氧化应激预适应减轻骨髓间充质干细胞氧化应激损伤

Oxidative stress preconditioning alleviates oxidative stress-induced damage of bone marrow mesenchymal stem cells

目的探讨氧化应激预适应对骨髓间充质干细胞(BMSC)氧化应激损伤的影响。方法通过密度梯度离心联合贴壁法分离培养BMSC,分为对照组(只加培养液)、预处理组(用50μmol/L H_(2)O_(2)预处理8 h,再用1000μmol/L H_(2)O_(2)持续处理24 h)、氧化损伤组(直接用1000μmol/L H_(2)O_(2)持续处理24 h)。采用2′,7′-二氯二氢荧光素二乙酸酯(DCFH-DA)染色检测细胞活性氧(ROS)水平;JC-1染色法检测线粒体膜电位;原位末端脱氧核糖核酸转移酶标记技术(TUNEL)检测DNA损伤;硫代巴比妥酸(TBA)法检测丙二醛(MDA)含量;水溶性四唑盐1(WST-1)法检测超氧化物歧化酶(SOD)活力;CCK-8法检测细胞活力及流式细胞术检测细胞凋亡。结果与氧化损伤组相比,预处理组ROS水平降低,线粒体膜电位升高,DNA损伤减轻,MDA含量明显减少,SOD活力及细胞活力提高,细胞凋亡率显著降低。结论氧化应激预适应可增强BMSC抗氧化应激能力,并促进其在氧化应激条件下的存活。

Objective To investigate the effect of oxidative stress preconditioning on the oxidative stress-induced damage of bone marrow mesenchymal stem cells(BMSCs).Methods BMSCs were isolated and cultured by density gradient centrifugation combined with adherence method.The cells were divided into three groups:control group(control medium),oxidative damage group(treated with 1000μmol/L H_(2)O_(2)for 24 hours),and preconditioning group(preincubated with 50μmol/L H_(2)O_(2)for 8 hours before treatment with 1000μmol/L H_(2)O_(2)for 24 hours).DCFH-DA staining was used to analyze the level of reactive oxygen species(ROS).Mitochondrial membrane potential was measured by JC-1 staining.DNA damage was detected by TUNEL.Malondialdehyde(MDA)content was detected by thiobarbituric acid(TBA)method,and superoxide dismutase(SOD)activity was detected by water soluble tetrazolium-1(WST-1)assay.CCK-8 assay was used to detect cell viability and flow cytometry to detect cell apoptosis.Results Compared with the oxidative damage group,the preconditioning group had reduced ROS level,reduced DNA damage,higher mitochondrial membrane potential,significantly decreased MDA content,increased SOD activity,increased cell viability,and significantly decreased apoptosis.Conclusion Oxidative stress preconditioning can enhance the anti-oxidative stress ability of BMSCs and promote its survival under oxidative stress.