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在体外培养的血管平滑肌细胞上 ,采用蛋白印迹杂交免疫沉淀的方法 ,研究尾加压素Ⅱ (UⅡ )与粘着斑激酶 (FAK)介导的信号传导之间的关系。结果表明 ,UⅡ在 10 -8、10 -7和 10 -6mol/L的浓度时 ,可分别使FAK活力增加 2 2、3 0 4和 0 73倍 ;加入UⅡ (10 -7mol/L) 5min后 ,FAK活力增加 95 % ,但与对照组相比无显著性差异(P >0 0 5 ) ,刺激 10min后 ,FAK活力升高 3 7倍 ,和对照组相比有显著性差异 (P <0 0 5 )。至 30min时活力显著升高 ,较对照组增加 4 8倍 (P <0 0 1)。UⅡ刺激平滑肌细胞 30min内 ,FAK的蛋白含量无明显变化 ;在UⅡ作用 2h后 ,FAK的含量增加 36 % ,至 4h达高峰 ,与对照组相比增加 5 3% ,6h后降低 ;细胞松弛素B不能抑制UⅡ对FAK的激活 ;丝裂素活化蛋白激酶的抑制剂PD980 5 9(5 0 μmol/L) ,钙调素依赖性蛋白激酶的抑制剂W7(5 0 μmol/L) ,对FAK的激活无影响 (P >0 0 5 ) ;蛋白激酶C的阻断剂H7(5 0 μmol/L) ,可与UⅡ发生协同作用 ,使FAK活力较单纯UⅡ组增加 1 98倍。因此 ,UⅡ与FAK介导的信号转导途径之间存在着密切的关系 ,且这种关系不依赖于细胞骨架的完整性 ,与蛋白激酶C介导的信号转导途径有密切的关系。
The relationship between urotensin Ⅱ (UⅡ) and focal adhesion kinase (FAK) -mediated signaling was studied by using Western blotting and immunoprecipitation on cultured vascular smooth muscle cells. The results showed that FA could increase the activity of FAK by 2 2,3 0 4 and 0 73 times when treated with 10 -8, 10 -7 and 10 -6 mol / L, respectively. When UⅡ (10 -7 mol / L) , FAK activity increased 95%, but there was no significant difference compared with the control group (P> 0.05). FAK activity increased 37% after stimulated for 10min, there was significant difference compared with the control group (P <0 0 5). The vitality increased significantly at 30 minutes, which was 48 times more than that of the control group (P <0.01). The protein content of FAK did not change significantly after UⅡ stimulated smooth muscle cells for 30min. After treated with UⅡ for 2h, the content of FAK increased 36%, peaked at 4h, increased by 53% compared with the control group, and decreased after 6h. The cytochalasin B could not inhibit the activation of FAK by UII; PD98059 (50μmol / L), inhibitor of calmodulin-dependent protein kinase (W050μmol / L) (P> 0.05). H7 (50 μmol / L), a blocker of protein kinase C, had a synergistic effect with UⅡ, which increased the FAK activity by 988 times compared with UⅡ alone. Therefore, there is a close relationship between UII and FAK-mediated signal transduction pathways, and this relationship does not depend on the integrity of the cytoskeleton, and is closely related to the protein kinase C-mediated signal transduction pathway.