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

Summary 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 tumor-promoting 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 μM) 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 μM) decreased³H-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 μM), a synthetic diacylglycerol analog, did not mimic the TPA action on³H-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 PGE₂ receptors to decrease their apparent affinity and thereby reduce the PGE₂-stimulated cAMP production in the clonal osteoblast-like MOB 3-4 cell line.