乳化异氟烷预处理对大鼠局灶性脑缺血再灌注损伤的影响

Effect of emulsified isoflurane preconditioning on focal cerebral ischemia-reperfusion injury in rats

目的 评价8%乳化异氟烷预处理对大鼠局灶性脑缺血再灌注损伤的影响.方法 健康雄性成年SD大鼠56只,体重250～280 g,随机分为4组（n=14）,假手术组（S组）仅分离血管,不留置线栓;脑缺血再灌注组（IR组）、乳化异氟烷预处理组（EIP组）和脂肪乳剂组预处理（FIP组）分别腹腔注射生理盐水、8%乳化异氟烷或30%脂肪乳剂7.5 ml/kg,24 h后制备局灶性脑缺血再灌注模型.于再灌注24 h各组取8只大鼠,进行神经功能缺陷评分,然后处死大鼠,取脑组织,测定脑梗死体积;各组处死6只大鼠,取脑组织,检测细胞凋亡情况,计算细胞凋亡率,并测定Bcl-2、Bax、caspase-3和Cyt C的蛋白表达水平.结果 与S组比较,IR组、EIP组和FIP组神经功能缺陷评分和细胞凋亡率升高,脑组织Bcl-2、Bax、caspase-3和Cyt C的蛋白表达上调（P＜0.01）;与IR组和FIP组比较,EIP组神经功能缺陷评分和细胞凋亡率降低,脑梗死体积减小,脑组织Bcl-2蛋白表达上调,Bax、caspase-3和Cyt C 的蛋白表达下调（P＜0.05）;IR组和FIP组各指标比较差异无统计学意义（P＞0.05）.结论 8%乳化异氟烷预处理可减轻大鼠脑缺血再灌注损伤,其机制可能与上调Bcl-2蛋白表达,下调Bax蛋白表达,减少线粒体释放Cyt C,降低caspase-3活化,抑制神经元凋亡有关.

Objective To investigate the effect of emulsified isoflurane preconditioning on focal cerebral ischemia-reperfusion （I/R） injury in rats and the mechanism. Methods Fifty-six healthy male adult SD rats weighing 250-280 g were randomly divided into 4 groups （n = 14）： group Ⅰ sham operation （group S）; group Ⅱfocal cerebral I/R; group Ⅲ emulsified isoflurane preconditioning and group Ⅳ fat emulsion preconditioning. Focal cerebral ischemia was induced by middle cerebral artery occlusion （MCAO） using a nylon thread with rounded tip inserted into internal carotid artery and advanced cranially until resistance was met. MCAO was maintained for 2 h followed by 24 h reperfusion. In group Ⅲ and Ⅳ 8% emulsified isoflurane 7.5 ml/kg and 3% fat emulsion 7.5 ml/kg were injected intraperitoneally at 24 h before MCAO respectively. Neurologic outcome was evaluated at 24 h of reperfusion and scored （0 = no deficit, 4 = unable to crawl, loss of consciousness）. The animals were then killed and brains removed. The infarct size was assessed. The apoptotic cells were detected by TUNEL and calculated. The expression of Bax, Bcl-2, cytochrome C and caspase-3 protein was detected by immunohistochemical staining. Results The neurologic deficit scores were significantly lower in emulsified isoflurane preconditioning group than in I/R group. Preconditioning with emulsified isoflurane significantly decreased the infarct size and the number of apoptotic cells and increased the expression of Bcl-2 protein and inhibited the expression of Bax, cytochrome C and caspace-3 protein. Conclusion Emulsified iscflurane preconditioning can protect the brain from focal cerebral I/R by increasing Bcl-2 protein expression and decreasing Bax protein expression, and reducing cytochrome C release from mitochondria, caspase-3 activation and neuronal apoptosis.