摘要
为了比较蛋白酶活化受体 1和 4(PAR1,PAR4)合成肽对血小板膜表面糖蛋白GPIbα表达及细胞骨架的影响 ,揭示蛋白酶活化受体在血小板信号传递中的作用 ,选择 2 5μmol/LPAR1与 2 50 μmol/LPAR4两类合成肽分别在不同时间段 (0 -60分钟 )活化血小板 ,应用流式细胞仪测定血小板膜糖蛋白GPIbα及P 选择蛋白的表达 ,并结合SDS PAGE与转移电泳技术比较血小板活化前后细胞骨架GPIbα、肌动蛋白、肌球蛋白的变化 ,同时对膜骨架成分进行GPIbα免疫沉淀分析。结果显示 ,PAR 1与PAR4两类合成肽均可促使血小板活化 ,其P 选择蛋白水平显著升高 ,GPIbα表达进行性减少又出现逐渐回升的可逆性变化 ,各时间段引起的GPIbα改变在两者之间都具有显著差异 (P <0 .0 5) ,其中PAR1首先发生作用 ,但PAR4维持时间更长。细胞骨架蛋白分析显示肌动蛋白与肌球蛋白协同GPIbα呈现先增加后减少的动态变化。免疫印迹也表明与GPIbα相连的肌动蛋白及肌球蛋白出现可逆性改变。结论 :PAR1与PAR4在血小板信号传递过程中发挥了重要作用 ,它们能各自独立地介导血小板活化 ,并导致GPIbα逆转 ,这与细胞骨架的积极参与相关。PAR1起效较快 。
This study was designed to compare the effects of protease-activated receptor 1 (PAR-1) and protease-activated receptor 4 (PAR -4) to the expression of platelet surface GPIbα and cytoskeleton reorganization, then to investigate the role of P ARs in platelet signal transmission. PAR1 (25 μmol/L) and PAR4 (250 μmol/L) we re used to stimulate platelet at different time points (0-60 minutes), and the p latelet surface GPIbα, actin and myosin and P-selectin were detected with flow cytometry, the alteration of GPIbα, actin and myosin in cytoskeleton was compared by Western blot, the membrane cytoskeleton followed GPIbα immunop recipitation was analyzed. The results showed that an increase of P-selectin an d reversible decrease of GPIbα expression were obtained after platelet activation by PAR1 o r PAR4, and a different kinetics of redistribution of GPIbα was found for the t wo peptides all over the time course (P<0 05). PAR1 acted more potently and rapidly than PAR4, but the effect of PAR4 persisted longer in the course of plat elet activation. Meanwhile, there was a transient change of actin, myosin and GP Ibα in cytoskeleton proteins. Similar redistribution was also found in GPIbα/m yosin and GPIbα/actin association. It is concluded that PAR1 and PAR4 possess a n important role in platelet signal transmission. Either of the receptors can me diate platelet activation and GPIbα redistribution, which is correlated with cy toskeleton reorganization. PAR1 acts more rapidly, and effect of PAR4 persists l onger.
出处
《中国实验血液学杂志》
CAS
CSCD
2003年第5期495-498,共4页
Journal of Experimental Hematology
