七氟烷预处理对局灶性脑缺血再灌注损伤大鼠线粒体通透性转换孔的影响

目的 探讨七氟烷预处理对局灶性脑缺血再灌注损伤大鼠线粒体通透性转换孔(mPTP)的影响.方法 成年雄性SD大鼠60只,体重250～300 g,随机分为5组(n=12):假手术组(S组)、缺血再灌注组(I/R组)、七氟烷预处理组(Sev组)、线粒体ATP敏感性钾离子通道(mito-KATP通道)阻断剂5-羟癸酸(5-HD)+Sev组和5-HD组.采用大脑中动脉阻断法制备局灶性脑缺血再灌注模型.S组只分离血管不置入线栓;I/R组制备局灶性脑缺血再灌注模型;Sev组吸入2.4%七氟烷60 min行预处理,24 h后制备局灶性脑缺血再灌注模型;5-HD+Sev组腹腔注射5-HD 40mg/kg,30 min后行七氟醚预处理,其余处理同Sev组;5-HD组腹腔注射5-HD 40 mg/kg,30 min后制备局灶性脑缺血再灌注模型.于再灌注24 h时断头取缺血侧顶叶皮层组织,测定mPTP活性,Western blot法测定Bcl-2、Bax表达水平,并计算Bcl-2/Bax比值,采用TUNEL法检测神经元凋亡情况.结果 与S组比较,I/R组、Sev组、5-HD+Sev组和5-HD组凋亡神经元计数升高,Bcl-2和Bax表达上调,Bcl-2/Bax比值升高,mPTP活性升高(P＜0.05);与I/R组比较,Sev组凋亡神经元计数减少,Bcl-2表达上调,Bcl-2/Bax比值升高,mPTP活性降低(P＜0.05);与Sev组比较,5-HD+Sev组和5-HD组Bcl-2表达下凋,Bcl-2/Bax比值降低,mPTP活性升高(P＜0.05);5-HD+Sev组与5-HD组上述指标比较差异无统计学意义(P＞0.05).结论 七氟烷预处理可能通过激活神经元mito-KATP通道,上调Bcl-2的表达,从而抑制mPTP的大量开放减轻大鼠局灶性脑缺血再灌注时的神经元凋亡.

Effect of sevoflurane preconditioning on mitochondrial permeability transition pore following focal cerebral ischemia-reperfusion injury in rats

Objective To investigate the effect of sevoflurane preconditioning on mitochondrial permeability transition pore (mPTP) following focal cerebral ischemia-reperfusion (I/R) injury. Methods Male adult SD rats weighing 250-280 g were randomly assigned into 5 groups ( n = 12 each): group Ⅰ sham operation(group S); group Ⅱ focal cerebral ischemia-reperfusion (group I/R); group Ⅲ sevoflurane preconditioning + I/R(group Sev); group Ⅳ 5-HD + Sev + I/R and group Ⅴ 5-HD + I/R. Focal cerebral I/R was induced by right middle cerebral artery occlusion (MCAO). A 50 mm long 4.0 nylon thread was inserted into right internal carotid artery and advanced cephalad until resistance was met. MCAO was maintained for 2 h followed by 24 h reperfusion in group Ⅱ-Ⅴ. In group Ⅲ the animals inhaled 2.4% sevoflurane for 60 min at 24 h before I/R. In group Ⅳ 5-HD 40 mg/kg was injected intraperitoneally at 30 min before sevoflurane preconditioning. In group Ⅴ 5-HD 40 mg/kg was injected IP at 30 min before MCAO. The animals were decapitated at 24 h of reperfusion. The parietal cortex of the ischemic side was isolated. Spectrophotometry was used to measure the mPTP activity. The expression of Bcl-2 and Bax protein (by Western blotting) and neuronal apoptosis (by TUNEL) were determined.Results Sevoflurane preconditioning significantly inhibited the activity of mPTP, reduced neuronal apoptosis induced by I/R and increased Bcl-2 protein expression but had no significant effect on Bax protein expression. The protective effects of sevoflurane preconditioning against cerebral I/R were blocked by 5-HD. Conclusion Sevoflurane could inhibit the mitochondrial permeability transition and reduce neuronal apoptosis via activation of mito-KATP channel and up-regulation of Bcl-2 expression.