemented with serum‐free medium which had been conditioned by BHK‐21 cells. The conditioned medium contained a factor(s) which enabled or stimulated

ISSN:0021-9541    

DOI:10.1002/jcp.1041140211

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1983

数据来源: WILEY