缺血后处理对大鼠肢体缺血再灌注后肾细胞凋亡的抑制作用

发布时间：2018-08-22 08:55

【摘要】：目的:观察缺血后处理对大鼠肢体缺血再灌注(LIR)后肾组织细胞凋亡的影响,探讨其可能机制。方法:30只SD大鼠随机分为对照组、缺血再灌注组(IR组)和缺血后处理加再灌注组(I-postC组),每组10只。建立大鼠肢体缺血再灌注(LIR)模型,即橡皮带环绕大鼠双后肢根部阻断血流4h再恢复血流灌注4h。对照组大鼠仅松弛环绕橡皮带不阻断血流,I-postC组大鼠则在再灌注前附加反复3次缺血5min-再灌注5min操作,即缺血后处理。全自动生化分析仪测各组大鼠血浆肌酐(Cr)、尿素氮(BUN)和C反应蛋白(CRP)水平;免疫组织化学法检测大鼠肾组织凋亡相关蛋白Bcl-2和Bax的表达,采用自动图像分析系统统计其定量结果并计算Bcl-2/Bax比值;TUNEL染色后在激光共聚焦显微镜下观察肾组织细胞凋亡情况,电镜下观察肾组织超微结构。结果:与对照组比较,IR组和I-postC组大鼠血浆Cr、BUN和CRP水平均明显增高(P0.01);与IR组比较,I-postC组大鼠血浆Cr、BUN和CRP水平均明显降低(P0.01)。与对照组比较,IR组和I-postC组大鼠肾组织中Bax和Bcl-2表达水平明显升高(P0.05或P0.01),Bcl-2/Bax比值降低(P0.05);与IR组比较,I-postC组大鼠肾组织中Bax表达水平降低(P0.05),Bcl-2表达水平升高(P0.01),Bcl-2/Bax比值升高(P0.05)。激光共聚焦显微镜下观察,与对照组比较,IR组大鼠肾组织中凋亡细胞明显增多;与IR组比较,I-postC组大鼠肾组织中凋亡细胞明显减少。透射电镜下观察,对照组大鼠肾组织细胞结构清晰完整;IR组大鼠肾组织中肾近曲小管上皮细胞核固缩,溶酶体和致密颗粒沉积增多,线粒体数目减少,部分线粒体嵴断裂或模糊,肾小球足突细胞突起不规则、融合,有空泡现象,粗面内质网扩张;与IR组比较,I-postC组大鼠肾组织中肾小管上皮细胞及肾小球损伤有一定程度改善。结论:LIR可诱发大鼠肾组织细胞凋亡,缺血后处理可抑制肢体缺血再灌注后的肾细胞凋亡,对改善大鼠肾功能有一定作用。
[Abstract]:Aim: to observe the effect of post-ischemic treatment on apoptosis of renal tissue after limb ischemia reperfusion (LIR) in rats and to explore its possible mechanism. Methods Thirty Sprague-Dawley rats were randomly divided into control group, ischemia reperfusion group (IR group) and post-ischemic plus reperfusion group (I-postC group) with 10 rats in each group. The (LIR) model of limb ischemia-reperfusion was established in rats, that is, the rubber band around the roots of the hind limbs blocked the blood flow for 4 h and then recovered the blood flow for 4 h. In the control group, the rats in the I-postC group were treated with repeated 5 min-reperfusion 5min for 3 times before reperfusion, that is, after ischemia. The levels of plasma creatinine (Cr), urea nitrogen (BUN) and C-reactive protein (CRP) were measured by automatic biochemical analyzer and the expression of apoptosis-related proteins Bcl-2 and Bax in renal tissue of rats were detected by immunohistochemical method. The quantitative results were analyzed by automatic image analysis system and the Bcl-2/Bax ratio was calculated by Tunel staining. The apoptosis of renal tissue was observed under laser confocal microscope and the ultrastructure of renal tissue was observed under electron microscope. Results: compared with the control group, the plasma levels of Cr bun and CRP in IR group and I-postC group were significantly increased (P0.01), and those in I-postC group were significantly lower than those in IR group (P0.01). Compared with the control group, the expression of Bax and Bcl-2 in renal tissue of IR group and I-postC group were significantly increased (P0.05 or P0.01) and the ratio of Bcl-2 / Bax was decreased (P0.05), and the expression of Bax in renal tissue of I-postC group was lower than that of IR group (P0.05). The expression level of Bcl-2 was increased (P0.01) and the ratio of Bcl-2 / Bax was increased (P0.05). Compared with the control group, the apoptotic cells in renal tissue of IR group were significantly increased, and those in I-postC group were significantly decreased compared with IR group under confocal laser microscope. Under transmission electron microscope, the renal cell structure of the control group was clear and intact. In IR group, the renal proximal convoluted tubule epithelium was pyknosis, lysosome and dense granules were increased, and the number of mitochondria was decreased. Some mitochondria cristae were broken or blurred, glomerular podocytes were irregular, fused, empty vesicles, rough endoplasmic reticulum dilated, and renal tubular epithelial cells and glomerular damage in I-postC group were improved to some extent compared with IR group. Conclusion: WLIR can induce renal cell apoptosis in rats, and post ischemic treatment can inhibit the apoptosis of renal cells after limb ischemia reperfusion, which can improve the renal function of rats to some extent.
【作者单位】：华北理工大学基础医学院病理生理学系;
【基金】：河北省卫计委医学研究重点项目资助课题(20130060)
【分类号】：R692

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