五味子酚上调SIRT1改善H_2O_2对SH-SY5Y细胞增殖的抑制作用研究

Study on the Inhibitory Effect of Schisanhenol Up-regulated SIRT1 Improve H_2O_2 on the Proliferation of SH-SY5Y Cells

目的:探讨SIRT1/LC3自噬通路在五味子酚(Schisanhenol,Sal)抗H_2O_2诱导的SH-SY5Y细胞增殖活力下降中的作用。方法:加入不同浓度Sal(50、10μmol/L)处理SH-SY5Y细胞4 h后,再加入H_2O_2 100μmol/L处理12 h,采用MTT法检测细胞增殖能力,Western blot检测SIRT1蛋白和自噬蛋白标志物(LC3蛋白、Beclin-1蛋白)的表达水平。结果:H_2O_2组细胞增殖活力较对照组显著下降(P<0.001);Sal组不同浓度(50、10μmol/L)干预后的细胞增殖活力均高于H_2O_2组(P=0.001、0.017),且随着Sal浓度增加,细胞增殖活力增强。H_2O_2组SIRT1蛋白含量较对照组明显降低(P<0.001),Sal组不同浓度(50、10μmol/L)的SIRT1蛋白含量均明显高于H_2O_2组(P=0.001、0.014)。H_2O_2组Beclin-1和LC3-Ⅱ蛋白表达均高于对照组,比较差异均有统计学意义(P<0.001);Sal组(50μmol/L)的Beclin-1的蛋白表达低于H_2O_2组,且不同浓度(50、10μmol/L)LC3-Ⅱ蛋白表达均低于H_2O_2组,比较差异均有统计学意义(P<0.05)。结论:Sal可通过上调SIRT1蛋白表达,拮抗H_2O_2引起的过度自噬,从而起到拮抗H_2O_2对SH-SY5Y细胞增殖的抑制作用。

Objective:To investigate the role of SIRT1/LC3 autophagy pathway in the decrease of proliferation activity of SH-SY5Y cells induced by H2O2 in Schisanhenol(Sal).Method:SH-SY5Y cells were treated with different concentrations of Sal(50 and 10μmol/L)for 4 h and then added H2O2 100μmol/L for 12 h.The cell proliferation activity was detected by MTT,and the expressions of SIRT1 protein and autophagic protein markers(LC3 protein,Beclin-1 protein)were detected by Western blot.Result:The cell proliferation activity of H2O2 group was significantly lower than that of control group(P<0.001),the proliferative activity of Sal group at different concentrations(50,10μmol/L)was higher than that of H2O2 group(P=0.001,0.017),and the cell proliferation activity increased with the increase of Sal concentration.The content of SIRT1 protein in H2O2 group was significantly lower than that of control group(P<0.001),and the content of SIRT1 protein in Sal group at different concentrations(50,10μmol/L)was significantly higher than that of H2O2 group(P=0.001,0.014).The expressions of Beclin-1 and LC3-Ⅱproteins in group H2O2 were higher than those of control group,the differences were statistically significant(P<0.001),the protein expression of Beclin-1 in Sal group(50μmol/L)was lower than that of H2O2 group,and the expression of LC3-Ⅱprotein in different concentrations(50,10μmol/L)was lower than that in H2O2 group,the differences were statistically significant(P<0.05).Conclusion:Sal can attenuate the excessive autophagy induced by H2O2 by up regulation of the expression of SIRT1 protein,thus antagonizing the inhibitory effect of H2O2 on the proliferation of SH-SY5Y cells.