肾缺血后处理对大鼠Kim-1表达的影响及对缺血再灌注损伤的保护作用

目的：探讨缺血后处理(ischemic postconditioning, IPO)对大鼠肾缺血再灌注损伤（ischemia reperfusion injury, IRI）的保护作用及与肾损伤分子1（kidney injury molecule-1, Kim-1）表达的关系，以及Kim-1能否作为敏感的标记物反映IPO早期的保护效果，为IPO应用于泌尿外科临床寻找早期、有效的监测指标。方法：雄性SD大鼠85只，体重240~300 g，缺血再灌注组（IRI）30只，缺血后处理组（IPO）30只，假手术组（Sham）25只。经腹正中切口切除大鼠右肾建立左侧孤立肾模型。IRI组夹闭左肾动脉45 min后开放，IPO组在IRI模型基础上，恢复血流前给予反复6次10 s供血-10 s缺血的后处理。各组于术后6 h、12 h、24 h、48 h、72 h每时点随机选取5只大鼠，分别取血和肾皮质标本。测定血尿素氮（BUN）、肌酐（Cr）水平，实时PCR（RT-PCR）检测肾组织Kim-1 mRNA表达量。IRI和IPO组各随机取5只大鼠，于0 h、6 h、12 h、24 h、48 h、72 h这6个时点取尿液样本，ELISA法检测尿Kim-1含量。肾病理组织切片观察3组间的差别。结果：ELISA检测显示，IRI与IPO组尿液Kim-1分子均在再灌注6 h开始上升，24 h达峰值，6 h、12 h、24 h、72 h各组间比较差异有统计学意义（P<0.05）。RT-PCR显示，Kim-1 mRNA于再灌注6 h开始上升，24 h达峰值，各时间点IRI组显著高于IPO组（P<0.05），24 h后IPO组Kim-1 mRNA下降速度较快（P<0.05）。BUN、Cr于术后12 h明显上升，比Kim-1分子升高时间延后，24 h后IRI组较IPO组显著增高（P<0.05）。肾组织病理切片结果提示，12 h、24 h、48 h IPO组肾损伤较IRI组明显减轻。结论：IPO能减轻大鼠肾IRI，并可以降低IRI后肾Kim-1表达量。Kim-1作为急性肾损伤的标记物和保护因子之一，较其他指标可更早、更有效地反映IPO的保护作用。

Expression of kidney injury molecule-1 in renal ischemic postconditioning and its protective effect against renal ischemia-reperfusion injury in rats

Objective:To investigate whether the protective mechanism of ischemic postconditioning(IPO) against renal ischemia reperfusion injury(IRI) is related to the expression of kidney injury molecule-1(Kim-1),and whether Kim-1 can reflect the protective effect of ischemic postconditioning as a sensitive biomarker in rats so as to find early and effective indicators to use IPO in clinical research of urology in future.Methods: Eighty-five male SD rats(240-300 g) were randomly divided into three groups(IRI: n=30;IPO: n=30;sham operation group: n=25).After the right nephrectomy through median abdominal incision to make the solitary kidney model,the left renal artery was separated from the renal pedicle.In IRI group,the left renal artery was blocked for 45 min before reperfusion.In IPO group,six cycles of 10 s reperfusion-10 s reocclusion of the left kidney were performed after 45 min ischemia.The blood samples and the kidney cortex tissues were taken at the time points of 6 h,12 h,24 h,48 h and 72 h after reperfusion,with 5 rats for each time point.The plasma samples were obtained for urea and creatinine detection.A modified quantitative RT-PCR was used to quantitate the Kim-1 mRNA expression of renal tissue.Five rats were chosen from groups IRI and IPO respectively to obtain the urine samples of each rat,0 h,6 h,12 h,24 h,48 h,and 72 h after reperfusion.The Kim-1 levels of urine were detected by ELISA.Pathological examinations were performed to check the differences between the three groups.Results: Urine tests and RT-PCR showed that,in IRI and IPO groups,Kim-1 and its mRNA levels began to rise after 6 h of reperfusion and reached the peak at 24 h synchronously.Urine Kim-1 levels in IRI group were significantly higher than those of the IPO group at 6 h,12 h,24 h,72 h(P<0.05).The Kim-1 mRNAs in IPO group were lower than those in IRI group at the time points of 6 h,12 h,24 h,and 48 h(P<0.05),and the levels decreased more rapidly after 24 h.BUN and Cr significantly increased at 12 h,which were later than the Kim-1 and Kim-1 mRNA.The BUN and Cr levels in IRI group were higher than those in IPO group after 24 h(P<0.05).The pathological examination showed that there was less epithelial injury in IPO group.Conclusion: Ischemic postconditioning can attenuate the renal ischemia reperfusion injury and downregulate the expression of Kim-1 and Kim-1 mRNA.Kim-1,as a biomarker and protective factor of acute kidney injury(AKI),can sensitively reflect the renal protective effect by ischemic postconditioning.