Plasma stimulation of prostacyclin production by rat smooth muscle cells requires previous induction of phospholipase activity

Abstract Human normal platelet poor plasma (PPPn) stimulates prostacyclin (PGI 2 ) production in a dose-dependent manner and after 15 to 60 min of incubation time when confluent rat smooth muscle cells (RSMC) were preincubated for 24 hours with fresh culture medium. This PGI 2 production was independent of new protein synthesis, and was not observed in the cells maintained only in exhausted medium. The serum of fresh culture medium also induced a significant and transient increase of prostaglandin endoperoxide synthase (PGHS) activity as a function of preincubation time, which was dependent of protein synthesis. However, neither PGHS activity nor arachidonic acid availability limited the PPPn induced PGI 2 synthesis in RSMC. Moreover, the previous addition of phorbol 13-myristate acetate also allowed the PPPn to induce PGI 2 synthesis, that was significantly inhibited by a specific phospholipase A 2 inhibitor. Furthermore, we found that PPPn induced a significant increase of intracellular calcium, and also stimulated PGI 2 production at short incubation times due to its effect on phospholipases, and not by a direct supply of substrate. We conclude that a previous activation of phospholipase A 2 was necessary to observe a significant and sustained PGI 2 synthesis induced by PPPn in RSMC, and that the increase of intracellular calcium observed with PPPn might stimulate these previously activated phospholipases.