PHORBOL ESTER (TPA) REDUCES PROSTAGLANDIN-E2-STIMULATED CAMP PRODUCTION BY DESENSITIZATION OF PROSTAGLANDIN-E2 RECEPTORS IN A CLONAL OSTEOBLAST-LIKE CELL-LINE, MOB 3-4

A clonal osteoblast-like cell line, MOB 3-4, increased cAMP production in response to prostaglandin E2 (PGE2) (5-500 ng/ml). The purpose of this study was to show the effects of tumorpromoting phorbol ester (e.g., 12-O-tetradecanoylphorbol 13-acetate, TPA) on basal and PGE2-stimulated cAMP production and the affinity of PGE2 receptors in the cells. Pretreatment with TPA (1 nM-10-mu-M) for 30 minutes increased basal cAMP production, whereas it markedly reduced the PGE2-stimulated cAMP production in the presence of 0.1 mM isobuthylmethyl xanthine. Both the TPA increase and reduction were dose- and time-dependent. However, TPA exerted no effect on forskolinor cholera toxin-stimulated cAMP production. Copretreatment with TPA and H-7, an inhibitor of protein kinase C (PKC), prevented the TPA-induced increase in basal cAMP production, whereas it did not prevent the reduction of the PGE2-stimulated cAMP production. On the other hand, TPA (0.1-10 mu-M) decreased H-3-PGE2 binding in a dose- and time-dependent manner. Scatchard analysis revealed that TPA decreased the apparent affinity of PGE2 receptors without effect on their apparent number. In addition, 1-oleoyl-2-acetylglycerol (12.6 mu-M), a synthetic diacylglycerol analog, did not mimic the TPA action on H-3-PGE2 binding. Thus, TPA at relatively high concentrations appeared to increase basal cAMP production by a PKC-mediated mechanism, and it appeared to directly act on and thereby reduce the PGE2-stimulated cAMP production in the clonal osteoblast-like MOB 3-4 cell line.