Role of EP(1) and EP(4) PGE(2) subtype receptors in serum-induced 3T6 fibroblast cycle progression and proliferation.

Recent studies have suggested that prostaglandin E(2) (PGE(2)) subtype receptors (EP) are involved in cellular proliferation and tumor development. We studied the role of EP(1) and EP(4) PGE(2) subtype receptor antagonists AH-6809 and AH-23848B, respectively, in serum-induced 3T6 fibroblast proliferation. This was significantly reduced in a dose-dependent manner (IC(50) approximately 100 and approximately 30 microM, respectively) to an almost complete inhibition, without any cytotoxic effect. However, the effect of each antagonist on 3T6 cell cycle progression clearly differed. Whereas the EP(1) antagonist increased the G(0)/G(1) population, the EP(4) antagonist brought about an accumulation of cells in early S phase. These effects were associated with a decrease in cyclin D and E levels in AH-6809-treated 3T6 cells and lower cyclin A levels in AH-23848B-treated fibroblasts with respect to control cells. The G(0)/G(1) accumulation caused by AH-6809 seems to be intracellular Ca(2+) concentration ([Ca(2+)](i)) dependent, because a 6-h 1 microM thapsigargin treatment allowed G(0)/G(1)-arrested cells to enter S phase. Similarly, treatment with 20 microM forskolin for 6 h allowed S-phase and G(2)/M progression of AH-23848B-treated cells. This study shows that the inhibitory effect of the EP(1) and EP(4) antagonists on serum-induced 3T6 fibroblast growth is due to their effect at various levels of the cell cycle machinery, suggesting that PGE(2) interaction with its different subtype receptors regulates progression through the cell cycle by modulating cAMP and [Ca(2+)](i).