甲基苯丙胺与HIV-Tat蛋白协同改变大鼠血脑屏障通透性的氧化应激作用机制

Mechanisms of Oxidative Stress in the Change of Blood-Brain Barrier Permeability by Methamphetamine and HIV-Tat Protein

目的观察甲基苯丙胺与HIV-Tat蛋白协同对大鼠血脑屏障的作用机制.方法雄性SD大鼠每天2次腹腔注射给予MA10 mg/kg的同时尾静脉注射给予HIV-Tat 400 ng/kg,连续7 d,给药结束24 h后随机取10只大鼠尾静脉注射伊文思蓝检测脑组织EB含量,随机取5只大鼠断颈取脑,用于SOD活力、GSH和MDA含量的测定.余下2只大鼠快速断颈取脑,戊二醛-锇酸溶液固定前额叶皮质部分,透射电镜观察结构变化.结果与正常对照组相比,实验组脑组织中EB含量不同程度增加,提示实验组血脑屏障通透性增加(P<0.05);MDA含量升高、SOD活力和GSH含量不同程度降低,提示实验组氧化应激反应增强(P<0.05).MA+Tat组与MA组、Tat组相比,EB含量升高明显,提示MA与HIV-Tat蛋白联用能协同增加血脑屏障通透性;MDA含量显著升高,SOD活力、GSH含量明显下降,提示MA与HIV-Tat蛋白联用能协同增强大鼠脑组织氧化应激反应(P<0.01);NAC+MA+Tat组与MA+Tat组相比,EB含量降低,提示NAC能一定程度拮抗MA与HIV-Tat蛋白对血脑屏障通透性的影响(P<0.01);MDA含量下降,SOD活力、GSH含量明显升高,反映NAC能在一定程度拮抗MA与HIV-Tat蛋白对大鼠脑内氧化应激反应增强作用(P<0.01).各实验组在电镜下观察到血脑屏障一系列超微结构改变,如脑微血管内皮细胞肿胀变薄,血管周围胶质细胞和星形胶质细胞肿胀,胞饮小泡增加等,这些改变以MA+Tat组最为显著.结论 MA和HIV-Tat蛋白能改变血脑屏障通透性,两者具有协同作用,协同作用机制可能与氧化应激有关.

Objective To investigate the changes of the blood-brain barrier and investigate the effects induced synergistically by MA and Tat protein. Methods Male SD rats were ip given MA 10 mg/kg twice daily followed by tail intravenous injection given Tat protein 0.4 μg/kg once daily for 7 days. After the last injection,we randomly chose 10 rats and given tail intravenous injection of EB, then we detected the EB content in brain tissues, chose 5 rats to test the ROS in brain tissues, the rest rats were prepared for transmission electronmicroscopy（TEM） observation. Results Compared with control group, EB contents in brain tissues of rats in the test group were significantly increased, suggesting blood-brain barrier permeability was increased（P〈 0.05）.Compared with control group,MDA in brain tissues of rats in the test group were significantly increased, while SOD and GSH activities were significantly decreased, suggesting oxidative stress was enhanced（P〈0.05）. Compared with MA and Tat group,EB and MDA in the MA＋Tat group were higher,suggesting that MA combined with Tat had synergistic effects on changing permeability of blood-brain barrier and the level of oxidate stress. EB and MDA in NAC＋MA＋Tat group were lower than MA＋Tat group（P 〈0.01）, whereas, SOD and GSH were higher than MA＋Tat group,suggesting NAC could protect BBB against injury induced by MA and Tat（P〈0.01）. And a series of ultrastructural changes of blood-brain barrier were found under transmission electron-microscopy, such as cerebral microvascular endothelial cells became swollen and thinner, the glia and astrocytes also were swollen.Conclusion MA and Tat protein have synergistic effects on changing the permeability of blood-brain barrier. In addition, the effects may attribute to oxidative stress.