Regulation of cyclooxygenase-2 expression by phospholipase D in human amnion-derived WISH cells

Prostaglandins (PGs) are known to play a key role in the initiation of labor, but the mechanisms regulating their synthesis in amnion are largely unknown. In this study, the regulatory mechanisms for PGE₂ production during phospholipase D (PLD) and p38-dependent activation of WISH cells were investigated. We found that the stimulation of WISH cells with interleukin (IL)-1β elicited dose-dependent synthesis of cyclooxygenase-2 (COX-2) mRNA, protein, and their products, PGE₂. Moreover, the treatment of [³H]myristate-labeled cells in the presence of 1-butanol caused the dose-dependent formation of [³H]phosphatidylbutanol (PBt), a product specific to PLD activity. Pretreating the cells with 1-butanol and Ro 31-8220 inhibited the IL-1β-induced COX-2 expression, but 3-butanol did not affect this response. In addition, evidence that PLD was involved in the stimulation of COX-2 expression was provided by the observations that COX-2 expression was stimulated by the dioctanoyl phosphatidic acid (PA) and that the prevention of PA dephosphorylation by 1-propranolol potentiated COX-2 expression by IL-1β. Moreover, IL-1β stimulation of the cells caused the phosphorylation of p38 and extracellular signal-regulated kinase (ERK), and IL-1β-induced COX-2 expression was inhibited by the pretreatment of WISH cells with a p38 inhibitor, in contrast ERK upstream inhibitor had no effect. Furthermore, Ro 31-8220 inhibited IL-1β-induced p38 phosphorylation but not ERK phosphorylation. The results of this study indicate that in human amnion cells, IL-1β might activate PLD through an upstream protein kinase C to elicit p38 and finally induce COX-2 expression.