锌离子对脂多糖作用下大鼠腹膜间皮细胞转分化的影响及机制  

Zinc protects rat peritoneal mesothelial cells from undergoing lipopolysaccharide-induced epithelial-tomesenchymal transition

作　　者：张秀丽[1,2] 梁单[3] 樊怡[1] 孙立霞[1] 杨丽娜[1] 王力宁[1] 马健飞[1] 

机构地区：[1]中国医科大学第一附属医院肾内科 [2]本溪市中心医院肾内科 [3]中国人民解放军95935部队

出　　处：《肾脏病与透析肾移植杂志》2013年第3期238-243,251,共7页Chinese Journal of Nephrology,Dialysis & Transplantation

摘　　要：目的:研究锌离子(Zn)对脂多糖(LPS)诱导的大鼠腹膜间皮细胞(RPMC)转分化(EMT)的影响及机制,从而为Zn在腹膜透析(PD)中的应用提供理论基础。方法:胰蛋白酶消化法原代培养腹膜间皮细胞、传代、经鉴定后分组:(1)正常对照组;(2)LPS组:100μmol/L作用48h;(3)ZnSO4作用组:30μmol/L ZnSO4作用24h后加100μmol/L LPS作用48h。应用RT-PCR方法和免疫荧光方法检测α平滑肌肌动蛋白(α-SMA)、上皮钙黏素(E-cadherin)、Ⅰ型胶原蛋白(CollagenⅠ)基因表达。应用Western Blotting检测核因子κB(NF-κB)、NF-κB抑制蛋白(I-κB)、磷酸化的NF-κB(p-NF-κB)、磷酸化的I-κB(p-I-κB)蛋白表达;应用流式细胞术,采用活性氧自由基(ROS)检测试剂盒检测各组别的ROS情况。结果:LPS培养48h后细胞形态开始发生变化,由典型的上皮逐渐变为梭形及不规则形,类似肌成纤维细胞,而硫酸锌(ZnSO4)作用组细胞表型改变明显减轻。与对照组相比,LPS组RPMC的α-SMA、CollagenⅠ基因表达明显升高,而E-cadherin基因表达降低;与LPS组相比,ZnSO4作用组RPMC的α-SMA、CollagenⅠ表达明显降低、E-cadherin基因表达增多;LPS组ROS显著高于对照组,ZnSO4作用组ROS显著低于LPS组;与LPS组相比,ZnSO4作用组p-NF-κB、p-I-κB蛋白表达降低。结论:ZnSO4可逆转LPS所致的RPMC的EMT,对腹膜透析过程中所导致的RPMC损伤具有保护作用,其作用机制可能是通过抑制氧化应激反应和NF-κB信号通路转导而发挥作用。Objective To investigate that the effects of Zn on lipopolysaccharide （LPS）-induced epithelial-to-mesenchymal transition （EMT） in rat PMCs （RPMCs） and examined the underlying molecular mechanisms. Methodology： RPMCs were isolated, cultured and passaged by enzymatic disaggregation, then identified by phase contrast inverted microscope, transmission electron microscope with immunocytochemistry method. RPMCs were incubated with 100μM LPS for 48 hours, or pre-incubated with 30μM ZnSO4 for 24 hours with stimulated by 100μM LPS for 48 hours. RPMCs in the control group were just incubated with medium. The expressions of α-SMA, E-cadherin , collagen I were detected by RT-PCR and the expressions of NF-κB, I-κB p-NF-κB, p-I-κB were determined by western blotting. In addition, reactive oxygen species （ROS） assay experiments were performed using the ROS assay kit. Results： Zn supplementation inhibited LPS-induced RPMC EMT significantly, by attenuating ROS assay production. Whereas this LPS-induced EMT could be attenuated by pre-treating the RPMCs with 30 μM ZnSO4, which were evidenced by the reduced up-regulation of α-SMA and collagen I and the ameliorated expression of epithelial protein E-cadherin. Further analysis revealed that Zn supplementation inhibited LPS-induced RPMC EMT likely through inhibition of NF-κB pathways. Conclusion： These results indicate that Zn can inhibit EMT in LPS-induced RPMCs by inhibition of oxidative stress as well as NF-κB pathways activation.

关 键 词：脂多糖 硫酸锌 转分化 大鼠腹膜间皮细胞 腹膜透析 

分 类 号：R965[医药卫生—药理学]