Revised Manuscript Received October 2, 1989 abstract: Eicosanoids have been implicated inthe regulation of arterial smooth muscle cell (SMC) cholesteryl ester (CE) metabolism. These eicosanoids, which include prostacyclin (PGI2), stimulate CE hydrolytic activities. High-density lipoproteins (HDL), which promote cholesterol efflux, also stimulate PGI2 production, suggesting that HDL-induced cholesterol efflux is modulated by eicosanoid biosynthesis. To ascertain the role of endogenously synthesized eicosanoids produced by arterial smooth muscle cells in the regulation of CE metabolism, we examined the effects of cyclooxygenase inhibition on CE hydrolytic enzyme activities, cholesterol efflux, and cholesterol content in normal SMC and SMC-derived foam cells following exposure to HDL and another cholesterol acceptor protein, serum albumin. Alterations of these activities were correlated with cholesterol efflux in response to HDL or bovine serum albumin (BSA) in the presence or absence of aspirin. HDL stimulated PGI2 synthesis and CE hydrolases in a dose-dependent manner. Eicosanoid dependency was established by demonstrating that HDL-induced acid cholesteryl ester hydrolase (ACEH) activity was blocked by aspirin. CE enrichment essentially abrogated HDL-induced PGI2 production in cells which also exhibited decreased lysosomal and cytoplasmic CE hydrolase activities. In CE-enriched cells whose cytoplasmicCE pool was metabolically labeled with [3H] oleate or cLDL containing [3H] cholesteryl linoleate, aspirin did not alter HDL-or BSA-induced net CE hydrolysis or efflux, respectively. Finally, aspirin treatment did not alter the mass of either free or esterified cholesterol content of untreated or CE-enriched SMC following exposure to acceptor proteins. These data demonstrated that CE enrichment significantly reduced HDL-induced activation of CE hydrolytic activity via inhibitionof endogenous PGI2 production. These novel findings support our hypothesis that CE hydrolysis is under eicosanoid metabolic control. We suggest that CE enrichmentablates eicosanoid-dependent control of CE hydrolysis. However, net cholesterol efflux from intracellular pools appeared to be independent of eicosanoid biosynthesis. Therefore, a dissociation exists between eicosanoid-dependent CE hydrolysis and net cholesterol efflux induced by plasma cholesterol acceptors. e mechanisms by whichcellular cholesterol content is regulated havebeen the subject of intense research. Cellular cholesterol content is stringently maintained through (1) uptake by the low-density lipoprotein (LDL) 1 receptor (Goldstein & Brown, 1977; Steinberg, 1987; Chen et al., 1988),(2) cholesterol synthesis via HMG CoA-reductase (Brown et al., 1981), and (3) efflux, which is mediated by plasma and interstitial acceptors including albumin (Fielding & Moser, 1981; Chau & Geyer, 1978; Bartholow & Geyer, 1982) and high-density lipoprotein (HDL)(Rothblat & Phillips, 1982; Oram, 1983; Stein et al., 1980; Innerarity et al., 1982). Recent evidence suggests that eicosanoids also influence cellular cholesterol metabolism. First, exogenous prostacyclin (PGI2) activates CE hydrolytic activities and reduces cellular cholesterol content in intact smooth muscle cells (Hajjar, 1985; tThis work was funded in part by the NIH SCOR in Thrombosis (HL-18828), NIH R01 HL-39701, NIH Biomedical Research Support Grant Program (BRSG S07 RR05396), an American Heart Association grant-in-aid (87-813), and an American Health Assistance Foundation Grant in Coronary Artery Disease Research (Rockville, MD). KBP is the recipient of an NIH New Investigator Award (HL-38277) and the …