摘要
目的探讨红细胞生成素(EPO)是否可以抑制高糖诱导的大鼠近端。肾小管上皮细胞凋亡及其相关机制。方法传代培养大鼠近端肾小管上皮细胞(NRK-52E),分为正常对照组(NC组)、渗透浓度对照组(OC组)、高糖组(HG组)、高糖+EPO50U/ml组(E1组)和高糖+EPO 100U/ml组(E2组)。免疫荧光检测NRK-52E细胞有无EPO受体(EPOR)表达。Western印迹检测高糖对EPOR表达的影响。流式细胞仪AnnexinV—FITC/PI双染法检测细胞凋亡指数。荧光探针CM—H2DCFDA检测细胞内活性氧(ROS)的水平。RT—PCR检测bcl-2、bax、capases-3mRNA的表达。结果(1)NRK.52E细胞表达EPOR,且高糖可刺激EPOR表达增加。(2)高糖可诱导NRK-52E细胞凋亡,与葡萄糖相同渗透浓度的甘露醇不能明显诱导细胞凋亡。E1、E2组细胞早、晚期凋亡率显著低于HG组(P〈0.05)。(3)高糖刺激NRK-52E细胞后,细胞内ROS产生增多,bcl-2 mRNA的表达下调,bax、caspase-3 mRNA的表达上调。EPO可以抑制细胞内ROS的产生,上调bcl-2 mRNA表达,下调bax、caspase-3mRNA表达。结论EPO可能通过EPOR的介导,缓解高糖诱导的氧化应激,上调bcl-2mRNA表达,下调bax、caspase-3 mRNA表达,抑制NRK-52E细胞凋亡。
Objective To investigate whether erythropoietin (EPO) can inhibit the proapoptotic effect of high glucose on rat proximal tubular epithelial cells, and the possible mechanisms in which EPO exerts its anti-apoptotic role. Methods Rat proximal tubular epithelial cells (NRK-52E) were divided into 5 groups: normal control group, osmolarity control group, high glucose group, high glucose with EPO (50 U/ml) group and high glucose with EPO (100 U/ml) group. The expression of EPO receptor (EPOR) in NRK-52E cells was examined by immunocytochemistry. The effect of high glucose on the expression of EPOR was detected by Western blotting. The rate of apoptosis was evaluated by flow cytometry Annexin V-FITC/PI double stains. The intracellular ROS was detected using fluorescent probe CM-H2DCFDA. The expression of bcl-2, bax and caspase-3 mRNA were examined by RT-PCR. Results The expression of EPOR was demonstrated in NRK-52E cells, and high glucose could up-regulate the expression of EPOR. High glucose could induce oxidative stress in NRK-52E cells, and up-regulate the mRNA expression of bax and caspase-3, down-regulate the mRNA expression of bcl-2. These effects of high glucose on NRK-52E cells could be reversed by EPO. Conclusion EPO inhibits NRK- 52E cells apoptosis induced by high glucose through attenuating oxidative stress,up-regulating the expression of bel-2 mRNA and down-regulating the expression of bax and caspase-3 mRNA, which may be mediated by EPOR.
出处
《中华肾脏病杂志》
CAS
CSCD
北大核心
2010年第7期537-542,共6页
Chinese Journal of Nephrology
关键词
红细胞生成素
活性氧
细胞凋亡
高糖
肾小管上皮细胞
Erythropoietin
Reactive oxygen species
Apoptosis
High glucose
Renal tubular epithelial cells