ROCKⅠ/Ⅱ在转化生长因子β1诱导的主动脉平滑肌细胞表型转化中的作用  

Effects of ROCKⅠ/Ⅱ on phenotype switch in aortic vascular smooth muscle cells induced by TGF-β1

作　　者：汪海波[1] 高旭辉[1] 朱健[1] 郗二平[1] 张瑜[1] 王正 刘子豪[1] 谢彪 朱水波[1] 

机构地区：[1]中国人民解放军广州军区武汉总医院心胸外科,湖北武汉430070

出　　处：《中国普通外科杂志》2017年第12期1568-1574,共7页China Journal of General Surgery

基　　金：湖北省武汉市科技局应用基础研究计划基金资助项目(215060101010053)

摘　　要：目的:探讨ROCKⅠ/Ⅱ在转化生长因子β1(TGF-β1)诱导的人主动脉平滑肌细胞(HA-VSMC)表型转化中的作用。方法:将HA-VSMC分别转染ROCKI和ROCKII的si RNA后荧光显微镜观察转染情况,并用Western blot方法检测不同处理的HA-VSMC(ROCKI si RNA转染、ROCKII si RNA转染、+TGF-β1、ROCKI si RNA转染+TGF-β1、ROCKII si RNA转染+TGF-β1)中ROCKI和ROCKII蛋白的表达;分别用Western blot和RT-PCR方法检测不同处理的HA-VSMC(+TGF-β1、ROCKI si RNA转染+TGF-β1、ROCKII si RNA转染+TGF-β1、ROCK非特异性抑制剂Y-27632预处理+TGF-β1)中细胞收缩表型标志物α-平滑肌肌动蛋白(α-SMA)、平滑肌22α(SM22α)与合成表型标志物骨桥蛋白(OPN)的蛋白与m RNA表达,均以无处理的HA-VSMC为空白对照。结果:免疫荧光观察与Western blot检测表明两种si RNA均成功转染;TGF-β1处理后,HA-VSMC中ROCKI蛋白表达明显升高(P<0.05),但ROCKII蛋白表达无明显变化(P>0.05),ROCKI si RNA转染后TGF-β1上调ROCKI的作用被明显抑制(P<0.05)。与空白对照组HA-VSMC比较,TGF-β1处理后的HA-VSMC中α-SMA、SM22α的蛋白和m RNA表达明显降低,而OPN蛋白与m RNA表达明显升高(均P<0.05),ROCKI si RNA转染或Y-27632预处理后,TGF-β1的上述作用均明显减弱(均P<0.05),ROCKII si RNA转染对TGF-β1的上述作用均无明显影响(均P>0.05)。结论:TGF-β1可诱导HA-VSMC由收缩表型向合成型表型转化,ROCKI表达的升高可能在这一转化中起主要作用。Objective： To investigate the actions of ROCKI/II in phenotypic transformation of human aortic vascular smooth muscle cells （HA-VSMCs） induced by transforming growth factor β1 （TGF-β1）. Methods： HA-VSMCs were respectively transfected with ROCKI and ROCKII, and the transfection results were observed by fluorescence microscope. The ROCKI and ROCKII protein expressions in HA-VSMCs with different treatments （ROCKI siRNA transfection, ROCKII siRNA transfection, ＋TGF-β1, ROCKI siRNAtransfection＋TGF-β1, and ROCKII siRNAtransfection＋TGF-β1） were determined by Western blot analysis. The protein and mRNA expressions of the contractile phenotype maker α-smooth muscle actin （α-SMA） and smooth muscle 22α （SM22α） and synthetic phenotype marker osteopontin （OPN） in HA-VSMCs with different treatments （＋TGF-β1, ROCKI siRNA transfection＋TGF-β1, ROCKII siRNA transfection＋TGF-β1, and pretreatment of ROCK non-specificity Y-27632＋TGF-β1） were determined by Western blot analysis and RT-PCR method, respectively. Untreated HA-VSMCs were used as blank control. Results： Both siRNAs were successfully transfected as evidenced by fluorescence observation and Western blot analysis. In HA-VSMCs after TGF-β1 treatment, the ROCKI protein expression level was significantly up-regulated （P〈0.05）, but the ROCKII protein expression level did not significantly change （P〉0.05）, while the ROCKI increasing effect of TGF-β1 was significantly inhibited by ROCKI siRNA transfection （P〈0.05）. In HA-VSMCs after TGF-β1 treatment, the protein and mRNA expressions of α-SMA and SM22α were decreased and those of OPN were increased significantly （allP〈0.05）, and these effects were signi昀cantly suppressed by ROCKI siRNA transfection or Y-27632 pretreatment （allP〈0.05）, but were not affected by ROCKII siRNA transfection （allP〉0.05）. Conclusion： TGF-β1 can induce the transformation of HA-VSMCs from contractile phenotype to synthetic phenotype, which may be associa

关 键 词：肌 平滑 血管 动脉瘤 夹层 RHO相关激酶类 ROCKI/II 表型 

分 类 号：R654.3[医药卫生—外科学]