Regulation of dog urothelial cell arachidonic acid release and prostaglandin E2 synthesis.
The effects of various agonists on prostaglandin E2 (PGE2) synthesis and arachidonic acid release were evaluated to identify factors which regulate urothelial cell responsiveness. Techniques for harvesting and culturing urothelial cells from the canine bladder were developed. Although serum had little effect on growth, it was required for arachidonic acid, 12-O-tetradecanoylphorbol-13-acetate (TPA) or A23187 to elicit increases in PGE2 production. Fetal calf serum (FCS), bovine calf serum (BCS) and heated BCS (each at 1%) were equally effective in supporting prostaglandin production. The optimum concentration range for the effect of FCS was 0.3-1.0% with 7.0% being less effective. The lack of agonist responsiveness observed with no serum could be reversed by adding serum at days 2 or 6 of a total of 9 days in culture. Subculturing cells dramatically reduced responsiveness to all stimulators tested. Bradykinin-stimulated release of arachidonic acid was maximal within 15 min, while the TPA release continued throughout the 120-min study. TPA response was inhibited by cycloheximide and actinomycin D. Neither agent altered the response to arachidonic acid. Combinations of arachidonic acid with TPA were neither additive nor synergistic. Responses to arachidonic acid, TPA and A23187 were optimum when the cells were near confluency. NaF, epidermal growth factor and mezerein also stimulated PGE2 synthesis. Tumor-promoting, but not the non-tumor-promoting phorbol esters (4 alpha-phorbol-12,13-didecanoate), increased PGE2 synthesis. Thus, the arachidonic acid cascade in dog urothelial cells is a hormonal responsive system which provides a method for evaluating transmembrane signaling.
Wong
Y H
YH
Geriatric Research, Education and Clinical Center, VA Medical Center, St Louis, MO 63125.
Zenser
T V
TV
Davis
B B
BB
D000818:Animals; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism*; D001920:Bradykinin / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002455:Cell Division; D002478:Cells, Cultured; D003470:Culture Media; D003513:Cycloheximide / Q000494:pharmacology; D015232:Dinoprostone / Q000096:biosynthesis*; D004285:Dogs; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007700:Kinetics; D008854:Microscopy, Electron; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D001743:Urinary Bladder / Q000187:drug effects / Q000378:metabolism*
D001095:Arachidonic Acids; D003470:Culture Media; D016718:Arachidonic Acid; D000001:Calcimycin; D003513:Cycloheximide; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin