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

我们研究了血小板膜蛋白CD36在CD36缺陷血小板信号转导中的作用。CD36的一个已报道的作用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较少。花生四烯酸(AA)是一种不饱和脂肪酸，通过其代谢产物TXA2在血小板聚集中发挥重要作用。我们观察到，在储存钙离子减少的情况下，CD36缺陷血小板在AA代谢物具有更重要作用的反应中表现出明显的差异。我们发现AA能抑制其代谢产物TXA2诱导的血小板聚集。AA对正常血小板的IC_(50)为26.5±12.2μM，对C36缺陷的IC_(50)为105±44μM。这些结果证实了CD36作为脂肪酸转运蛋白的作用。这是有关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激活胞浆中不依赖于膜脂的非钙依赖的PKC，胞浆中的PKC亚型的生理意义还不是很清楚。因此，接下来对AA的抑制作用机制进行了研究。已证实AA通过钙非依赖性蛋白激酶C的丝氨酸磷酸化作用抑制PLCKC的激活，因此与聚集密切相关的钙依赖性蛋白激酶C不被激活。因此，PKC亚型相互关联，导致了血小板生理上的正确反应。更重要的一点是，AA本身通过PKC亚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.