Altered regulation of arachidonic acid metabolism by SV40 immortalized human urothelial cells.
Regulation of arachidonic acid metabolism was investigated in an SV40 immortalized, non-tumorigenic human urothelial cell line (SV-HUC). This cell line is being used to evaluate the multistage carcinogenic process. Media from confluent cultures were analyzed for radioimmunoassayable prostaglandin E2 (PGE2). A variety of agonists, including 12-O-tetradecanoylphorbol-13-acetate (TPA) and A23187 were tested and did not increase PGE2 synthesis within 2 h of addition. This was not due to the lack of prostaglandin H synthase activity because addition of exogenous arachidonic acid increased PGE2 synthesis. Cultures prelabeled overnight with [3H]arachidonic acid failed to increase the release of radioactivity following agonist addition. Thus, the lack of an early response in SV-HUC appears to be due to decreased release of endogenous arachidonic acid by phospholipase(s). After a 24 h incubation with 0.1 microM TPA or 1.0 microM A23187, the addition of arachidonic acid elicited significantly more PGE2 synthesis in agonist-treated cells than it did in control cells. Microsomes from 24 h TPA-treated cells produced approximately 15-fold more PGE2 than did those from control cells. In addition, the PGE2 content of overnight media was significantly greater in TPA-treated cells than in control cells. The 24 h agonist response was blocked by cycloheximide and staurosporine--inhibitors of protein synthesis and protein kinase C respectively. Pretreatment of cells with aspirin, an irreversible inhibitor of prostaglandin H synthase, prior to addition of TPA did not prevent the late 24 h TPA-mediated increase in PGE2 synthesis. Results suggest that this late effect of TPA is due to de novo synthesis of prostaglandin H synthase. Thus, SV-HUC has lost the early but retains the late response to agonists.
Thomas
D J
DJ
VA Medical Center, St Louis, MO.
Jacob
A K
AK
Reznikoff
C A
CA
Zenser
T V
TV
Davis
B B
BB
D016718:Arachidonic Acid / Q000378:metabolism*; D001241:Aspirin / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology*; D002461:Cell Line, Transformed; D015232:Dinoprostone / Q000096:biosynthesis*; D004224:Diterpenes; D008861:Microsomes / Q000378:metabolism; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis; D013539:Simian virus 40; D013729:Terpenes / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D001743:Urinary Bladder / Q000166:cytology
D004224:Diterpenes; D013729:Terpenes; D016718:Arachidonic Acid; C011309:mezerein; D000001:Calcimycin; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate; D001241:Aspirin; D001920:Bradykinin