P85磷酯肌醇-3激酶-蛋白激酶C-ζ复合物对血管紧张素Ⅱ激活平滑肌细胞p70核蛋白体S6激酶的调节作用  

Phosphatidylinositol 3-kinase/Protein Kinase C-ζ Complex Regulates Angiotensin Ⅱ Activation of p70 Ribosomal S6 Kinase in Vascular Smooth Muscle Cells

作　　者：廖端芳[1] BerkBC 关永源[3] 

机构地区：[1]衡阳医学院心肺药理研究室,衡阳421001 [2]Center for Cardiovascular Research University of Rochester Medical Center,Rochester [3]中山医科大学药理学教研室,广州510089

出　　处：《中国动脉硬化杂志》2000年第3期193-198,共6页Chinese Journal of Arteriosclerosis

基　　金：国家自然科学基金!资助课题 (39970 847)

摘　　要：作者以前的研究发现蛋白激酶C ζ介导血管紧张素Ⅱ经Ras MEK途径激活血管平滑肌细胞丝裂素活化的蛋白激酶MAPK或细胞外信号调节激酶ERK1/ 2。本文研究了P85磷酯肌醇 3激酶 蛋白激酶C ζ复合物核蛋白体激酶p70S6激酶活性的调节作用及其信号传递途径。WesternBlot分析显示正常培养的血管平滑肌细胞表达p70S6激酶、p85磷酯肌醇 3激酶和蛋白激酶C ζ。 10 0nmol血管紧张素Ⅱ和 10 μg/L血小板源生长因子刺激并不影响p70S6激酶、p85磷酯肌醇 3激酶和蛋白激酶C ζ表达。p70S6激酶磷酸转移酶活性分析发现血管紧张素Ⅱ与血管平滑肌细胞作用 5min即开始激活p70S6激酶 ,2 0min达高峰 ,并呈剂量依赖性。磷酯肌醇 3激酶抑制剂wort mannin (10nmol)、蛋白激酶C ζ抑制剂PseudoZ (5 0 μmol)和p70S6激酶抑制剂rapamycin (10 0 μg/L)均可明显阻断血管紧张素Ⅱ诱导的p70S6激酶激活。有趣的是 ,我们观察到p85磷酯肌醇 3激酶和蛋白激酶C ζ受血管紧张素Ⅱ和血小板源生长因子刺激后均向Ras转位并与之结合 ,抗Ras抗体可将p85磷酯肌醇 3激酶或蛋白激酶C ζ混合沉淀 ,抗p85磷酯肌醇 3激酶抗体亦可使蛋白激酶C ζ沉淀下来。提示Ras、p85磷酯肌醇 3激酶和蛋白激酶C ζ三种蛋白结合在一起形成一个功能复合体 。Aim To investigate the effects of phosphatidylinositol 3-kinase (PI3K)/protein kinase C-ζ (PKC-ζ)complex on regulating angiotensin Ⅱ(AngⅡ) activation of p70 ribosomal S6 kinase (p70S6K) in vascular smooth muscle cells(VSMC). Methods VSMC isolated from 200 g to 250 g male Sprague-Dawley rats were cultured to 70% to 80% confluence and were growth-arrested by incubation in 0.1% calf serum/DMEM for 48 h before use. Both 3 H-TdR incorporation and cell counting were used to estimate the proliferation of VSMC. Western blot, immunoprecipitation and immunoblot analyses were performed to valuate the AngⅡ-stimulated kinase expression and Ras-PI3K-PKC-ζ association of VSMC. Phosphotransferase assay by using S6 peptide as substrate was employed to measure the p70 S6K activity. Results VSMC expressed p85PI3K, PKC-ζ and p70S6K; AngⅡ and PDGF stimulation did not affect the p85PI3K, PKC-ζ and p70S6K expression. 2) 100 nmol AngⅡ

关 键 词：磷酯肌醇-3激酶 血管紧张素Ⅱ 蛋白激酶C 

分 类 号：Q55[生物学—生物化学]