MG132对高糖作用下大鼠腹膜间皮细胞ERK1／2及结缔组织生长因子表达的影响

Effect of MG132 on the expression of ERK1/2 and connective tissue growth factor in rat peritoneal mesothelial cells induced by high glucose

目的探讨在高糖作用下泛素蛋白酶体途径（UPP）对大鼠腹膜间皮细胞（RPMC）细胞外信号调节激酶1／2（ERK1／2）、结缔组织生长因子（CTGF）蛋白表达的影响，并用蛋白酶体抑制剂MG132为阻断剂，探讨泛素蛋白酶体途径在腹膜纤维化中的作用。方法胰蛋白酶消化法原代和传代培养RPMC，经鉴定后第2代用于实验。随机分为对照组、不同浓度葡萄糖组（1．5％、2．5％、4．25％葡萄糖作用24h）、高糖作用不同时间组（2．5％葡萄糖分别作用于RPMC0、12、24、48h）和MG132组（0．5、1、2umol／LMG132预孵30min后加2．5％葡萄糖作用24h）。Western印迹法检测ERK1／2蛋白的表达。ELISA法检测细胞培养上清液中CTGF蛋白的含量。结果与对照组相比，高糖可刺激RPMC磷酸化（P）ERKI／2蛋白表达上调，24h时达高峰（P〈0．01），48h时下调，但仍高于正常组（P〈0．01）。高糖诱导RPMCCTGF蛋白表达增高，且呈时间、剂量依赖性（P〈0．05）。与高糖组相比，MG132能明显降低高糖所致的腹膜间皮细胞p-ERK1／2、CTGF蛋白的表达（P〈0．05）。结论MG132可降低由葡萄糖诱导的腹膜间皮细胞p-ERK1／2、CTGF蛋白的表达；泛素蛋白酶体途径参与了由高糖诱导的腹膜纤维化，阻断泛素蛋白酶体途径有利于防治腹膜纤维化。

Objective To observe the effect of MG132 on the expression of extracellular regulated kinase 1/2 （ERK1/2） and connective tissue growth factor （CTGF） in rat peritoneal mesothelial cells （RPMCs） induced by high glucose. Methods RPMCs were isolated, cultured and passaged by trypsin, then identified. The second generation of cultured RPMCs were used in the experiment. RPMCs were divided into normal control group, high glucose （1.5%, 2.5%, 4.25%） for 24 hours, high glucose （2.5%） for 0, 12, 24, 48 hours, incubated with MG132 （0.5, 1, 2 umol/L） for half an hour and then with high glucose （2.5%） for 24 hours. ERK1/2 protein was detected by Western blotting, and CTGF protein in supernatant was detected by ELISA. Results Compared with the control group, the expression of p- ERK1/2 was significantly increased in the groups stimulated by high glucose （P 〈 0.01）, reached the peak at 24th hour （P 〈 0.01）, and then the expression decreased at 48th hour, but still was higher than that in the normal control group （P 〈 0.01）. CTGF protein expression of RPMCs induced by high glucose increased, in time- and dose-dependent manner （P 〈 0.05）. MG132 could significantly decrease the expression of ERK1/2 and CTGF induced by high glucose （P 〈 0.05）. Conclusions MG132 can decrease the expression of p-ERK1/2 and CTGF in RPMCs induced by high glucose. The ubiquitin proteasome pathway participates in the development of peritoneal fibrosis, and blocking the way may contribute to the prevention of peritoneal fibrosis.