Rapid inactivation of cyclooxygenase activity after stimulation of intact platelets

Abstract

Try sin, thrombin, and ionophore A23187 activate phospholipid breakdown of conditions that lead to use of the latter also results in a similarly rapid inhibition of cyclooxygenase activity, as determined by subsequent challenge with thrombin. Under these conditions lipoxygenase activity is much less markedly inactivated. The arachidonate-induced inhibition of cyclooxygenase activity is not prevented by cyclic AMP. Trypsin does not induce platelet aggregation, and platelets whose cyclooxygenase activity has been inactivated are intact insofar as they are still able to undergo aggregation. These studies demonstrate that operation in intact platelets of the cyclooxygenase pathway, through use of endogenous or exogenous substrate, leads to a very rapid, irreversible inactivation of this enzyme. The lipoxygenase pathway is also progressively impaired, but much less rapidly than the cyclooxygenase enzyme and much less markedly on use of exogenous compared to endogenous substrate. The possible consequences of these physiological processes of spontaneous inactivation are considered.

Arachidonate is a potent modulator of human heat shock gene transcription.

“…Indeed, there is experimental evidence that arachidonate is an inducer of oxidative stress (43)(44)(45), thus leading to damage of preexisting or newly synthesized proteins. Arachidonate affects the activity of numerous proteins as evidenced by its inhibition of enzymes associated with eicoasanoic acid metabolism (33) and its ability to act as an allosteric modifier of the RasGTPase-activating protein and the neurofibromatosis 1 gene product (46). Whether arachidonate or heat shock have in common their effects on the accumulation of malfolded proteins as the mediating signal for HSF-DNA binding remains unclear.…”

Regulation of vascular prostaglandin synthesis by metabolites of arachidonic acid in perfused rabbit aorta.

“…In preparations of ram seminal vesicle microsomes, for example, the oxidation of arachidonic acid leads to a rapid, irreversible inactivation of both cyclooxygenase and prostacyclin synthetase (13,14). The cyclooxygenase of intact platelets undergoes a similar substrate-induced autodeactivation (15). It has been postulated that a highly reactive free radical, probably generated in the reduction of PGG2 to PGH2, is responsible for the irreversible inactivation of these enzyme systems (14,16).…”

Release of platelet-derived growth factor from human platelets by arachidonic acid

“…Whereas these investigators utilized ram seminal vesicle preparations, Lapetina and Cuatrecasas (30) have recently demonstrated a similar phenomenon for the platelet cyclooxygenase. Furthermore, as suggested by the work of Hammarstrom and Falardeau (31), platelet thromboxane synthetase may similarly be sensitive to inactivation during the course of arachidonate metabolism.…”

“…Aggregation was allowed to proceed for 3 min after the addition of the fatty acid salt and was terminated by immersing the samples in an ice-water bath. After aggregation, samples were centrifuged at 12,350 X g for 30 min at 4VC and the supernatant fluids were decanted and retained for assay of the released products. For inhibition studies, a weighed amount of indomethacin (Sigma) was dissolved in 60% (vol/vol) ethanol.…”

“…The platelet dense granule pool was labeled with ['4C]serotonin, and labeled gelfiltered platelets (GFP) were prepared as described (3). The procedure yielded [30][31][32][33][34][35][36][37][38][39][40] ml of labeled GFP containing 1-3 X 108 platelets per ml.…”