肢体远隔缺血后适应通过磷脂酰肌醇3激酶／Akt通路保护大鼠局灶性脑缺血再灌注损伤

Remote limb ischemic postconditio ning protects focal cerebral ischemia-reperfusion injury in rats via phosphatidyUnositol 3 kinase/Akt pathway

目的 通过检测肢体远隔缺血后适应（limb ischemic postconditioning, LIP ）后p-Akt蛋白及胱天蛋白酶-9和Bcl-2 mRNA表达,探讨磷脂酰肌醇3激酶（phosphatidyl inositol 3 kinase, PI3K ）/Akt通路在LIP保护大鼠局灶性脑缺血再灌注损伤中的作用.方法 42只Wistar大鼠随机分为假手术组、脑缺血再灌注组和LIP组.缺血再灌注组和LIP组均采用线栓法制作大脑中动脉缺血再灌注模型. LIP组在脑缺血2 h后再灌注前实施3个循环健侧股动脉LIP（5 min缺血/5 min再灌注）.采用2,3,5-氯化三苯基四氮唑染色测定脑梗死体积,免疫组化染色法检测p-Akt蛋白表达,原位杂交法检测胱天蛋白酶-9和Bcl-2 mRNA表达.结果 与脑缺血再灌注组比较,LIP组脑梗死体积显著缩小（P〈0.05）,p-Akt蛋白和Bcl-2 mRNA表达水平显著增高（P均〈0.05）,胱天蛋白酶-9 mRNA表达水平显著降低（P〈0.05）.结论 LIP可缩小局灶性脑缺血再灌注大鼠脑梗死体积,其机制可能与上调p-Akt蛋白和Bcl-2 mRNA表达以及下调胱天蛋白酶-9 mRNA表达有关,提示LIP可通过PI3K/Akt通路减轻脑缺血再灌注损伤.

Objective To investigate the role of phosphatidyl inositol 3 kinase （PI3K）/Akt pathway in the protection of focal cerebral ischemia reperfusion injury in rats with limb ischemic postconditioning （LIP） by detecting the expression levels of p-Akt protein, and caspase-9 and Bcl-2 mRNAs after remote LIP. Methods Forty-two Wistar rats were randomly assigned to 3 groups： sham operation, ischemia-reperfusion （IR） and LIP groups. The middle cerebral artery ischemia-reperfusion injury model was induced by the suture method in the IR group and the LIP group. In the LIP group, three circulatory LIP （ 5 min ischemia/5 min reperfusion） in the contralateral femoral artery were performed before reperfusion 2 h after cerebral ischemia. Infarct volume was measured by 2,3,5-triphenyltetrazolium chloride staining. The expression of p-Akt protein was detected by immunohistochemical staining and the expression levels of cystin-9 and Bcl-2 mRNAs were detected by in situ hybridization. Results Compared with the IR group, the infarct volume in the LIP group was significantly reduced （ P〈0.05）. The expression levels of p-Akt protein and Bcl-2 mRNA significantly increased （all P〈0.05）, and the expression level of caspase-9 mRNA significantly decreased （P〈0.05）. Conclusions LIP can reduce the volume of cerebral infarction in focal cerebral ischemia-reperfusion injury in rats. Its mechanism may be involved in up-regulation of p-Akt protein and Bcl-2 mRNA expression and down-regulation of caspase-9 mRNA expression, suggesting that LIP can alleviate cerebral ischemia-reperfusion injury through PI3K/Akt pathway.