GSK-3β抑制剂对肾移植冷缺血再灌注损伤的保护作用  

Inhibition of glycogen synthase kinase 3β attenuates cold ischemia/reperfusion injury in rat kidney graft

作　　者：苏帅 张琪琳 王云龙 冉瑞图 尹志康[1] SU Shuai;ZHANG Qilin;WANG Yunlong;RAN Ruitu;YIN Zhikang(Department of Urology,the First Affiliated Hospital of Chongqing Medical University,Chongqing,400016,China)

机构地区：[1]重庆医科大学附属第一医院泌尿外科,重庆400016

出　　处：《第三军医大学学报》2018年第18期1653-1662,共10页Journal of Third Military Medical University

基　　金：重庆市科委基础研究与前沿探索(CSTC2014jcyj A0210)~~

摘　　要：目的分析GSK-3β抑制剂TDZD-8对大鼠肾移植缺血再灌注损伤(ischemia reperfusion injury,IRI)中NF-κB的活化、炎症反应和氧化应激的作用。方法选用250~280 g雄性SD大鼠,建立大鼠肾移植模型和冷IRI模型。肾移植模型:将大鼠分为假手术组、肾移植组、肾移植+5 mg/kg TDZD-8组和肾移植+1 mg/kg TDZD-8组,每组6只;冷缺血再灌注模型:将大鼠分为假手术组、IRI组、IRI+5 mg/kg TDZD-8组和IRI+1 mg/kg TDZD-8组,每组6只。采用Western blot和IHC检测两个模型各组大鼠p-Rel A/p65、p-GSK-3β、GSK-3β、Rel A/p65变化;ELISA检测血清TNF-α、IL-1β、SOD和MDA的变化;HE染色观察肾脏的病理改变,进行肾小管坏死评分;检测血清肌酐和尿素氮。结果与假手术组相比,肾移植组或冷IRI组血清肌酐、尿素氮、TNF-α、IL-1β和MDA活性明显增高(P<0.01),血清SOD活性降低(P<0.01),肾小管坏死评分明显升高(P<0.01);免疫组化检测结果显示p-GSK-3β和p-Rel A/p65磷酸化水平上调(P<0.01),Western blot检测结果显示NF-κB p65活性升高和GSK-3β活性降低(P<0.01),病理损伤加重。使用TDZD-8干预后,大鼠血清肌酐、尿素氮、TNF-α、IL-1β和MDA活性降低(P<0.05),血清SOD活性升高(P<0.05),肾小管坏死评分降低(P<0.05);免疫组化检测结果显示p-GSK-3β和p-Rel A/p65磷酸化水平下调(P<0.05),Western blot检测结果显示NF-κB p65活性降低并且GSK-3β活性增高(P<0.05),病理损伤减轻。结论 GSK-3β抑制剂能降低NF-κB活性,下调NF-κB的磷酸化,减少下游炎症因子的释放,减轻肾脏病理和氧化应激损伤,减轻肾移植缺血再灌注损伤。Objective To investigate the effect of TDZD-8,a glycogen synthase kinase 3β（GSK-3β） inhibitor,on the activation of nuclear factor-κB（NF-κB） and oxidative stress in ischemia reperfusion injury（IRI） in the kidney graft of rats. Methods Male SD rats were randomized into renal transplantation（RT） model group and cold IRI model group. In both groups,the rats were subdivided into sham-operated group,RT group,RT ＋ TDZD-8（5 mg/kg） group and RT ＋ TDZD-8（1 mg/kg） group（n = 6）. Western blotting and immunohistochemistry were used to observe the changes in p-Rel A/p65,p-GSK-3β,GSK-3β and Rel A/p65 in the renal graft,and the changes in serum tumor necrosis factor-α（TNF-α）,interleukin-1β（IL-1β）,superoxide dismutase（SOD） activity and malondialdehyde（MDA）content were determined with enzyme-linked immunosorbent assay（ELISA）. The pathological changes in the graft were observed with HE staining and renal tubular necrosis scores were assessed. A colorimetric assay was used to detect the changes in serum creatinine and urea nitrogen of the rats. Results Compared with shamoperated group,the recipient rats in RT model and cold IRI model groups showed significantly increased serum levels of creatinine,urea nitrogen,TNF-α,IL-1β,and MDA（P 0. 01）,lowered serum SOD activity（P 0. 01）,and increased renal tubular necrosis scores（P 0. 01）; the expression levels of p-Rel A/p65 and p-GSK-3β and NF-κB p65 activity were significantly increased（P 0. 01） and GSK-3β activity was decreased（P 0. 01） in the renal graft. Interventions with TDZD-8 in the 2 models significantly decreased serum levels of creatinine,urea nitrogen,TNF-α,IL-1β,and MDA（P 0. 05）,elevated serum SOD activity（P 0. 05） and lowered the renal tubular necrosis score（P 0. 05）,and decreased p-Rel A/p65 and p-GSK-3β expression levels and NF-κB p65 activity in the graft（P 0. 05）,Western blotting showed decreased NF-κB p65 activity but enhanced GSK-3β activity（P 0. 05）. Conclusion

关 键 词：糖原合成酶3β 转录核因子κB 肾移植 缺血再灌注损伤 

分 类 号：R617[医药卫生—外科学] R692[医药卫生—临床医学]