23 kDa亚型肝再生增强因子在肾纤维化过程中表达水平检测及意义研究

Expression and significance of 23 kDa ALR in renal fibrosis

目的通过构建转化生长因子β1(TGF-β1)诱导的人肾小管上皮细胞(HK-2)上皮-间质转化(EMT)模型及单侧输尿管梗阻术(UUO)模型,观察内源性ALR表达情况。方法体外培养HK-2细胞,给予不同浓度TGF-β1处理,以蛋白质印迹法(Western blotting)检测E-钙黏附蛋白、波形蛋白表达水平,实时荧光定量聚合酶链反应、Westernblotting、细胞免疫荧光检测ALR表达水平。构建UUO模型大鼠,以Western blotting检测不同组别ALR表达水平。结果与对照组相比,TGF-β1处理HK-2细胞E-钙黏附蛋白表达水平降低、波形蛋白表达水平增加(P<0.05),内源性23 k Da ALR m RNA及蛋白表达水平增加(P<0.05),ALR免疫荧光增强。UUO模型大鼠中,与假手术组相比,UUO术后7 d内23 kDa ALR表达水平增加(P<0.05),术后14 d 23 kDa ALR表达水平较术后7 d下降(P<0.05)。结论 TGF-β1处理HK-2细胞及UUO模型大鼠23 kDa ALR表达水平增加;ALR可能参与了线粒体的氧化应激及凋亡过程。

Objective To investigate the expression changes of endogenous augmenter of liver regeneration(ALR)in the pathological process of renal tubular epithelial-to-mesenchymal transition(EMT)induced by transforming growth factor-β1(TGF-β1)and unilateral ureteral obstruction(UUO)rat model.Methods HK-2 cells were exposed to different concentra-tions of TGF-β1,the expression of E-cadherin and Vimentin were detected by Western blotting,and the expression of ALR was analyzed by real time polymerase chain reaction,Western blotting and immunofluorescence.The level of ALR was assessed by Western blotting in different groups of UUO model.Results The expression of E-cadherin was decreased and Vimentin was increased significantly in TGF-β1 groups compared to control group(P<0.05).TGF-β1 activated the mRNA and protein expres-sion of 23 kDa ALR in HK-2 significantly(P<0.05).The level of 23 kDa ALR protein of operated group was markedly higher than sham-operated group within 7 days after surgery,but the level of ALR was significantly decreased in operated group on 14 days after surgery(P<0.05).Conclusions 23 kDa ALR might be highly expressed in TGF-β1 stimulated HK-2 cells and UUO rats,which could be associated with mitochondrial oxidative stress and apoptosis.