槲皮素对过氧化氢损伤小鼠成纤维细胞的保护作用

目的探讨槲皮素对过氧化氢(H_2O_2)损伤小鼠成纤维细胞(NIH-3T3)的保护作用。方法体外培养NIH-3T3细胞,取对数生长期的NIH-3T3细胞分成四个实验组。对照组(C)仅加含有10%胎牛血清的DMEM培养基。实验1(Q_1)组先用0．5mmol/LH_2O_2作用细胞30min,然后再用含有50μmol/L的槲皮素培养基作用24h。实验2(Q_2)组先用含有501μmol/L的槲皮素培养基作用24h,再换含有0．5mmol/L H_2O_2作用细胞30min。H_2O_2实验(H_2)组先用含有0．5mmol/L H_2O_2作用细胞30min,再换仅含有10%胎牛血清的DMEM培养基作用24h。取培养细胞提取DNA和制备1×10~7/ml细胞悬液的滴片,应用TUNEL和Ladder电泳检测细胞凋亡率；应用Weaster blod和细胞免疫组化技术检测Bcl-2、Bax、Caspase-3、HSP-70、NF- kB蛋白质的表达。结果由TUNEL和Ladder实验得出,Q_1组的细胞凋亡率明显低于H_2组和Q_2组。由Weaster blod和细胞免疫组化技术检测分析得出,Q_1组与Q_2、H_2组相比,Bcl-2的表达明显增高,Bax、Caspase-3、NF-kB、HSP-70的表达减低。结论槲皮素可能主要是通过抑制NF-kB的表达,提高HSP-70的表达,对H_2O_2诱导的NIH-3T3细胞损伤产生保护作用。

Antioxidative effects of quercetin on damage induced by H2O2 in NIH-3T3 cells of mice

Objective To explore antioxidative effects of quercetin on damage induced by H_2O_2 in NIH-3T3 cells.Methods The NIH-3T3 cells in logarithmic growth phase incubated in DMEM were divided into 4 groups as follows:(1)Q pre-protection group(Q_1);prior to incubation with 0.5mmol/LH_2O_2 for 30 min,the NIH-3T3 cells were incubated with 50μmol/L quercetin for 24h;(2)Q post-protection group(Q_2):after incubation with 0.5mmol/L H_2O_2 for 30 min,the NIH-3T3 cells were incubated with 50umol/L quercetin for 24h;(3)H_2O_2 damage group(H_2):the NIH-3T3 cells were incubated with 0.5mmol/L H_2O_2 for 30 rain followed by subsequent incubation with 10% FBS for 24h;(4)The control group(C):the NIH-3T3 cells were incubated with DMEM medium supplemented with 10% FBS.The apoptosis was detected by DNA ladder and TUNEL.The expressions of Bcl-2,Bax,Caspase-3,HSP-70 and NF-kB were demonstrated with immunocytochemistry and Western blotting methods.The NIH-3T3 cell survival rate was detected by MTT assay.Results The apoptosis in the Q_1 group was decreased than that in the Q_2 and H_2 groups.The Q_1 group compared with Q_2 and H_2 groups,the expressions of Bcl-2 was increased;Bax,Caspase-3,HSP-70 and NF-kB were attenuated;and the NIH-3T3 cell survival rate was promoted.Conclusion The antioxidative effect of quercetin on damage in the NIH-3T3 cells may act through down-regulated expressions of NF-kB and up-regulated expression of HSP-70.