BMSCs对猪胰岛缺氧再给氧损伤的保护作用  

PROTECTIVE EFFECT OF BONE MARROW MESENCHYMAL STEM CELLS ON ISLETS FROM HYPOXIA/REOXYGENATION-INDUCED INJURY

作　　者：赵久茗[1] 何斯荣[1] 王成世[1] 刘敬平[1] 李胜富[1] 陈又南[1] 陆燕蓉[1] 程惊秋[1] 

机构地区：[1]四川大学华西医院再生医学研究中心,成都610041

出　　处：《中国修复重建外科杂志》2013年第8期910-915,共6页Chinese Journal of Reparative and Reconstructive Surgery

摘　　要：目的研究恒河猴BMSCs在缺氧再给氧环境下对新生猪胰岛活性和功能的保护作用。方法取健康成年恒河猴5只,体重6～10 kg,取骨髓采用骨髓单核细胞贴壁培养法筛选获得BMSCs;取3～5日龄新生长白猪5只,取胰腺以Ⅴ型胶原酶消化制备新生猪胰岛。将实验分为胰岛正常组(A组)、胰岛加BMSCs正常组(B组)、胰岛缺氧再给氧组(C组)、胰岛加BMSCs缺氧再给氧组(D组),观察比较常规培养和缺氧(1%O2)12 h再给氧24 h或48 h条件下,胰岛的功能活性变化:二乙酸荧光素/碘化丙啶(propidium iodide,PI)染色计算胰岛成活率;细胞计数试剂盒8法检测新生猪胰岛的代谢活性;膜联蛋白V-FITC/PI标记后流式细胞仪检测胰岛凋亡率;葡萄糖刺激法分析胰岛功能糖刺激指数(stimulation index,SI)。结果 C组较A、B组胰岛团更为分散,死亡细胞增多;D组较C组胰岛团完整,成活率更高。A、B、C、D组胰岛成活率分别为90.2%±9.1%、88.3%±5.9%、52.3%±12.1%、71.4%±11.5%,C、D组显著低于A、B组(P<0.05),D组显著高于C组(P<0.05)。D组BMSCs以1∶10和1∶20比例与胰岛缺氧再给氧共培养后,代谢活性显著高于C组(P<0.05)。A、B、C、D组胰岛凋亡率分别为27.1%±3.2%、24.0%±1.0%、64.3%±1.8%、46.2%±1.4%,C、D组显著高于A、B组(P<0.05),但D组显著低于C组(P<0.05)。各时间点A、B组SI比较差异均无统计学意义(P>0.05),C组显著低于A、B组(P<0.05),在24 h和72 h时D组显著高于C组(P<0.05)。结论 BMSCs对缺氧再给氧诱导的新生猪胰岛活性和功能损伤具有保护作用。Objective To study the protective effects of bone marrow mesenchymal stem cells（BMSCs） of rhesus monkeys on porcine islets from hypoxia/reoxygenation（H/R）-induced injury. Methods BMSCs were isolated and cultured from the marrow of 5 adult rhesus monkeys（weighing,6-10 kg） by adherent monocytes.Islets were isolated and purified from the pancreas of 5 neonatal porcine（3-5 days old） by collagenase V digestion method,and were cultured with or without BMSCs,and exposed to hypoxia（1%O2） for 12 hours and reoxygenation for 24 or 48 hours,respectively.The experiment was divided into 4 groups： normal islet group（group A）,normal islet ＋ BMSCs group（Group B）,H/R islet group（group C）,and H/R islet ＋ BMSCs group（group D）.The survival rate of islets was calculated by fluorescein diacetate/propidium iodide（PI） staining.The viability of the islet cells was detected by cell counting kit 8.Apoptotic rate of islet cells was tested using Annexin V-FITC/PI labeling and flow cytometry.The stimulation index（SI） of islet function was analyzed by glucose-stimulated insulin secretion assay. Results The islet cell cluster of group C was more dispersed than that of groups A and B,and group C had more death cells;and the islet cell cluster of group D was more complete and the survival rate was higher than those of group C.The survival rate of islet was 90.2% ± 9.1%,88.3% ± 5.9%,52.3% ± 12.1%,and 71.4% ± 11.5% in groups A,B,C,and D respectively,it was significantly lower in groups C and D than in groups A and B（P 0.05）,but it was significantly higher in group D than in group C（P 0.05）.After coculture of BMSCs and islet at the ratio of 1∶10 and 1∶20 in group D,the viability of islet cells was significantly higher than that in group C（P 0.05）.The apoptotic rate was 27.1% ± 3.2%,24.0% ± 1.0%,64.3% ± 1.8%,and 46.2% ± 1.4% in groups A,B,C,and D respectively,it was significantly higher in groups C and D than that in groups A and B（P 0.05）,but it was significantly lower in g

关 键 词：BMSCS 胰岛 缺氧再给氧 恒河猴 猪 

分 类 号：R587.1[医药卫生—内分泌]