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目的检测小鼠血管平滑肌细胞(VSMCs)中Sirtuin3(Sirt3)的表达,探讨Sirt3在血管紧张素Ⅱ(AngⅡ)诱导小鼠VSMCs增殖中的作用。方法用RT-PCR、Western blot法检测细胞中是否有Sirt3基因和蛋白表达;用不同浓度AngⅡ(10-7mol/L、10-6mol/L、10-5mol/L)分别刺激细胞,用RT-PCR、Western blot法检测AngⅡ对Sirt3表达的影响;用Sirt3 siRNA转染干扰Sirt3mRNA水平后5-乙炔基-2’脱氧尿嘧啶核苷(Edu)试剂盒检测细胞增殖水平。结果 Western blot法、RT-PCR结果均显示正常C57小鼠VSMCs中有一定量Sirt3蛋白和mRNA表达;10-7mol/L、10-6mol/L、10-5mol/L AngⅡ刺激细胞后Sirt3 mR-NA水平及蛋白表达量均明显升高(P<0.01),其中10-6mol/L组比10-7mol/L组和10-5mol/L组升高更明显,差异有统计学意义(P<0.05);Sirt3敲减组细胞增殖较对照组明显增加(P<0.01),Sirt3沉默+AngⅡ(10-6mol/L)组较AngⅡ组细胞增殖明显增加(P<0.01)。结论小鼠VSMCs中有一定量的Sirt3表达,AngⅡ刺激可使Sirt3 mRNA水平和蛋白表达量明显升高;Sirt3可抑制AngⅡ诱导的VSMCs增殖。
Objective To detect the expression of Sirtuin3 (Sirt3) in mouse vascular smooth muscle cells (VSMCs) and to explore the role of Sirt3 in the proliferation of mouse VSMCs induced by Ang Ⅱ. Methods Sirt3 gene and protein expression were detected by RT-PCR and Western blot. Cells were stimulated with different concentrations of AngⅡ (10-7mol / L, 10-6mol / L, 10-5mol / L) PCR and Western blot were used to detect the effect of AngⅡ on Sirt3 expression. Sirt3 siRNA transfected Sirt3 siRNA transfected Sirt3 siRNA transfected Sirt3 siRNA into Sirt3 mRNA. Results Western blot and RT-PCR showed that a certain amount of Sirt3 protein and mRNA were detected in VSMCs of normal C57 mice. Sirt3 mR-NA was stimulated by 10-7 mol / L, 10-6 mol / L and 10-5 mol / (P <0.01). The levels in 10-6mol / L group were significantly higher than those in 10-7mol / L group and 10-5mol / L group (P <0.05) (P <0.01). Sirt3 silencing + AngⅡ (10-6mol / L) group significantly increased cell proliferation compared with AngⅡgroup (P <0.01). Conclusion Sirt3 expression in mouse VSMCs is a certain amount. Sirt3 mRNA and protein expression are significantly increased by AngⅡ stimulation. Sirt3 can inhibit the proliferation of VSMCs induced by AngⅡ.