肝硬化大鼠血管平滑肌细胞Gq蛋白信号转导功能的研究

The impairment of Gq protein coupled receptor in aortic vascular smooth muscle cells of cirrhotic rats

目的 为观察肝硬化大鼠动脉血管平滑肌细胞对血管紧张素Ⅱ (AT Ⅱ )是否存在受体后Gq蛋白信号转导通路的异常。方法 采用总胆管结扎制备大鼠肝硬化模型 ,分离主动脉血管平滑肌细胞后进行细胞培养 ,并予免疫组化鉴定。分别采用细胞感应微生理探测系统、三磷酸肌醇(IP3)分析法、Western印迹法检测血管平滑肌细胞AT Ⅱ受体介导的细胞外液酸化速率 (ECAR)、IP3生成、P42 / 44丝裂原激活的蛋白激酶 (MAPK)磷酸化功能。结果 大鼠胆管结扎后 4 5周肝硬化形成。AT Ⅱ可剂量依赖性增强假手术组ECAR及IP3生成 ,但肝硬化组的ECAR和IP3形成明显降低 ,肝硬化组中AT Ⅱ刺激P42 / 44MAPK的磷酸化亦显著减少 ,但 2组的表达总量不变。结论 肝硬化时 ,动脉血管平滑肌细胞对AT Ⅱ受体介导的Gq蛋白信号转导功能发生障碍 。

Objective To test angiotensin Ⅱ (AT Ⅱ) receptor mediated signal transduction in aortic vascular smooth muscle cells (VSMCs) of cirrhotic rats. Methods Cirrhosis was induced by bile duct ligation. VSMCs were characterized by immunohistochemistry. Cytosensor microphysiometry,inositol 1,4,5 triphosphates (IP 3) assay, and Western blotting analysis were used to evaluate AT Ⅱ stimulated cellular responsiveness of VSMCs, such as the extracelluar acidification rate (ECAR),IP 3 formation and phosphorylation of P42/44 mitogen activated protein kinase (MAPK) in sham operated or cirrhotic rats. Results The secondary biliary cirrhosis was developed 4 5 weeks after operation. Our results showed that ECAR and IP3 formation stimulated by AT Ⅱ were significantly reduced in the aortic VSMCs from the cirrhotic rats. Furthermore, the maximal p42/44 MAPK phosphorylation stimulated by AT Ⅱwas also considerably suppressed in the cirrhotic VSMCs in contrast to control cells. Conclusion Our results demonstrated that Gq coupled receptor signaling pathway stimulated by AT Ⅱ was severely impaired in aortic VSMCs of cirrhotic rats, which may lead to arterial hyporesponsiveness to vasoconstrictors in cirrhosis.