Studies on the function and the regulation of expression of platelet membrane protein GPIV (CD36)

我们利用CD36缺陷血小板研究了血小板膜蛋白CD36对血小板信号转导的影响。据报道，CD36的作用之一是不饱和脂肪酸的转运体。与正常血小板相比，CD36缺陷血小板的特征性反应很少。花生四烯酸（AA）是一种不饱和脂肪酸，通过其代谢物TxA2在血小板聚集中起关键作用。我们观察到，在Ca2+储存减少的条件下，CD36缺陷血小板在AA代谢物发挥更重要作用的反应中表现出明显的差异。我们发现AA抑制其代谢物TxA2诱导的血小板聚集。正常血小板AA IC50为26.5±12.2 μM， C36缺陷血小板AA IC50为105±44 μM。这些结果证实了CD36作为脂肪酸转运蛋白的作用。这是关于CD36缺陷血小板功能受损的首次报道。AA和其他不饱和脂肪酸独立于膜脂和胞质Ca2+激活胞浆中Ca2+独立的PKC， PKC同型的生理意义尚不清楚。因此，下一步研究了AA的抑制作用机制。证实AA通过Ca2+非依赖性PKC的丝氨酸磷酸化抑制PLCβ活化。因此，与聚集密切相关的Ca2+依赖性PKC不被激活。因此，PKC同型相互联系，导致血小板的生理适当反应。更重要的一点是，AA本身通过PKC同型调节其后代诱导的反应。

We have studied the effects of platelet membrane protein CD36 on the platelet signal transduction using CD36 defective platelets. One of the reported roles of CD36 is the transporter of unsaturated fatty acids. Compared to normal ones little characteristic responses of CD36 defective platelets have been reported. Arachidonic acid (AA), one of the unsaturated fatty acid, has pivotal role in platelet aggregation through its metabolite TxA2. We observed that under the decreasing store Ca2+ conditions CD36 defective platelets showed clear differences in the reactions where AA metabolites have more essential roles. We found that AA inhibits platelet aggregation induced by its metabolites TxA2. The IC50 of AA for normal platelets was 26.5±12.2 μM,and that for C36 defective platelets was 105±44 μM respectively. These results corroborate the role of CD36 as fatty acid transporter. This is the first report on the impaired functions of CD36 defective platelets. AA and other unsaturated fatty acids activate Ca2+ independent PKC in the cytosol independently of membrane lipids and cytosolic Ca2+ Physiological significances of the PKC isotypes have not been clearly understood. Hence the mechanisms of the inhibiting effects of AA were next studied. It was confirmed that AA inhibits PLCβ activation through serine phosphorylation by Ca2+ independent PKC.Consequently the Ca2+ dependent PKC closely connected to aggregation are not activated. Thus PKC isotypes are linked to each other leading to the physiologically proper responses of platelets. More important point is that AA itself regulates reactions induced by its descendants through the PKC isotypes.