Somatomedin synthesis by a subclone of Buffalo rat liver cells: characterization and evidence for immediate secretion of de novo synthesized hormone.
Previous studies have reported that cultured Buffalo rat liver (BRL-3A) cells synthesize and release a somatomedin called multiplication-stimulating activity, a major component of which is rat insulin-like growth factor-II (rIGF-II). We have isolated a subclone of hepatocytes (BRL-3SC) from these cells, which release rIGF-II into serum-free medium at a similar but considerably more uniform rate than BRL-3A cells, and have described their morphology and defined some of the conditions regulating the rate of synthesis and secretion of rIGF-II. Time-course experiments indicate that after a 2-h lag, rIGF-II is released at a nearly constant rate of 70 microU insulin-like activity/ml . 24 h for 5 days. These cells demonstrate considerable resistance to variations in environmental pH, with extremes of pH 6.8-8.0 in the initial culture medium causing a negligible effect on rIGF-II release. We found no evidence for the existence of an appreciable amount of intracellular rIGF-II, consistent with the lack of secretion granules in these cells. Moreover, at concentrations not causing irreversible cell damage, inhibitors of protein synthesis, cycloheximide and actinomycin D, produced 87% and 76% reductions, respectively, in rIGF-II release in 24 h, and inhibitors of oxidative phosphorylation, dinitrophenol and sodium azide, reduced rIGF-II release by 72% and 78%, respectively. We conclude from these studies that BRL-3A cells, and particularly those of the subclone BRL-3SC herein described, may provide an excellent model for investigations of somatomedin synthesis and release and that such studies are facilitated by the fact that rIGF-II released into serum-free medium appears to be the immediate product of de novo peptide synthesis.
D000818:Animals; D001386:Azides / Q000494:pharmacology; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002999:Clone Cells; D003513:Cycloheximide / Q000494:pharmacology; D004247:DNA / Q000032:analysis; D004140:Dinitrophenols / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D010085:Oxidative Phosphorylation; D051381:Rats; D011915:Rats, Inbred BUF; D019810:Sodium Azide; D013002:Somatomedins / Q000096:biosynthesis* / Q000378:metabolism
D001386:Azides; D004140:Dinitrophenols; D013002:Somatomedins; D004247:DNA; D019810:Sodium Azide; D003513:Cycloheximide