SIRT3在高糖引起的细胞衰老过程中的表达变化及意义

Changes and significance of SIRT3 expression in cellular senescence induced by high glucose

目的探讨沉默信息调节因子3(SIRT3)在高糖引起的人二倍体成纤维细胞(WI-38)衰老过程中的表达变化和意义。方法通过低糖(3.34mmol/L)、正常糖(5.56mmol/L)、高糖(25mmol/L)三种不同条件培养WI-38细胞,使之由20群体倍增水平(PDs)增至32PDs。采用Western blotting检测p21、p27、Catalase、SIRT3和MnSOD的蛋白表达;免疫荧光共染方法检测各组WI-38细胞中SIRT3和MnSOD和衰老相关核异染色质灶(SAHF)的表达和定位;用荧光探针检测活性氧(ROS)的表达。结果 Western blot-ting结果显示,高糖组SIRT3、Catalase和MnSOD的表达与低糖组及正常糖组比较明显降低(P<0.05);高糖组的p21p、27的蛋白表达与正常糖组比较明显增加(P<0.05);与低糖组比较,高糖组的p21表达上调,而p27表达差异无统计学意义(P>0.05)。免疫荧光共染结果显示,高糖组中的SIRT3、Catalase和MnSOD表达较其他两组明显降低(P<0.05);SIRT3和MnSOD主要存在于胞质中。荧光探针显示,高糖组中的ROS水平与低糖组和正常糖组比较明显增加。结论高糖能加速WI-38细胞的衰老过程,SIRT3可能与高糖引起的细胞衰老有着密切联系。

Objective To investigate the role of the silent information regulator 3（SIRT3） in the decrepitude process of human diploid fibroblasts（WI-38） induced by high glucose.Methods The WI-38 cells [population doublings（PDs）,20-32] were cultured in media containing different concentrations of glucose as follows： low glucose（3.34mmol/L,LG）,normal glucose（5.56mmol/L,NG）,and high glucose（25mmol/L,HG）.The protein expression levels of p21,p27,catalase,MnSOD,and SIRT3 were evaluated through Western blot.The double-label immunofluorescence assay was used to detect the location and expression of SIRT3,MnSOD,and senescence-associated heterochromatin foci（SAHF） in the WI-38 cells.The ROS level was determined with fluorescent probe.Results The results from the Western blot showed that the protein expression of SIRT3,catalase,and MnSOD decreased significantly in the HG group compared with the LG and NG groups（P0.05）.The expression of p21 and p27 significantly increased in the HG group compared with the NG group（P0.05）.Compared with the LG group,the expression of p21 protein increased in the HG group,but the expression of p27 protein was not statistically significant（P0.05）.SIRT3 and MnSOD were highly expressed in the cytoplasm.The ROS levels in the HG group were elevated compared with those in the LG and NG groups.Conclusion SIRT3 may play an important role in cellular senescence induced by high glucose in human diploid fibroblasts.