[Arachidonic acid distribution and release in human and rat blood platelets: importance of transacylases]. / Distribution et libération de l'acide arachidonique dans les plaquettes humaines et de Rat: importance des transacylases.

Human platelets release about 5 fold more arachidonate than rat platelets when they ar triggered with a high dose of thrombin. Total arachidonate content of the phospholipids was not significantly different between the two species. In contrast, phosphatidylcholine (PC) from human platelets exhibited twice more arachidonate than PC from rat platelet and opposite arachidonate contents were found in phosphatidylethanolamine. Moreover, rat platelet PC was very rich in disaturated species, primarily dipalmitoyl whereas high levels of oleate and linoleate were present in human platelets PC. The differences in the fatty acids content of the two species are the result of CoA-independent and CoA-dependent transacylase activities which are more efficient in rat than in human platelets and could account for the low level of arachidonate released from rat platelets.

Very high proportion of disaturated molecular species in rat platelet diacyl-glycerophosphocholine: involvement of CoA-dependent transacylation reactions.

The molecular species composition of rat platelet diacyl-glycerophosphocholine (GPC) was investigated by reverse-phase HPLC and by mass spectrometry. The two methods gave the same very high proportion of fully saturated phospholipids, the 16:0-16:0 and 16:0-18:0 species representing together about 40% of the overall molecular species. [14C]Palmitoyllyso-GPC was found to be acylated by resting platelets in equal amounts into 16:0-16:0 and into 16:0-20:4 species. The acylation rate of this lysophospholipid was increased by 3-fold and 14-fold when platelets were stimulated for 10 min with thrombin and the ionophore A23187, respectively. Essentially the same two molecular species were synthesized upon stimulation but with a higher preference for arachidonate than for palmitate. We investigated the mechanisms responsible for the incorporation of palmitate and arachidonate by examining the enzymatic acylation of [14C]palmitoyllyso-GPC by platelet homogenates. The percentage of the various molecular species formed when CoA, ATP, and Mg2+ were added excludes the CoA, ATP-dependent pathway as being involved in the acylation reactions previously observed. In the absence of ATP, CoA-independent transacylations appear to play a crucial role in the synthesis of the 16:0-20:4 species whereas the addition of CoA greatly favored dipalmitoyl-GPC synthesis. The involvement of CoA-dependent mechanisms in the synthesis of dipalmitoyl-GPC was demonstrated as follows: (i) the labeling in the sn-2 position of the dipalmitoyl-GPC synthesized in the presence of CoA was not modified when free unlabeled palmitic acid was added to the incubation medium and (ii) platelet homogenates were unable to esterify lysolecithin with added labeled palmitic acid in the presence of CoA only.

Ca2+ effect on ATP-independent lysophospholipid transacylases is indissociable from Ca2+ effect on phospholipase A2 activity in rat platelet sonicates.

CoA-dependent transacylation and phospholipid hydrolysis were studied in parallel experiments using rat platelet sonicates. The decrease observed in palmitoyllyso-sn-glycero-3-phosphocholine (palmitoyllyso-GPC) transcylation as a function of Ca2+ concentration was found to be correlated with appearance of endogenous lysoderivatives. We also demonstrated that endogenously produced acyllyso-sn-glycero-3-phosphoethanolamine (acyllyso-GPE) induced CoA-dependent arachidonate transfer from diacyl-GPC. These results further argue for a two-step arachidonate release from diacyl-GPC when platelets are stimulated with thrombin.