Interleukin-1 beta induces the synthesis and activity of cytosolic phospholipase A2 and the release of prostaglandin E2 in human amnion-derived WISH cells.

The objective of this study was to examine the expression and activity of cytosolic phospholipase A2 (cPLA2) in relation to prostaglandin E2 (PGE2) synthesis in human amnion-derived WISH cells in response to stimulation by interleukin-1 beta (IL-1 beta). cPLA2 activity was characterized by sensitivity to heat and acid treatment, stability to dithiothreitol, and inhibition by the specific inhibitor, arachidonyl trifluoromethyl ketone (AACOCF3). Treatment of WISH cells with IL-1 beta (0.01-1 ng/mL) for up to 24 h resulted in a significant increase in PGE2 release in a concentration- and time-dependent manner accompanied by increases both in total cellular cPLA2 activity and in cPLA2 protein levels detected by Western blot analysis. The parallel increase in total cellular cPLA2 activity and cPLA2 protein level indicates that IL-1 beta may induce the synthesis of cPLA2. Incubation of the cells with 10 microM AACOCF3 for 24 h significantly inhibited IL-1 beta-induced PGE2 production strongly suggesting that cPLA2 mediates IL-1 beta-induced PGE2 formation. In unstimulated cells, there is appreciable total cellular cPLA2 activity and protein, but these cells produce low amounts of PGE2 until stimulated by IL-1 beta, suggesting that cPLA2 translocation from cytosol to the membrane is necessary for its bioactivity. In contrast to IL-1 beta, treatment with phorbol ester (12-O-tetradecanoyl phorbol-13-acetate, TPA, 10(-10)-10(-6)M) for 24 h significantly inhibited total cellular cPLA2 activity in a concentration-dependent manner. The amount of total cellular cPLA2 protein seen on Western blot remained unchanged following TPA treatment. These data suggest that in WISH cells, IL-1 beta induces both translocation to the membrane and de novo synthesis of cPLA2 protein to sustain prostaglandin (PG) synthesis. In contrast, TPA may only cause cPLA2 translocation but no increase in cPLA2 protein synthesis, resulting in limited PG synthesis. Our results provide a mechanism for the effect of IL-1 beta on prostaglandin synthesis in human amnion cells and provide support for a role of cPLA2 in the mechanism initiating human parturition.