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目的探讨自噬在缺血预处理减轻大鼠肢体缺血再灌注心肌损伤中的作用。方法雄性SD大鼠28只,8周龄,体重200~250 g,采用随机数字表法,将其分为4组(ni=7):假手术组(S组)、肢体缺血再灌注组(IR组)、肢体缺血预处理组(IPR组)、自噬抑制剂处理组(3-MA组)。采用夹闭双侧股动脉缺血4 h后恢复灌注4 h的方法建立肢体缺血再灌注模型;采用股动脉夹闭前30 min实施缺血5 min,再灌注5 min,循环3次后建立肢体缺血预处理模型;3-MA组于缺血预处理前30 min腹腔注射3-甲基腺嘌呤1.5 ml/kg。再灌注4 h后处死大鼠取心肌组织,电镜下观察自噬小体形成情况和病理学结果,采用HE染色和TUNEL法检测心肌细胞凋亡情况,采用Western blot法检测LC3-Ⅱ的表达情况。结果与S组相比,IR组、IPR组、3-MA组心肌细胞凋亡比例明显升高,LC3-Ⅱ/LC3-Ⅰ表达上调(P<0.05);与IR组相比,IPR组细胞凋亡降低、LC3-Ⅱ/LC3-Ⅰ表达下调(P<0.05);与IPR组相比,3-MA组细胞凋亡比例升高、LC3-Ⅱ/LC3-Ⅰ表达下调(P<0.05)。结论自噬参与了肢体缺血预处理减轻大鼠肢体缺血再灌注心肌损伤的作用。
Objective To investigate the role of autophagy in ischemic preconditioning to reduce myocardial ischemia-reperfusion injury in rats. Methods Male Sprague-Dawley rats, 8 weeks old, weighing 200-250 g, were randomly divided into 4 groups (n = 7): sham operation group (S group), limb ischemia-reperfusion group (IR group), limb ischemic preconditioning group (IPR group) and autophagy inhibitor treatment group (3-MA group). The model of limb ischemia-reperfusion was established by means of 4 h reperfusion after 4 h occlusion of bilateral femoral artery occlusion. The ischemic reperfusion was performed 5 min before reperfusion and reperfusion for 30 min before femoral artery occlusion Limb ischemic preconditioning model. The 3-MA group was injected intraperitoneally with 1.5 ml / kg 3-methyladenine 30 min before ischemic preconditioning. After 4 h of reperfusion, the rats were killed to take the myocardial tissue. The autophagosome formation and pathological results were observed under electron microscope. The apoptosis of cardiomyocytes was detected by HE staining and TUNEL method. The expression of LC3-Ⅱ was detected by Western blot . Results Compared with S group, the percentage of apoptosis of cardiomyocytes in IR group, IPR group and 3-MA group was significantly increased and the expression of LC3-Ⅱ / LC3-Ⅰ was up-regulated (P <0.05). Compared with IR group, (P <0.05). Compared with IPR group, the proportion of apoptosis in 3-MA group was increased and the expression of LC3-Ⅱ / LC3-Ⅰ was down-regulated (P <0.05) . Conclusion Autophagy participates in the protective effect of limb ischemic preconditioning on myocardial ischemia-reperfusion injury in rats.