Klotho蛋白在缺血再灌注急性肾损伤中的动态变化

Dynamic changes of Klotho protein in ischemia-reperfusion of acute kidney injury patients

目的观察Klotho蛋白在缺血再灌注急性肾损伤(acute kidney injury,AKI)中的动态变化。方法将120只Wistar雄性大鼠随机分为正常+生理盐水组(Norm+NS)、假手术+生理盐水组(Sham+NS)、假手术+Klotho组(Sham+Klotho)、缺血再灌注手术+生理盐水组(I/R+NS)、手术+Klotho组(I/R+Klotho),建立缺血-再灌注AKI模型,同时各组于造模成功1h、5h、12h、24h分别处死大鼠6只,分别测定血清尿素氮(blood urea nitrogen,BUN)和血肌酐(serum creatinine,Scr),检测血清Klotho蛋白水平,Klotho mRNA表达情况;应用Pearson直线相关法分析AKI时大鼠肾组织以及血清的Klotho蛋白与Scr水平之间的关系。结果缺血再灌注24h后,与Norm+NS组、Sham+NS组相比,I/R+NS组Scr、BUN以及肾小管损伤评分上升(tScr=3.767,4.145,tBUN=14.533,15.047,t肾小管评分=153.093,7.261;P值分别为0.013,0.009,<0.001,<0.001,<0.001,0.008),肾小管上皮细胞明显变性、坏死,而外源性给与Klotho蛋白的I/R+Klotho组Scr、BUN和肾小管损伤评分较I/R+NS组降低(t值分别为10.220,9.571,11.076;P值均<0.001),空泡变性减少;I/R+NS组血清、肾组织Klotho蛋白以及Klotho mRNA的表达低于Norm+NS组、Sham+NS组。结论缺血再灌注急性肾损伤后小鼠血清及肾组织局部Klotho蛋白表达均下调,Klotho蛋白水平的下降可能与急性肾损害密切相关,Klotho蛋白可能是AKI的保护性因素。

Objective To observe the dynamic changes of Klotho protein in acute kidney injury(AKI)during ischemia-reperfusion,and to further explore the role of Klotho protein in AKI.Methods A total of 120 Wistar male rats were randomly divided into Norm+NS group(normal rats+normal saline),Sham+NS group(sham operation+normal saline),Sham+Klotho group(sham operation+Klotho),I/R+NS group(ischemia-reperfusion+normal saline),and I/R+Klotho group(ischemia-reperfusion+Klotho)to establish the ischemia-reperfusion AKI groups and control groups.After the establishment of the models for 1h,5h,12h and 24h,six rats in each group were sacrificed.Blood urea nitrogen(BUN),serum creatinine(Scr),serum and kidney Klotho protein(ELISA method)and Klotho mRNA(RT-PCR method)were assayed.Pearson linear correlation method was used to analyze the relationship between serum and kidney Klotho protein levels and Scr level during AKI.Results After ischemia-reperfusion for 24h in I/R+NS group,Scr and BUN levels and renal tubular injury score increased as compared with those in Norm+NS group and Sham+NS group(for Scr:t=3.767 and 4.145,P=0.013 and 0.009;for BUN:t=14.533 and 15.047,P<0.001;for renal tubular injury score:t=153.093 and 7.261,P<0.001 and t=0.008),and renal tubular epithelial cells showed significant degeneration and necrosis.In contrast in I/R+Klotho group in which exogenous Klotho protein was supplied,Scr and BUN levels and renal tubular injury score decreased(t=10.220,9.571 and 11.076;P<0.001)and vacuolar degeneration of the renal tubular epithelial cells decreased,as compared with those in I/R+NS group.The content of Klotho protein and the expression of Klotho mRNA in sera and kidney tissues were lower in I/R+NS group than in Norm+NS group and Sham+NS group(P<0.05).Conclusion In rats with ischemia-reperfusion AKI,Klotho protein and the expression of Klotho gene in serum and kidney decreased significantly.The decrease of Klotho protein level is closely related to AKI.Klotho protein may be a protective factor for AKI.