内质网应激在白蛋白诱导肾小管上皮细胞凋亡中的作用  

Effects of endoplasmic reticulum stress on albumin-induced apoptosis in renal proximal tubular cells

作　　者：吴小玮[1] 何娅妮[1] 王惠明[1] 李开龙[1] 丁涵露[1] 吕顺巧[1] 

机构地区：[1]第三军医大学大坪医院肾内科野战外科研究所,重庆400042

出　　处：《中华肾脏病杂志》2009年第3期198-203,共6页Chinese Journal of Nephrology

基　　金：基金项目：国家自然科学基金（30370534,30771002）

摘　　要：目的探讨内质网应激在白蛋白超负荷诱导肾小管上皮细胞凋亡中的作用和分子机制。方法Western印迹法检测肾小管上皮细胞（HKC）内质网伴侣蛋白糖调节蛋白78（GRP78）和内质网应激蛋白CHOP（CCAAT增强子结合蛋白同源蛋白，也称为GADD153）表达与人血清白蛋白（HSA）作用时间和浓度的关系。实时荧光定量PCR法检测CHOP siRNA对CHOP基因转录的抑制情况。Western印迹法检测CHOP siRNA转染后CHOP蛋白水平的变化。膜联蛋白V和碘化丙锭（Annexin V-FITC和PI）双染的流式细胞术检测白蛋白诱导HKC凋亡的变化以及CHOP siRNA对HKC凋亡的影响。结果（1）分别以0、5、10、20g／L白蛋白作用于HKC 24h，GRP78、CHOP蛋白表达及细胞凋亡均随白蛋白浓度的增加而上调，各组间差异有统计学意义（P〈0．01）；以20g／L白蛋白分别作用于HKC 0、6、12、24、36h，GRP78蛋白表达在6h即显著增加，CHOP蛋白表达及HKC凋亡则在12h显著增加，各组间差异有统计学意义（P〈0．01）。（2）CHOP siRNA显著抑制白蛋白诱导的CHOP基因转录及蛋白表达（P〈0．01），对白蛋白诱导的HKC凋亡有显著抑制作用（P〈0．01）。结论白蛋白超负荷可诱导肾小管上皮细胞发生内质网应激，并通过上调促凋亡因子CHOP表达引起肾小管上皮细胞内质网相关的细胞凋亡，这可能是蛋白尿引起肾小管间质病变的重要机制。Objective To investigate the effects and molecular mechanism of endoplasmic reticulum stress （ERS） on albumin-induced apoptosis in renal proximal tubular cells （HKCs）. Methods Western blot was performed to detect the relationship of the expression of glucose-regulated protein 78 （GRP78） and CCAAT/enhaneer-binding protein-homologous protein （CHOP） with the action time and concentration of human serum albumin （HSA）. Expression levels of CHPO mRNA and protein in HKCs after CHOP siRNA transfection were examined by real-time fluorescence quantitative PCR and Western blot respectively. Annexin-V-FITC and PI double staining cytometry was used to detect the apoptosis of HKCs induced by HSA and influenced by CHOP siRNA. Results （1）After HKCs were stimulatde by 0, 5, 10, 20 g/L albmnin for 24 hours respectively, the expression of GRP78, CHOP and HKCs apoptosis were increased with the albumin concentration （P〈0.01）. After HKCs were stimulated by 20 g/L albumin for 0, 6, 12, 24,36 hours respectively, the expression of GRP78 was up-regulated at 6-hour, while CHOP and HKCs apoptosis were increased at 12-hour, and significant differences were found among groups （P〈0.01）. （2） CHOP siRNA significantly inhibited albumin-induced HKC CHOP mRNA and protein expression, as well as HKC apoptosis （P〈0.01）. Conclusions Renal tubular cells exposed to high protein load result in ERS. ERS may subsequently lead to tubular damage by activation of pro-apoptosis factor CHOP.

关 键 词：白蛋白类 内质网 细胞凋亡 CCAAT增强子结合蛋白同源蛋白 肾小管上皮细胞 

分 类 号：R692[医药卫生—泌尿科学]