中介素对急性肾衰竭的肾脏保护作用  

The renal protective effect of IMD on acute renal failure

作　　者：崔艳丽[1] 何利珍[1] 李小亮 CUI Yanli;HE Lizhen;LI Xiaoliang(Laboratory of Second People’s Hospital of Jiaozuo City,JiaoZuo,Henan 454000,China)

机构地区：[1]焦作市第二人民医院检验科

出　　处：《安徽医药》2020年第1期73-77,共5页Anhui Medical and Pharmaceutical Journal

摘　　要：目的探讨中介素在急性肾衰竭病人血清中的表达及其对肾脏的保护机制。方法选取焦作市第二人民医院2015年12月至2018年6月间确诊的急性肾衰竭病人45例和健康对照组30例,采用酶联免疫吸附试验检测血清中介素水平,寻找血清中中介素水平与急性肾衰竭的相关性。另外,建立大鼠肾小管上皮细胞(NRK⁃52E)缺氧/复氧模型,模拟急性肾衰竭肾脏缺血再灌注过程,MTT法检测细胞活性,流式细胞术检测细胞凋亡情况,RT⁃PCR和蛋白质印迹法检测自噬标志蛋白beclin⁃1、微管相关蛋白轻链3(LC3)和磷脂酰肌醇3⁃激酶/蛋白激酶B/雷帕霉素靶蛋白(phosphatidylinositol 3⁃kinase/protein kinase B/mam⁃malian target of rapamycin,PI3K/Akt/mTOR)信号相关蛋白表达水平,以进一步探索中介素对肾脏的保护作用。结果急性肾衰竭病人血清中中介素表达水平明显高于健康对照组者。大鼠肾小管上皮细胞经过缺氧/复氧处理后,细胞的存活率为(45.3±4.5)%,与对照组(100±1.4)%相比显著下降(P<0.05)。采用不同浓度的中介素(2μmol/L、4μmol/L、6μmol/L)对细胞进行干预后,细胞存活率分别为(55.9±2.6)%,(75.4±3.2)%,和(86.7±2.9)%,与缺氧/复氧模型组相比显著上升(P<0.05)。细胞在缺氧再复氧后发生了严重的细胞凋亡现象,凋亡率为(42.3±3.6)%,与对照组(10.2±1.7)%相比显著上升(P<0.05),而经过中介素(4μmol/L)预孵育4 h的细胞凋亡率为(27.6±3.1)%,与缺氧/复氧模型组相比显著下降(P<0.05)。随着中介素浓度升高,PI3K/Akt/mTOR信号通路关键蛋白磷酸化水平显著降低(P<0.05),beclin⁃1、LC3⁃Ⅱ/LC3⁃Ⅰ表达量升高,细胞自噬水平显著升高;与中介素单独处理组相比,PI3K抑制剂LY290042与中介素(2μmol/L)联合用组,PI3K、Akt、mTOR磷酸化水平极显著下降(P<0.05),beclin⁃1、LC3⁃Ⅱ/LC3⁃Ⅰ表达量极显著上升,细胞自噬水平极显著升高。结论中介素在急性肾衰竭病人血清中高Objective To investigate the expression of intermedin(IMD)in serum of patients with acute renal failure(ARF)and its protective mechanism to the kidney.Methods Form December 2015 to June 2018,The Second People’s Hospital of Jiaozuo was selected a total of 45 cases of ARF patients and 30 cases of healthy volunteers were selected.The serum IMD levels were de⁃tected by enzyme⁃linked immunosorbent assay(ELISA)to explore the correlation between serum IMD levels and ARF.In addition,the in vitro hypoxia/reoxygenation(I/R)model was established in the rat renal tubular epithelial cells(NRK⁃52E)to further ex⁃plore the protective effect of IMD on ARF.Cell viability was detected by MTT assay.Cell apoptosis was measured by flow cytometry.The protein levels of two marker molecules of autophagy including beclin⁃1 and LC3,as well as phosphatidylinositol 3⁃kinase/pro⁃tein kinase B/mammalian target of rapamycin(PI3K/Akt/mTOR)related proteins,were detected by RT⁃PCR and western blotting.Results The serum levels of IMD in patients with ARF was significantly higher than that in healthy controls.After I/R stimulation,the cell viability of NRK⁃52E cells was significantly decreased(45.3±4.5)%as compared to the control group(100±1.4)%.After IMD intervention(2μmol/L,4μmol/L,6μmol/L),cell viability was significantly increased to(55.9±2.6)%,(75.4±3.2)%,and(86.7±2.9)%,respectively.Cell apoptotic rate in I/R group(42.3±3.6)%was significantly upregulated when compared with control group(10.2±1.7)%,while treatment with IMD(4μmol/L)markedly reduced the apoptotic rate(27.6±3.1)%.In addition,the phos⁃phorylation levels of key proteins in PI3K/Akt/mTOR signaling pathway were significantly decreased by IMD intervention in a dose⁃dependent manner(P<0.05).The expression levels of beclin⁃1 and LC3⁃Ⅱ/LC3⁃Ⅰwere also increased after IMD intervention.The PI3K inhibitor LY290042 caused significant decrease in phosphorylation levels of PI3K,Akt and mTOR,as well as increase in be⁃clin⁃1 and LC3⁃Ⅱ/LC3⁃Ⅰi

关 键 词：降钙素基因相关肽 肾功能不全 磷酸肌醇3⁃激酶类 凋亡抑制蛋白质类 自噬 再灌注损伤 中介素 磷脂酰肌醇⁃3⁃激酶/蛋白激酶B/哺乳动物雷帕霉素蛋白信号通路 

分 类 号：R73[医药卫生—肿瘤]