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Biomedical subjects

N Savion

Publications and source records attributed to N Savion.

121 records · Page 7Linked to original sources
PubMed1981

Effect of cell density on thrombin binding to a specific site on bovine vascular endothelial cells.

We studied thrombin binding to proliferating and confluent endothelial cells derived from bovine vascular endothelium. [125]thrombin was incubated with nonconfluent or confluent endothelial cells and both the total amount bound and the amount linked in a 77,000-dalton thrombin-cell complex were determined. Approximately 230,000 molecules of thrombin bound per cell in nonconfluent cultures compared to 12,800 molecules per cell in confluent cultures. Approximately 67,7000 thrombin molecules were bound in an apparently covalent complex, Mr = 77,000, with each cell in sparse cultures, whereas only 4,600 thrombin molecules per cell were bound in this complex with confluent cultures. Similar studies with [125I]thrombin and endothelial cells derived from bovine cornea revealed no difference either in the total amount of thrombin bound or in the amount bound in the 77,000-dalton complex using sparse or confluent cultures. When confluent vascular endothelial cultures were wounded, additional cellular binding sites for the 77,000-dalton complex with thrombin appeared within 24 h. A 237% increase in the amount of thrombin bound to these sites was induced by a wound which resulted in a 20% decrease in cell number in the monolayer. There was no significant increase in thrombin binding to other cellular sites at 24 h. These experiments provide evidence that the first change in thrombin binding after injury is an increase in the cellular sites involved in the 77,000-dalton complex, and suggest that thrombin binding to endothelial cells may be important in the vascular response to injury.

Animals↗
PubMed1981

Factors controlling proliferation and progesterone production by bovine granulosa cells in serum-free medium.

Bovine granulosa cells seeded in the presence of serum on extracellular matrix-coated dishes proliferate actively when exposed to serum-free medium supplemented with insulin (2 microgram/ml), fibroblast growth factor (FGF, 100 ng/ml), and high density lipoprotein (HDL, 30 microgram protein/ml). The final density of the cultures is 80-120% that of cultures grown in the presence of medium supplemented with optimal concentration (10%) of calf serum. Insulin has the greatest effect on cell proliferation when added alone to serum-free medium, since it induced an increase in cell number that was 35-60% that observed with optimal serum concentration. Somatomedin C can replace insulin when added alone. FGF, epidermal growth factor, or HDL had no significant effect on cell proliferation by themselves. When these factors were added together with insulin, they acted synergistically in stimulating cell proliferation. When cultures were seeded in the total absence of serum, the addition of transferrin (10 microgram/ml) to serum-free medium was required in order for insulin and FGF to be mitogenic. Cultures maintained on extracellular matrix and exposed to serum-free medium alone have a lifespan in culture of 4 generations. Addition of insulin, FGF, and HDL increases the lifespan of the cultures to 12 generations. Bovine granulosa cells, which proliferate in a defined medium, respond to dibutyryl cAMP by releasing progesterone into the medium. Addition of FSH to the defined medium resulted in a 30% decrease in cell proliferation and in a 2.1-fold increase in the amount of progesterone released into the medium in response to dibutyryl cAMP. This release of progesterone reached a level similar to that observed with cultures grown in medium supplemented with optimal concentration of serum and exposed or not to FSH during their growth phase and at confluence. These results demonstrate that bovine granulosa cells can actively proliferate in a serum-free medium and maintain their differentiated function, as indicated by their ability to produce progesterone.

Animals↗
PubMed1980

Role of the degradation process in the mitogenic effect of epidermal growth factor.

The protease inhibitor leupeptin inhibits the degradation process of (125)I-labeled epidermal growth factor ((125)I-EGF) by cultured bovine granulosa cells. At 80 mug/ml, leupeptin inhibited the appearance of degradation products of (125)I-EGF in the medium by 95% during 1 hr of incubation and by 90% during 24 hr of incubation when the cells were exposed to 5 ng of (125)I-EGF per ml. In contrast, cultures exposed to either saturating (10 ng/ml) or nonsaturating (0.1 ng/ml) concentrations of EGF in the presence of leupeptin (80 mug/ml) exhibited an increase in DNA synthesis that was 70-80% that of cultures exposed to EGF alone. Cultures responded to either EGF or fibroblast growth factor with a logarithmic increase in cell number and, over a period of 8 days, the number of cells increased 10- to 18-fold. Addition of leupeptin did not diminish the growth rate of the cultures. In the presence of leupeptin, (125)I-EGF accumulated within the granulosa cells and was in a form that was precipitable with antiserum against EGF and that comigrated on isoelectric focusing with native (125)I-EGF. That a full mitogenic response can be obtained despite a 90-95% inhibition of EGF degradation at either saturating or nonsaturating concentrations of the mitogen suggests that a proteolytic degradation of a given mitogen may not be involved in the induction of a proliferative response.

Animals↗
PubMed1980

Patterns of cellular peptide synthesis by cultured bovine granulosa cells.

