骨髓间质干细胞输注对庆大霉素致急性肾小管损伤细胞增殖和凋亡的影响

Effects of infusion of bone marrow mesenchymal stem cells on proliferation and apoptosis in gentamicin-induced acute renal failure in rat

目的观察骨髓间质干细胞(MSCs)对庆大霉素导致的急性肾功能衰竭(ARF)的疗效,并探讨其可能的机制。方法建立庆大霉素皮下注射致大鼠ARF模型。雄性SD大鼠MSCs用4,6-二乙酰基-2-苯基吲哚(DAPI)标记。雌性SD大鼠随机分为对照组(C组)、MSCs组(M组)和DMEM-F12组(D组)。C组未给予任何处理;M组在每次给予庆大霉素的同时经尾静脉静注DAPI标记的MSCs;D组在每次给予庆大霉素的同时注入无血清DMEM-F12。7d后观察血尿素氮(BUN)和血肌酐(Scr)以及肾脏病理改变(HSK评分),采用原位末端标记法检测细胞凋亡指数(AI),免疫组化法检测增殖细胞核抗原(PCNA)、Bcl-2、Fas/FasL的表达,并观察DAPI标记的MSCs在受体大鼠肾脏的分布情况。结果M组的HSK评分、BUN、Scr(分别为5.5±0.8、11.6±2.2mmol/L、93.5±13.1μmol/L)均低于D组(分别为10.2±0.8、22.2±6.3mmol/L、204.4±53.1μmol/L,P<0.05);M组肾小管上皮细胞PCNA+细胞数、AI(分别为20.8%±3.7%、15.73%±2.43%)低于D组(分别为42.1%±5.2%、23.76%±4.30%,P<0.05)。M组Bcl-2表达(35.27%±2.93%)高于D组(25.28%±3.10%,P<0.05);M组Fas、FasL(分别为21.53%±3.72%、20.50%±3.71%)均低于D组(37.28%±11.14%、37.66%±10.59%,P<0.05)。整个实验期间未发现MSCs定位于肾组织中。结论外源性MSCs可以促进ARF损伤后肾小管上皮细胞的增殖并减少细胞凋亡,促进Bcl-2和抑制Fas/FasL表达,从而有利于肾小管损伤的早期恢复。

Objective To investigate the effects of mesenchymal stem cells （MSCs） on the recovery of gentamicin-induced acute re hal failure （ARF）, and explore its possible mechanism. Methods MSCs were isolated from a male SD rat and labeled with DAPI before in jection. ARF rat model was reproduced by hypodermic injection of gentamicin. At the time that gentarncin was injected, MSCs were injected through tail vein. Thirty SD rats were randomly divided into three groups （10 each）： MSCs injection group （group M）, DMEM-F12 infusion group （group D） and control group （group C）. Rats were sacrificed on day 7 after gantamiein injection. The levels of serum Scr and BUN were determined, and the renal tissue was sampled for pathological observation. Cell apoptotic index （AI） was calculated after terminal deoxynucleotidyl transferase mediated d-UTP nick end labeling （TUNEL）. The expressions of Bcl-2, Fas and FasL proteins and PCNA were determined by immunohistochemical technique. The distribution of DAPI positive MSCs in kidney of recipient SD rats was examined by immunohistochemistry. Results HSK score （5.5±0.8 vs 10.2±0.8, P〈0.05）, BUN （11.6±2.2 mmol/L vs22.2±6.3mmol/L, P〈0. 05） and Scr （93.5±13. 1μmol/L vs 204.4±53.1μmol/L, P〈0.05） were lower in group M than those in group D. PCNA positive cells （20. 8%±3. 7% ws 42. 1%±5. 2%, P〈0. 05） and AI （15. 73%±2. 43% vs 23. 76%±4. 30%, P〈0. 05） in group M were lower than those in group D. The expressions of Bcl-2 in group M were higher than those in group D （35.27% ± 2.93% vs 25. 28%±3. 10%, P〈0. 05）, and the expressions of Fas （21.53±3. 72% vs 37. 28%±11.14%, P〈0. 05）, FasL （20. 50±3.71% vs 37. 66%±10. 59%, P〈0.05） were lower in group M than those in group D. MSCs were not trapped in renal tissue of MSC-treated rats. Conclusions MSCs can accelerate proliferation and decrease apoptosis of renal tubular epithelial cells after ARF, promote the expression of Bcl-2 and restrain the expressions of Fas and FasL, and improve the recovery of gentamicin-induced acute renal tubular epithelial ceils damage.