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雷帕霉素在小鼠肾脏缺血再灌注损伤中对自然杀伤T细胞的调控作用

The regulatory effect of rapamycin to natural killer T cells in the murine ischemia-reperfusion injury model

摘要:

目的 观察新型免疫抑制剂雷帕霉素在小鼠肾脏缺血再灌注损伤早期的保护作用及其对自然杀伤T(NKT)细胞募集过程的影响.方法 45只C57BL/6小鼠随机分为假手术(Sham)组、缺血再灌注损伤(IR)组和雷帕霉素给药(Rapa)组,后两组建立双侧肾脏缺血再灌注损伤模型,Rapa组给予雷帕霉素灌胃,术后1d采集标本,评价肾功能和肾组织病理学损伤,采用原位缺口末端标记法(TUNEL)测定肾实质细胞凋亡水平并采用流式细胞术检测NKT细胞在脾脏、肾脏及外周血的淋巴细胞中所占比例的变化,采用实时定量反转录聚合酶链反应(RT-qPCR)检测与NKT细胞迁移相关的趋化因子的表达水平.结果 Rapa组小鼠在肾脏缺血再灌注损伤的早期比较于IR组血清肌酐和尿素氮水平明显下降,病理损伤减轻,肾脏实质细胞凋亡减少.Rapa组的NKT细胞在脾脏总淋巴细胞中所占比例为(0.0764 ±0.007 8)%,较IR组[(0.1169 ±0.0060)%]显著下降(P<0.05).而Rapa组小鼠的外周血中NKT细胞所占比例为(0.205 6±0.002 8)%,较IR组[(0.1320 ±0.001 2)%]显著上升(P<0.05).Rapa组的肾脏总淋巴细胞中NKT细胞所占比例为(0.1169 ±0.0006)%,较IR组[(0.022 8 ±0.001 3)%]也显著上升(P<0.05).Rapa组的肾脏组织中CXC趋化因子配体9(CXCL9)和CXC趋化因子配体10(CXCL10) mRNA表达水平分别0.74±0.04和0.72 ±0.01,其受体CXC趋化因子受体3(CXCR3)阳性的NKT细胞在肾脏总淋巴细胞中的比例为(1.561 0±0.062 8)%,较IR组均明显增加(P<0.05).结论 雷帕霉素可以通过趋化因子CXCL9、CXCL10及其受体CXCR3向肾脏募集具有保护作用的NKT细胞发挥对小鼠肾脏缺血再灌注损伤的保护作用.

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abstracts:

Objective To investigate the effects of rapamycin on kidneys exposed to ischemia-reperfusion (IR) injury in early stage and on trafficking of natural killer T (NKT) cells in a murine model.Methods 45 C57BL/6 mice were randomly divided into sham operation group,IR group and rapamycin-treated group.Mice were subjected to kidney 30 min ischemia followed by 24 h reperfusion and rapamycin (2.5 ml/kg) was administered by garage.Renal function and histological changes were assessed.Renal cell apoptosis was detected by TdT-mediated dUTP nick end labeling (TUNEL) assay.The proportion of NKT cells in peripheral blood,spleen and kidney was detected by flow cytometry.The chemokines and corresponding receptor involved in NKT cell trafficking were determined respectively.Results Compared with IR group,the serum creatinine and urea nitrogen level in Rapa group declined,pathological injury ameliorated and apoptotic renal cells decreased.In rapamycin-treated group,the frequency of NKT cells in spleen was (0.076 4 ± 0.007 8) %,significantly decreased than that in IR group [(0.116 9 ± 0.006 0) %,P < 0.05].While the frequency of NKT cells in peripheral blood was (0.205 6 ± 0.002 8)%,which is augmented in comparison with that in IR group [(0.132 0 ± 0.001 2)%,P <0.05].Similarly,the frequency of NKT cells in renal lymphocytes was (0.116 9 ± 0.000 6) %,more than that in IR group [(0.022 8 ± 0.001 3) %,P < 0.05].In addition,the frequency of CXC chemokine receptor 3 (CXCR3) positive NKT cell in the kidney was (1.561 0 ± 0.062 8)% and mRNA expression of chemokines CXC chemokine ligand 9 (CXCL9) and CXC chemokine ligand 10 (CXCL10),the ligands of CXCR3,were 0.74 ±0.04 and 0.72 ±0.01 respectively,which were evidently augmented compared with IR group (P < 0.05).Conclusion Rapamycin may recruit NKT cells from spleen to the IR-induced kidney to ameliorate renal IR injury in the early stage,which is mediated by the interaction of chemokine CXCL9,CXCL10 and their receptor CXCR3.

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作者: 张潮 [1] 王硕 [1] 王宣传 [1] 李佳蔚 [2] 戎瑞明 [1]
作者单位: 复旦大学附属中山医院泌尿外科, 上海,200032 [1] 200032,上海市器官移植重点实验室 [2]
期刊: 《中华实验外科杂志》2016年33卷12期 2678-2681页 ISTICPKU
栏目名称: 泌尿外科
DOI: 10.3760/cma.j.issn.1001-9030.2016.12.017
发布时间: 2017-02-13
基金项目:
国家自然科学基金 上海市自然科学基金(12ZR1405500)National Natural Science Foundation of China Science and Technology Commission of Shanghai Municipality
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