SEGREGATED COUPLING OF PHOSPHOLIPASES A(2), CYCLOOXYGENASES, AND TERMINAL PROSTANOID SYNTHASES IN DIFFERENT PHASES OF PROSTANOID BIOSYNTHESIS IN RAT PERITONEAL-MACROPHAGES

We examined herein the functional linkage of enzymes regulating the in itial, intermediate, and terminal steps of PG biosynthesis to provide PGs in rat peritoneal macrophages stimulated with LPS and/or A23187, Q uiescent cells stimulated with A23187 produced thromboxane B-2 (TXB2) in marked preference to PGE(2) within 30 to 60 min (constitutive immed iate response), which was mediated by preexisting cytosolic phospholip ase A(2) (cPLA(2)), cyclooxygenase-1 (COX-1), and TX synthase. Cells t reated with LPS predominantly produced PGE(2) during culture for 3 to 24 h (delayed response), where cPLA(2) and secretory PLA(2) functioned cooperatively with inducible COX-2, which was, in turn, coupled with inducible PGE(2) synthase, Cells primed for 12 h with LPS and stimulat ed for 30 min with A23187 produced PGE(2) in marked preference to TXB2 (induced immediate response), in which three inducible enzymes, cPLA( 2), COX-2, and PGE(2) synthase, were functionally linked, Preferred co upling of the two inducible enzymes, COX-2 and PGE(2) synthase, was fu rther confirmed by the ability of LPS-treated cells to concert exogeno us arachidonic acid to PGE(2) optimally at a time when both enzymes we re simultaneously induced, These results suggest that distinct PG bios ynthetic enzymes display segregated functional coupling following diff erent transmembrane stimulation events even when enzymes that catalyze similar reactions in vitro coexist in the same cells.