Positive and negative regulation of cell proliferation through prostaglandin receptors in NIH-3T3 cells

Among major eicosanoids and their analogs, prostaglandin (PG) F-2 alpha > PGD(2) > PGE(1) greater than or equal to PGE(2) > iloprost, a stable agonist of PGI(2), dose-dependently stimulated DNA synthesis in quiescent NIH-3T3 cells. PGF(2 alpha), PGD(2), and PGE(2), in that order, formed inositol phosphates and elevated intracellular Ca2+ ([Ca2+](i)) but did not form cAMP nor inhibit forskolin-induced cAMP formation. Iloprost, PGI(2), and PGE(1) induced cAMP formation dose-dependently with an ED(50) Of around 10(-7) M, and PGE(2) at more than 10(-6) M did it. [H-3]PGF(2 alpha) and [H-3]PGD(2) bindings to membranes from NIH-3T3 cells were displaced in the order of PGF(2 alpha) > PGD(2) greater than or equal to PGE(2), while [H-3]PGE(2) binding was displaced by PGE(2) > PGD(2) greater than or equal to PGF(2 alpha). Expression of mRNA encoding EP1 and EP4 (EP2) subtypes could be detected by reverse transcription-polymerase chain reaction using primers specific for EP1 and EP4 (EP2) cDNAs, but not that of EP3 subtype mRNA. The dose dependence of cAMP formation on iloprost and PGI(2) and that of [Ca2+](i) elevation on PGF(2 alpha), D-2, and E(2) were similar to that of [H-3]thymidine incorporation on the corresponding agonists. Fluprostenol (1 mu M), a PGF(2 alpha) receptor agonist > 17-phenyl-trinor-PGE(2) (1 mu M), an EP1 receptor agonist stimulated [H-3]thymidine incorporation, but an EP3 receptor agonist, ONO-AP-324, nor an EP4 (EP2) receptor agonist, 11-deoxy-PGE(1) (1 mu M) did not. iloprost, dibutyryl cAMP, forskolin, or cholera toxin, when applied alone, enhanced [H-3]thymidine incorporation, while they inhibited [H-3]thymidine incorporation induced by submaximal concentrations of PGF(2 alpha) or epidermal growth factor (EGF), when applied within 12 hr after agonist stimulation. These results suggest that the proliferation of NIH-3T3 cells is stimulated by PGs via the PGF(2 alpha) receptor, EP1 subtype of PGE receptor, and the PGI(2)/PGE(1) receptor through [Ca2+](i)- and cAMP-dependent pathways, and that cAMP pathway negatively cross-talks with [Ca2+](i)- or receptor tyrosine kinase-mediated DNA synthesis in a cell cycle-dependent manner. (C) 1996 Wiley-Liss, Inc.