Conjugated linoleic acid exhibits stimulatory and inhibitory effects on prostanoid production in human endothelial cells and platelets

摘要

In addition to their reported antitumorigenic properties, various conjugated linoleic acid (CLA) isomers have also been shown to decrease prostanoid synthesis as a result of inhibiting the cyclooxygenase (COX) enzyme. We have previously reported that several CLA isomers inhibited both platelet aggregation and formation of thromboxane A_2 (TXA_2), a proaggregatory and vasoconstrictive agent. Since the interaction between platelets and vascular endothelial cells is essential to maintaining vascular homeostasis, we decided to investigate the effects of various CLA isomers on the production of endothelial prostacyclin (PGI_2), a potent vasodilator and inhibitor of platelet function. Using interleukin 1-β (IL1-β)-stimulated human umbilical vein endothelial cells (HUVECs), we initially established that HUVECs of passage #2 should be used since these cells were most responsive to thrombin-induced conversion of endogenous arachidonic acid to PGI_2, as monitored by the formation of its stable, inactive metabolite, 6-ketoPGF_(1α). In the first part of the study, the effects of CLA isomers in the free fatty acid form were tested. The 10(E), 12(Z)- and 9(Z), 11(E)-CLA isomers inhibited thrombin-induced 6-ketoPGF_(1α) formation with I_(50)'s of 2.6 and 5.5 μM, whereas the 9(Z), 11(Z)- and 9(E), 11(E)-CLA were ineffective at concentrations up to 60 μM. The inhibitory effect of the 10(E), 12(Z)-CLA was irreversible. Next, the effects of CLA incorporation into HUVECs on PGI_2 generation was determined. An average 8-fold stimulation of 6-ketoPGF_(1α) formation was obtained with quiescent IL1-β-exposed HUVECs pretreated for 18 h with 25 μM 9(Z), 11(Z)-CLA, whereas cells preincubated with the 10(E), 12(Z) isomer enhanced this eicosanoid 3-fold. Such IL1-β-treated HUVECs prelabed with 25 μM 9(Z), 11(Z)-CLA became refractory to thrombin stimulation, as measured by 6-ketoPGF_(1α) production, whereas a small, statistically insignificant, inhibition was observed upon thrombin treatment of HUVECs prelabeled with the 10(E), 12(Z) isomer. Qualitative similar results were obtained with resting or thrombin-stimulated platelets containing these esterified CLA isomers indicating that these effects occur with cells that contain either the COX-1 or COX-2 isozymes. The results of this in vitro study indicate that the effects of CLA on cellular prostanoid formation in endothelial cells and platelets can be either inhibitory or stimulatory, and this seems to depend not only on the specific CLA isomer and whether or not the CLA is in the free fatty acid form or esterified into cellular lipids, but also whether cells are in the resting or stimulated state. These findings suggest that in vivo, CLA might have multiple, complex effects on vascular homeostasis.