低氧诱导因子1α基因转染的人脂肪源性干细胞对肾小管上皮细胞损伤的影响  

Effect of human adipose-derived stem cells overexpressing HIF-1α on the cisplatin-indcued renal tubular cells

作　　者：王巍巍[1] 李慧凛[1] 张金元[1] 王葳[1] 

机构地区：[1]中国人民解放军第四五五医院肾脏科南京军区肾脏专科中心,上海200052

出　　处：《肾脏病与透析肾移植杂志》2011年第5期438-443,共6页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：上海市青年科技启明星计划(09QA1407500);上海市自然科学基金(10ZR1440000);南京军区医学科技创新项目重点课题(10Z008)

摘　　要：目的:将低氧诱导因子1α(HIF-1α)基因质粒高效感染人脂肪源性干细胞(hADSCs),并与顺铂诱导的人近端肾小管上皮细胞(HK-2)共孵育,观察hADSCs对HK-2细胞损伤的影响。方法:以慢病毒为载体,将HIF-1α转染hADSCs,与顺铂诱导的HK-2细胞共孵育。电镜观察HK-2细胞形态、结构的变化;原位末端标记(TUNEL)检测细胞凋亡指数;Westernblot检测HK-2细胞半胱氨酸天冬氨酸蛋白酶3(caspase-3)、B细胞淋巴瘤/白血病2基因(Bcl-2)的变化及hADSCs细胞促红细胞生成素(EPO)、血红素氧化酶1(HO-1)及血管内皮生长因子(VEGF)的表达,ELISA检测培养液中中性粒细胞明胶酶相关脂质运载蛋白(NGAL)和肾损伤分子1(KIM-1)的浓度,EPO、HO-1、VEGF和HIF-1α的浓度变化。结果:(1)与顺铂诱导组相比,共孵育后的HK-2细胞结构损伤减轻、凋亡细胞减少,caspase-3的表达低于顺铂诱导组、Bcl-2的表达水平升高;其中HIF-1α转染的hADSCs与HK-2细胞共孵育后,HK-2细胞损伤明显减轻,凋亡蛋白的变化更为明显。(2)顺铂诱导组培养液中NGAL、KIM-1浓度升高,以KIM-1变化明显(P<0.05);细胞共孵育后培养液中KIM-1浓度降低,转染组较明显。(3)转染后的hADSCs细胞内EPO、HO-1、VEGF的蛋白表达升高;培养液中HO-1、VEGF含量增高(P<0.05),HIF-1α、EPO的含量变化不明显。结论:hADSCs与HK-2细胞共孵育后,可以减轻顺铂诱导的HK-2细胞结构损伤、抑制细胞凋亡,HIF-1α转染hADSCs后其作用更为明显,可能与细胞保护因子的高表达有关。Objective：To transfect hypoxia inducible factor-1α （HIF-1α） to human adipose-derived stem cells （hADSCs） and then coincubated with HK-2 cells induced by cisplatin. Methodology：The hADSCs were transtfeted by lentiviral vector containing EGFP-HIF-lα and coincubated with HK-2 cells induced by cisplatin. The ultrastructural changes of HK-2 cells were observed by electron microscopy. Apoptosis index of HK-2 cells was tested by TUNEL. Western blot was performed to examine the expression of caspase-3 and Bcl-2 in HK-2 cells and also the expression of EPO, ttO-1 and VEGF in hADSCs cells. The concentrations of NGAL, KIM-1, EPO, HO-1, VEGF and HIF-lcx in media were tested by EL1SA. Results： The apoptosis in coincubation groups was decreased compared with the cisplatin group. The expression of caspase-3 in HK-2 cells in coincubation groups was lower than that in cisplatin group and the expression of Bcl-2 in HK-2 cells in eoineubation groups was increased. The injury of HK-2 cells coincubated with hADSCs tranfected with HIF-1α was lessened. The concentration of KIM-1 increased markedly in cisplatin-indueed HK-2 group and that was decreased after eoineubated with hADSCs. The expression of EPO, HO-1 and VEGF in hADSCs ceils tranfected with HIF- 1α was increased compared with the vector transfected group. The concentration of HO-1 and VEGF in media of gene- transfected group was higher than that in vector transfeeted group and the variation of HIF-1 ct and EPO were not obvious.Conclusion：The hADSCs could lessen the apoptosis of HK-2 cells induced by cisplatin, especially after transfected with HIF-1α and the protection may be related to the higher expression of growth factors.

关 键 词：低氧诱导因子1Α 人脂肪源性干细胞 基因转染 肾小管上皮细胞 

分 类 号：R363[医药卫生—病理学]