重组人MG53蛋白对人脐带间充质干细胞氧化损伤的保护作用  

MG53 protects umbilical cord mesenchymal stem cells from oxidative damage

作　　者：黄团结 宋及时 马珊珊 程康 邢衢 李鹏 刘腾飞 杨波[2] 关方霞 Huang Tuanjie;Song Jishi;Ma Shanshan;Cheng Kang;Xing Qu;Li Peng;Liu Tengfei;Yang Bo;Guan Fangxia(Department of Stem Cell Research,School of Life Science,Zhengzhou University,Zhengzhou 450001,China;Stem Cell Research Laboratory,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]郑州大学生命科学学院,450001 [2]郑州大学第一附属医院神经外科,450052

出　　处：《中华细胞与干细胞杂志（电子版）》2017年第1期35-44,共10页Chinese Journal of Cell and Stem Cell（Electronic Edition）

基　　金：国家自然科学基金(81601078;81471306);河南省国际人才合作项目(2016GH03;2016GH15)

摘　　要：目的建立人脐带间充质干细胞(h UC-MSCs)的H_2O_2氧化损伤模型,探讨重组人MG53蛋白(rh MG53)对h UC-MSCs氧化损伤的保护作用。方法使用组织块培养法从健康人脐带沃顿胶组织中分离培养h UC-MSCs,流式细胞仪检测细胞表型;CCK-8法检测h UC-MSCs的增殖和H_2O_2损伤程度。实验分为正常对照组(NC组)、rh MG53组、H_2O_2组和H_2O_2+rh MG53组。分别采用CCK-8法、Transwell小室、流式细胞术和β-半乳糖苷酶染色检测rh MG53蛋白对h UC-MSCs增殖、迁移、周期凋亡和衰老的影响。各组、各个时间点的吸光值采用析因设计的方差分析,组间比较采用单因素方差分析。结果 h UC-MSCs贴壁呈梭型生长,高表达间充质干细胞标志CD44、CD133、HLA-ABC,低表达CD34、CD45;H_2O_2对h UC-MSCs具有浓度和时间依赖性的损伤作用,确定200μmol/L的H_2O_2处理16 h为h UC-MSCs氧化损伤的最适条件。与NC组相比,H_2O_2组细胞增殖(0.994±0.011 vs 1.331±0.014)、迁移率(8.15﹪±2.47﹪vs 33.34﹪±3.62﹪)和S期细胞比例(29.67﹪±3.69﹪vs 34.33﹪±4.25﹪)显著降低,细胞衰老(44.07﹪±5.26﹪vs 5.7﹪±1.42﹪)和凋亡比例(52.63﹪±5.76﹪vs 4.65﹪±1.23﹪)显著增加,差异具有统计学意义(均P<0.05);rh MG53组细胞增殖(1.509±0.086 vs 1.331±0.014)和迁移率(52.13﹪±5.46﹪vs 33.34﹪±3.62﹪)显著提高,S期比例(35.27﹪±4.79﹪vs 34.33﹪±4.25﹪)变化差异无统计学意义(P>0.05),细胞衰老(3.92﹪±1.31﹪vs 5.7﹪±1.42﹪)和凋亡比例(3.96﹪±1.06﹪vs 4.65﹪±1.23﹪)显著降低,差异具有统计学意义(均P<0.05)。与H_2O_2组相比,H_2O_2+rhMG53组细胞增殖(1.252±0.056 vs 0.994±0.011)、迁移率(18.93﹪±3.12﹪vs8.15﹪±2.47﹪)和S期比例(34.84﹪±3.45﹪vs 29.67﹪±3.69﹪)显著提高,细胞衰老(17.89﹪±2.64﹪vs 44.07﹪±5.26﹪)和凋亡比例(4.65﹪±1.23﹪vs 17.63﹪±2.56﹪)显著降低,差异具有统计学意义(均P<0.05)。结论 rhMG53对H_2O_2造成的h UC-MSCs氧化损伤�Objective To establish a H2O2 oxidative damage model of human umbilical cord mesenchymal stem cells(hUC-MSCs),and to investigate the protective effects of recombinant human MG53 protein(rhMG53)on hUC-MSCs oxidative damage.Methods hUC-MSCs were isolated from human umbilical cord tissue collected from healthy donors,and the cell phenotype was determined by flow cytometry;the H2O2-treated P3 hUC-MSCs were evaluated by CCK-8 method.The cells were allocated into four groups:normal control group(NC group),rhMG53 group,H2O2 group and H2O2+rhMG53 group.The effect of rhMG53 on hUC-MSC proliferation,migration,senescence,cell cycle and apoptosis was respectively detected by CCK-8 assay,Transwell assay,flow cytometry andβ-galactosidase staining.Cell proliferation was analyzed by the variance analysis of factorial design,and the single factor variance analysis was used to compare the differences between groups.Results The P3 hUC-MSCs were adherent and spindle shaped,and expressed mesenchymal stem cell markers CD44,CD133,HLA-ABC,with low expression of CD34 and CD45.H2O2 inhibited hUC-MSC growth in a concentration and time-dependent manner.When hUC-MSCs were treated with 200 pmol/L H2O2 for 16 h,the proliferation(0.994±0.011 vs 1.331±0.014),migration(8.15%士2.47%vs 33.34%±3.62%)and percentage of cells in S phase(29.67%±3.69%vs 34.3%3士4.25%)decreased,while the proportion of senescent cells(44.07%±5.26%vs 5.7%±1.42%)and apoptosis(52.63%±5.76%vs 4.65%±1.23%)significantly increased in the H2O2 group compared with the NC group.The proliferation(1.509±0.086 vs 1.331±0.014)andmigration(52.13%±5.46%vs 33.34%±3.62%)of cells in the rhMG53 group increased significantly.The proportion of cells in S phase(35.27%±4.79%vs 34.33%±4.25%)was similar(P>0.05),and the proportion of senescent cells(3.92%±1.31%vs 5.7%±1.42%)and apoptosis(3.96%±1.06%vs 4.65±1.23%)was significantly decreased in the rhMG53 group(P<0.05).Compared with the H2O2 group,the cell proliferation(1.252±0.056 vs 0.994±0.011),migration ability(18.93%±3.12%

关 键 词：磷蛋白类 脐带 间质干细胞 创伤和损伤 保护 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]