乳化异氟醚预处理对大鼠局灶性脑缺血再灌注时海马CA1区神经元凋亡的影响

Effect of emulsified isoflurane preconditioning on neuronal apoptosis in hippocampal CA1 region induced by focal cerebral ischemia-reperfusion injury in rats

目的 评价乳化异氟醚预处理对大鼠局灶性脑缺血再灌注时海马CA1区神经元凋亡的影响。方法健康成年雄性SD大鼠48只，体重250～300g，月龄4—6月，采用随机数字表法，将大鼠随机分为6组（n=8），假手术组（s组）腹腔注射生理盐水10．5ml／kg，24h后只分离血管，不置入线栓；缺血再灌注组（I／R组）和乳化异氟醚预处理组（EI组）分别腹腔注射生理盐水或8％乳化异氟醚10．5ml／kg（120mg／ml）；LY294002＋乳化异氟醚预处理组（L＋EI组）缺血侧侧脑室注射LY294002（磷脂酰肌醇-3激酶特异性抑制剂）25mmol／L5μl，30min后腹腔注射8％乳化异氟醚10．5ml／kg；LY294002组（L组）和DMSO（LY294002溶剂）组缺血侧侧脑室注射LY29400225mmol／L（5阻）或DMSO5m。于给药后24h采用大脑中动脉缺血2h恢复再灌注的方法制备局灶性脑缺血再灌注模型。于再灌注24h时行神经功能缺陷评分，检测海马CA1区神经元凋亡情况和磷酸化丝氨酸．苏氨酸蛋白激酶（p-Akt）表达，观察海马CA1区病理学改变。结果与s组比较，其余各组神经功能缺陷评分、凋亡细胞计数升高，p-Akt表达上调（P〈0．05）；与I／R组比较，EI组神经功能缺陷评分、凋亡细胞计数降低，p-Akt表达上调（P〈0．05），L＋EI组、L组、DMSO组上述各指标差异无统计学意义（P〉0．05）；与EI组比较，L＋EI组神经功能缺陷评分、凋亡细胞计数升高，p-Akt表达下调（P〈0．05）。EI组病理学损伤程度明显轻于I／R组，L＋EI组、L组、DMSO组与I／R组相似。结论乳化异氟醚预处理可减少大鼠局灶性脑缺血再灌注时海马CA1区神经元凋亡，其神经元保护作用与P13K／Akt通路激活有关。

Objective To investigate the effect of preconditioning with emulsified isoflurane （elSO） on neuronal apoptosis in hippocampal CA1 region induced by focal cerebral isehemia-reperfusion （I/R） injury in rats. Methods Forty-eight healthy adult male SD rats weighing 250-300 g were randomly divided into 6 groups （ n = 8 each） ： sham operation group （group S） ; I/R group; eISO ＋ I/R group （group EI） ; LY294002 （a specific PI3K inhibitor） ＋ eISO ＋ I/R group （group L ＋ EI）; LY294002 ＋ I/R group（ group L） and DMSO （solvent for LY294002） ＋ I/R group（group DMSO）. Focal cerebral I/R was induced by 2 h midd/e cerebra/artery occlusion （MCAO）. A nylon thread （0.26 mm in diameter） with rounded tip was inserted into internal carotid artery and advanced cranially until resistance was met （depth of insertion about 18-20 mm）. eISO 10.5 ml/kg （ 120 mg/ml） was injected intraperitoneally （IP） in groups EI and L ＋ EI. LY294002 （25 mmol/L） 5 /d was injected into cerebral ventricle on the ischemic side in group L＋ EI （at 30 min before eISO） and group L. DMSO 5μl was injected into the cerebral ventricle on ischemic side before MCAO in group DMSO. Neurologic deficit was assessed and scored （0 = normal, 4 = unconscious） at 24 h of reperfusion. The animals were then killed and their brains were removed for detection of neuronal apoptosis （by TUNEI） and p-Akt expression （by immuno-histochemistry） in hippocampal CA1 region. Results Cerebral I/R significanlly increased the neurologic deficit scores, the number of apoptotic cells and p-Akt expression in group I/R as compared with group S. Preconditioning with eISO attenuated the I/R- induced increase in neurologic deficit scores and number of apoptotic cells but further increased p-Akt expression. The neuroprotective effect of eISO preconditioaing against I/R-induced changes was counteracted by LY294002. Conclusion eISO preconditioning can attenue.te focal cerebral I/R-induced neuronal apoptosis in rats by activating P13K/Akt pathway.