重组大鼠肝再生增强因子抑制庆大霉素诱导的肾小管上皮细胞凋亡的机制研究

Mechanism responsible for inhibitory effect of recombinant rat augmenter of liver regeneration on renal turbular epithelial cell apoptosis by gentamycin

目的:探讨重组大鼠肝再生增强因子(rrALR)在体外对庆大霉素(GM)诱导的大鼠近端肾小管上皮细胞(NRK-52E细胞)凋亡的影响及其作用机制。方法:将NRK-52E细胞分为正常对照组、庆大霉素处理组(GM1.6g/L)和rrALR干预组(分别加入GM和不同浓度的rrALR),培养细胞24h、48h。吖啶橙/溴乙啶(AO/EB)染色、DNA琼脂糖凝胶电泳及Annexin V/PI双染流式细胞术检测各组细胞的凋亡状态。Western blotting和RT-PCR检测各组细胞不同时点Bcl-2和Bax蛋白、mRNA的表达。结果:(1)rrALR对庆大霉素诱导的NRK-52E细胞凋亡具有抑制作用(P<0.05)。(2)rrALR能够呈剂量依赖方式增加细胞Bcl-2蛋白及核酸的表达(P<0.05)、降低细胞Bax蛋白及核酸的表达(P<0.05),使Bcl-2/Bax比值升高(P<0.05)。结论:rrALR可以通过调节Bcl-2家族Bax和Bcl-2蛋白及核酸的表达水平,抑制庆大霉素诱导的肾小管上皮细胞凋亡,减轻肾毒性药物对小管上皮细胞的损伤。

AIM ： To investigate the effects of recombinant rat augmenter of liver regeneration （rrALR） on apoptosis of renal tubular cells （ NRK - 52E cells） induced by gentamycin sulfate （GM）. METHODS： The cultured NRK -52E cells were divided into four groups： normal control cells, cells with GM （GM 1. 6 g/L） or GM and rrALR （15 mg/L or 25 mg/L） treatments. The apoptosis of cultured cells were assessed at 24 h, 48 h by AO/EB staining, DNA agarose gel electrophoresis analysis and flow cytometry using Annexin V - FITC and propidium iodide （PI） staining. The protein and mRNA expressions of Bcl - 2 and Bax were detected by Western blotting and RT - PCR, respectively. RESULTS： （ 1 ） rrALR inhibited NRK - 52E cells apoptosis induced by GM （ P 〈 0. 05 ）. （2） rrALR promoted the expression of Bel - 2 protein and mRNA, but inhibited the Bax protein and mRNA expression （P 〈 0. 05） in cultured NRK - 52E cells in a dose -dependent manner. The value of Bcl -2/Bax increased. CONCLUSION： rrALR inhibits renal tubular epithelial cell apoptosis and ameliorates cell injury induced by nephrotoxic drug GM presumably via the regulation of Bcl - 2 and Bax protein and mRNA expressions.