Legumain/TRAF6通路在肾脏缺血-再灌注损伤中的作用研究

The Role of Legumain/TRAF6 Pathway in Renal Ischemia-Reperfusion Injury

目的本研究旨在探讨Legumain/TRAF6通路在肾脏缺血-再灌注损伤中的作用机制。方法将50只SPF级C57BL/6小鼠随机分为5组,每组10只,A组:假手术组,B组:模型组,C组:Legumain低表达,D组:Legumain低表达+TRAF6过表达组,E组:Legumain低表达+TRAF6低表达组,再灌注24 h时处死各组小鼠。ELISA测定小鼠血清肌酐(SCr)、尿素氮(BUN)、乳酸脱氢酶(LDH)、丙二醛(MDA)、肾损伤分子1(KIM-1)含量。TUNEL检测试剂盒检测各组小鼠肾组织细胞凋亡水平,Western blot和RT-PCR法检测Legumain、TRAF6蛋白及mRNA表达水平。结果与A组相比,B组SCr、BUN、LDH、MDA、Kim-1水平、肾组织细胞凋亡率、Legumain、TRAF6蛋白及mRNA表达水平均升高(P<0.05),与B组相比,C组逆转上述五种标志物、细胞凋亡率、Legumain、TRAF6蛋白及mRNA表达均降低(P<0.05),与C组相比,D组逆转了上述五种标志物、细胞凋亡率、Legumain、TRAF6蛋白及mRNA表达的降低,呈现升高趋势(P<0.05),而E组上述五种标志物、细胞凋亡率、Legumain、TRAF6蛋白及mRNA表达均继续降低(P<0.05)。结论Legumain/TRAF6通路参与肾脏缺血-再灌注损伤,抑制Legumain表达可通过TRAF6减轻肾脏缺血-再灌注损伤。

Objective To investigate the role of Legumain/TRAF6 pathway in renal ischemia-reperfusion injury.Methods A total of 50 SPF grade C57BL/6 mice were randomly divided into 5 groups with 10 mice in each group,group A:sham operation group,group B:model group,group C:Legumain low expression group,group D:Legumain low expression group+TRAF6 overexpression group,group E:Mice with low Legumain expression and low TRAF6 expression were killed after 24 h reperfusion.serum creatinine(SCr),blood urea nitrogen(BUN),lactate dehydrogenase(LDH),Malonodialdehyde(MDA),Kidney injury molecule 1(KIM-1)content were determined by ELISA.TUNEL assay kit was used to detect apoptosis of renal cells in each group,and Western blot and RT-PCR were used to detect Legumain and TRAF6 protein and mRNA expression levels.Results Compared with group A,SCr,BUN,LDH,MDA,Kim-1 levels,renal cell apoptosis rate,Legumain,TRAF6 protein and mRNA expression levels in group B were increased(P<0.05).Group C reversed the above five markers,apoptosis rate,Legumain,TRAF6 protein and mRNA expression were decreased(P<0.05),Compared with group C,group D reversed the decrease of the above five markers,apoptosis rate,Legumain,TRAF6 protein and mRNA expression,and showed a increase trend(P<0.05).In group E,the above five markers,apoptosis rate,Legumain,TRAF6 protein and mRNA expressions continued to be decreased(P<0.05).Conclusions The Legumain/TRAF6 pathway is involved in renal ischemia-reperfusion injury,and inhibition of Legumain expression can attenuate renal ischemia-reperfusion injury through TRAF6.