骨髓间充质干细胞向急性肾损伤小鼠的肾脏归巢现象及功能探讨  

Mesenchymal stem cells homing to the injured kidneys and their function assay in mice

作　　者：刘楠梅[1] 田军[1] 程劲[1] 胡大勇[1] 张金元[1] 

机构地区：[1]解放军第四五五医院肾内科南京军区肾脏病专科中心,上海200052

出　　处：《肾脏病与透析肾移植杂志》2009年第5期430-437,共8页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：南京军区医药卫生基金(07M023);南京军区122工程学科带头人培养基金

摘　　要：目的:观察在缺血再灌注(ischemia/reperfusion,I/R)模拟急性肾损伤的小鼠模型中,骨髓间充质干细胞(mesenchymal stem cells,MSCs)能否向损伤的肾小管迁移并促进其修复。方法:Percoll密度梯度离心结合贴壁培养法有效分离纯化出C57BL/6小鼠的骨髓间充质干细胞(mMSCs),5-溴脱氧尿嘧啶核苷(BrdU)标记并采用免疫组化法鉴定。雄性C57BL/6小鼠45只,分为正常对照组(15只)、I/R组(15只、夹闭双侧肾蒂30 min开放)、I/R+BrdU-mMSCs组(15只、夹闭双侧肾蒂30 min开放的同时尾静脉注射经BrdU标记的mMSCs)。于建模后1d、2d、3d、7d、14d分别处死部分小鼠(每次每组均处死3只),留取动脉血及肾组织,检测血肌酐(SCr)及尿素氮(BUN)水平,观察肾组织病理变化,荧光组织化学及免疫组织化学观察MSCs在受体小鼠肾组织的分布并对其进行鉴定。结果:经免疫组化证实BrdU标记mMSCs的阳性率可达(98.71±0.32)%。I/R组、I/R+BrdU-mMSCs组小鼠的BUN及SCr均显著高于正常对照组,但I/R+BrdU-mMSCs组小鼠的BUN及SCr水平却较同一时间点的I/R组为低,肾小管损伤程度评分也有着显著降低。免疫组化检测显示I/R+BrdU-mMSCs组小鼠肾脏中可检测到BrdU+细胞的分布,以术后第3天最多[(19.36±6.94)%]。经荧光组织化学及激光共聚焦显微镜观察证实该BrdU+细胞定位于小鼠的肾小管上皮细胞(renal tubular epithelial cells,RTECs)处,且表达RTECs特异性的标志物角蛋白18(cytokertain-18,CK18)。结论:小鼠发生I/R诱导的急性肾损伤后可诱导MSCs向损伤的肾小管上皮迁移并转化,参与RTECs的更新,促进肾损伤的修复。Objective：To observe whether mesenchymal stem cells （MSCs） can transfer to the injured renal tubules and contribute to its recovery in mice with acute renal ischemia-reperfusion （I/R） injury model. Methodology： C57BL/6 mice＇s mMSCs had been successfully isolated by percoll density gradient centrifugation and adherence cultivation, then marked with BrdU, the marking ratio was identified by immunohistochemistry. 45 healthy male C57BL/6 mice were divided into control group （ n = 15 ） , I/R group （ n = 15, clamping bilateral renal pedicles and then reopening after 30 minutes）, I/R ＋ BrdU-mMSCs group （n = 15, clamping bilateral renal pedicles and then reopening after 30 minutes, meanwhile, BrdU marked mMSCs were injected through caudal vein into the body of model mice）. 1, 2, 3, 7, and 14 days after, 3 mice in each group were killed, their blood and kidneys were collected. The blood was used to detect serum creatinine （SCr） and urea nitrogen（ BUN）, and mice＇s kidneys were dyed with HE to observe their pathological changes. The mMSCs＇ desposition in mice ＇s kidneys was observed by fluorescence histoehemistry and immunohistochemistry. Results：The positive BrdU marking ratio was （98.71 ±0. 32） %. The level of BUN and SCr in I/R group and I/R ＋ BrdU- mMSCs group was much higher than that in control group. While comparing with I/R group, the level of BUN and SCr in I/R ＋ BrdU-mMSCs was much lower, acute tubular necrosis score was also significantly lower. The immunohistochemistry results showed that BrdU marked mMSCs deposited in I/R ＋ BrdU-mMSCs group＇s kidney, the highest level was on the third day（ 19. 36±6. 94）%. Furthermore, by fluorescence histochemistry and laser confocal microscopy, BrdU marked mMSCs were confirmed to deposit in renal tubule epithelium and it could express marker cytokertain-18 （CK18） specified in renal tubule epithelial cells （RTECs）. Conclusion：Acute renal injury by I/R can induce MSCs transferring to injured kidney and makin

关 键 词：骨髓间充质干细胞 缺血再灌注 肾小管上皮细胞 

分 类 号：R692[医药卫生—泌尿科学]