肝再生增强因子在缺血再灌注肾损伤中的表达及其对p38信号通路的影响  

Hypoxia stimulated augmenter of liver regeneration in rat proximal tubular epithelial cells(NRK52E)

作　　者：周娟[1] 张玲[1] 刘杞[1] 廖晓辉[1] 张臣丽[1] 

机构地区：[1]重庆医科大学病毒性肝炎研究所,重庆400010

出　　处：《重庆医科大学学报》2011年第3期266-269,共4页Journal of Chongqing Medical University

基　　金：国家自然科学基金项目资助(编号:30971364;81000299);重庆市科委自然科学基金计划项目资助(编号:CSTC2;010BB5385);重庆市卫生局医学科学技术研究重点项目资助(编号:2008-1-44);重庆市卫生局医学科研计划项目资助(编号:2010-2-126)

摘　　要：目的:观察缺氧及缺氧复氧状态下大鼠肾小管上皮细胞中肝再生增强因子(Augmenter of liver regeneration,ALR)的表达变化,以及对细胞p38丝裂原活化蛋白激酶(Mitogen-activated protein kincses,MAPK)信号通路的影响,探讨ALR对肾脏保护的作用机理。方法:将体外培养的大鼠肾小管上皮细胞(NRK52E)分组,在三气培养箱中缺氧不同的时间(2、4、8、12、18、24 h),复氧(12、24 h),Western blot法检测细胞ALR的蛋白表达,RT-PCR法检测细胞ALR mRNA的表达。缺氧不同的时间(0、15、30、60 min),并在缺氧15 min的细胞中设立对照组r,ALR干预组(20、100μg的rALR)及抑制剂干预组,Western blot检测细胞p38MAPK蛋白的表达。结果:(1)正常培养NRK52E细胞中有ALR mRNA表达,缺氧4 h后,ALR mRNA表达较正常对照显著增加(n=7,P<0.01)。(2)缺氧培养细胞12 h,细胞中ALR蛋白表达量显著高于正常对照组,于18 h达高峰(n=7,P<0.01)。(3)缺氧4 h,复氧12、24 h,ALR mRNA及蛋白表达较正常对照组明显增加。(4)缺氧15 min时p38MAPK蛋白表达最强,低剂量rALR(20μg)能够显著抑制p38MAPK通路的激活,而高剂量rALR(100μg)的抑制作用减弱。结论:缺氧及缺氧复氧均能诱导大鼠肾小管上皮细胞表达ALR增加,其中缺氧复氧能较早激活ALR表达,低浓度的外源性rALR减弱p38MAPK通路的表达,提示ALR对缺氧再灌注损伤大鼠肾小管上皮细胞有保护作用。Objective：To investigate the effect of hypoxia and reoxygenation on augmenter of liver regeneration expression and secretion in NRK52E cells,and conduct initial observation and analysis of exogenous rALR on p38 MAPK signal pathway.Methods： The cultured rat proximal tubular epithelial cell line（NRK52E cells）was divided into several groups,incubated in three gas incubators for different periods of hypoxia（2,4,8,12,18,24 h） and of reoxygenation（12,24 h）,or for various amounts of rALR.ALR mRNA levels were measured by semi-quantitative RT-PCR method;ALR protein expression were measured by Western blot.Hypoxia at different times（0,15,30,60 min）,and 15 min hypoxic cells in the control group,rALR group（20,100 μg of rALR） and SB203580 group,Western blot assayed p38MAPK protein expression cells.Results：（1） Normally cultured NRK52E cells had ALR mRNA expression.After hypoxia for 4 h,ALR mRNA expression was significantly increased compared with normal control（n=7,P0.01）;（2）Hypoxic cells being cultured for 12 h,cell ALR protein was significantly higher than that in the control group,and peaked in 18 h（n=7,P 0.01）.（3）Hypoxia for 4 h,and reoxygenation for 12,24 h,ALR mRNA and protein expression increased significantly compared with the control group;（4）P38MAPK protein expression during hypoxia 15 min was the strongest;low-dose rALR（20 μg） significantly inhibited the activation of p38MAPK pathway,while the inhibition of high dose rALR（100 μg） decreased.Conclusion：Hypoxia and hypoxia/reperfusion can induce the expression of renal ALR.Low concentration and exogenous rALR can reduce the expression of p38MAPK pathway,which indicates that in hypoxia-reperfusion injury,ALR may have protective effects on rat renal tubular epithelial cell.

关 键 词：肝再生增强因子 肾小管 上皮细胞 

分 类 号：R322.6[医药卫生—人体解剖和组织胚胎学]