The electrophoretic distribution of the polypeptides synthesized by bovine granulosa cell cultures after metabolic labeling with [35S]methionine has been analyzed by double gel electrophoresis. The fluctuations of 35 polypeptides have been followed as a function of the size of the follicles from which cultures originated, as a function of the cultures' proliferative stage (sparce, actively growing vs. confluent, resting cultures), and finally as a function of whether cells were exposed to either epidermal or fibroblast growth factor. When the patterns of protein synthesis in sparce vs. confluent granulosa cell cultures derived from small-sized follicles were compared, only a few differences were observed. In confluent cultures, 6 new peptides appeared, while 1 peptide present in sparce cultures disappeared. Cultures maintained in the presence of fibroblast or epidermal growth factor synthesized 20 new peptides upon reaching confluence. Among these were the 6 new peptides present in confluent but not in sparse granulosa cell cultures maintained in the absence of growth factors. The changes in protein synthesis observed in cultures grown in the presence of growth factors may reflect their direct effect on the cellular metabolism. A comparison between the protein distribution in cells derived from small- vs. large-sized follicles showed that fewer proteins were ultimately produced at confluence in cells derived from large-size follicles than in cells derived from small-sized follicles. This could be related to the process of cellular differentiation taking place within granulosa cells. The patterns of [35S]methionine-labeled proteins secreted by the granulosa cells into the incubation medium were analyzed and found to be similar regardless of the size of the follicle from which the culture originated. Little similarity between the proteins present in the follicular fluid and the pattern of the labeled proteins secreted into the incubation medium by cultured granulosa cells was observed. Only three proteins were identified which comigrate with proteins present in the follicular fluid. One of these was identified as fibronectin. This raises the possibility that the fibronectin present in the follicular fluid originated from granulosa cells and is not derived from plasma.

Animals↗
PubMed1978

Morphological changes in rat pancreatic slices associated with inhibition of enzyme secretion by high concentrations of secretagogues.

Stimulation of enzyme secretion in rat pancreatic slices by cholinergic agonists or by cholecystokinin-pancreozymin (CCK-PZ) and its peptide analogs showed a biphasic dose response curve. The optimal concentrations eliciting an efficient rate of enzyme secretion were 1 microM for carbamylcholine or acetylcholine, and 5 nM and 20 nM for CCK-PZ octapeptide and CCK-PZ, respectively. At higher concentrations of secretagogues, however, the rate of secretion progressively declined, and almost complete inhibition was achieved at 1 mM of carbamylcholine or acetylcholine and at 0.1 microM of CCK-PZ or its octapeptide analog. Atropine displaced the dose-response curve for carbamylcholine to the right so that in the presence of 7 microM atropine a concentration of 1 mM carbamylcholine now gave an optimal rate of enzyme secretion. The ionophore A-23187 which bypasses the receptor and elicits enzyme secretion did not relieve the inhibition caused by supraoptimal concentrations of secretagogues, indicating that the inhibition occurs at the cellular rather than at the receptor level. Secretin had no effect on the inhibition of enzyme secretion by a high concentration of carbamylcholine, indicating that the inhibition was not caused by lack of water and electrolyte secretion. The energy-producing metabolism was not affected since the ATP level in the pancreatic slices was the same in the presence of either inhibitory or optimal concentrations of secretagogues. The inhibition of enzyme secretion was reversible since restoration of efficient enzyme secretion occurred after removal of carbamylcholine (1 mM) by washing, followed by addition of an optimal concentration of CCK-PZ octapeptide. Morphological studies revealed that the presence of inhibitory concentrations of secretagogues caused severe distortion of the lumen structure: disruption of the filamentous system surrounding the lumen, disappearance of microvilli, and production of distended evaginations of the luminal membrane containing cellular material. These changes eventually caused a reduction in the size of the lumen which becomes plugged with secretory material. It is suggested that these changes in the microtubular microfilamentous system could account for the inhibition of enzyme secretion.

Acetylcholine↗
PubMedDate not supplied

Binding of apolipoprotein A-I and apolipoprotein A-IV to cultured bovine aortic endothelial cells.

Adult bovine aortic endothelial (ABAE) cells specifically bound 125I-labelled high density lipoprotein3 (125I-HDL3), and saturation of the binding was observed at a concentration of about 50 to 100 micrograms protein/ml. The binding of purified apolipoproteins (apo) A-I and A-IV to cultured ABAE cells and the correlation of this binding with the binding of HDL to the cells was studied. ABAE cells bound 125I-labelled apo A-I (125I-apo A-I) and after a 3-hour exposure to the ligand, most of the radioactivity (82%) was associated with the cell surface and was accessible for trypsinization. Saturation of 125I-apo A-I binding was observed at a concentration of 3 micrograms/ml; each cell possessed 1.38 X 10(5) high affinity binding sites (Kd = 2.04 X 10(-8) M). The cultures also bound 125I-labelled apo A-IV (125I-apo A-IV), and saturation of the binding was observed at a concentration of 2 micrograms/ml. Each cell possessed 5.4 X 10(4) high affinity binding sites for apo A-IV (Kd = 1.45 X 10(-8) M). Addition of either excess unlabelled apo A-I or excess unlabelled apo A-IV competed with the binding of both iodinated apo A-I and apo A-IV. It suggests that apo A-I and A-IV share the same binding site on endothelial cells. HDL also successfully competed with the binding of apo A-I and A-IV to ABAE cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