艾司洛尔预先干预对脑缺血再灌注损伤大鼠Toll样受体4/核因子-κB信号通路的影响

Effect of esmolol pretreatment on Toll like receptor-4/nuclear factor-kappa-B pathway in rats with repeated cerebral ischemia/reperfusion injury

目的探讨艾司洛尔预先干预对脑缺血再灌注损伤大鼠Toll样受体4（TLR4）/核因子-κB（NF-κB）信号通路的影响。方法将48只健康成年雄性SD大鼠按随机数字表法分为假手术组、脑缺血再灌注损伤组和艾司洛尔组，每组16只。后2组采用双侧颈总动脉夹闭术制备脑缺血再灌注损伤模型，艾司洛尔组和脑缺血再灌注损伤组分别于术前经静脉泵注艾司洛尔200 g/（kg·min）或等容量生理盐水1 h。假手术组只分离双侧颈总动脉而不夹闭，并于分离双侧颈总动脉后输注等容量生理盐水1 h。采用Morris水迷宫实验测试各组大鼠术前及术后1 d、3 d、7 d时的学习记忆能力，处死大鼠后取海马组织并测定其湿干重比，采用伊文斯蓝（EB）法检测脑组织血脑屏障通透性，采用酶联免疫吸附试验（ELISA）检测海马组织中肿瘤坏死因子-α（TNF-α）、白介素（IL）-6和IL-1β水平，采用逆转录-聚合酶链式反应（RT-PCR）检测海马组织中NF-κB p65、TLR4 mRNA表达，采用Western blotting检测海马组织中NF-κB p65、TLR4蛋白表达。结果与假手术组比较，脑缺血再灌注损伤组术后1 d、3 d及7 d时的逃避潜伏期及游泳距离均明显延长，海马组织湿干重比和脑组织EB含量均明显升高，海马组织中TNF-α、IL-6、IL-1β水平均明显升高，海马组织中NF-κB p65、TLR4 mRNA及蛋白表达均明显升高，差异均有统计学意义（P〈0.05）。与脑缺血再灌注损伤组比较，艾司洛尔组术后1 d、3 d及7 d时的逃避潜伏期及游泳距离均明显缩短，海马组织湿/干重比和脑组织EB含量均明显降低，海马组织中TNF-α、IL-6、IL-1β水平均明显降低，海马组织中NF-κB p65、TLR4 mRNA及蛋白表达均明显降低，差异均有统计学意义（P〈0.05）。结论艾司洛尔预先干预可减轻大鼠脑缺血再灌注损伤并改善其学习记忆能力，其机制与艾司洛尔减轻海马炎症反应、抑制海马TLR4/NF-κB信号通路表达有关。

ObjectiveTo investigate the effect of esmolol pretreatment on Toll like receptor-4 （TLR4）/nuclear factor-kappa-B （NF-κB） pathway in rats with repeated cerebral ischemia/reperfusion （IR） injury.MethodsForty-eight adult male Sprague-Dawley rats were randomly allocated into sham-operated group, IR group and esmolol group （n=16）. Rats in the IR group and esmolol group were injected intravenously with esmolol at a dose of 200 g/（kg·min） or normal saline for one h before surgery, and then, bilateral common carotid arteries were clipped to establish the repeated IR injury models. Bilateral common carotid arteries in rats of sham-operated group were only isolated but not clipped, and injected intravenously with normovolemic normal saline for one h. Learning and memory abilities of rats were measured by Morris water maze test before, and one, 3 and 7 d after surgery. Rats were euthanized and hippocampus tissues were excised. The wet to dry （W/D） ratio of the hippocampus was tested. The permeability of blood-brain barrier was determined by Evans blue （EB） method. The levels of tumor necrosis factor-alpha （TNF-α）, interleukin （IL）-6 and IL-1β in the hippocampus were tested by enzyme linked immunosorbent assay （ELISA）. The NF-κB p65 and TLR4 mRNA expressions in the hippocampus were determined by reverse transcription-polymerase chain reaction （RT-PCR）. The NF-κB p65 and TLR4 protein expressions in hippocampus were detected by Western blotting.ResultsAs compared with those in the sham-operated group, the escape latency was significantly prolonged and swimming distance was signficantly longer in rats of IR group one, 3 and 7 d after surgery （P〈0.05）, the W/D ratio of the hippocampus and the content of EB in brain tissues were significantly increased in the IR group （P〈0.05）, the levels of TNF-α, IL-6 and IL-1β in hippocampus were significantly increased in the IR group （P〈0.05）, and the NF-κB p65 or TLR4 mRNA and protein expressions in the hippocampus of IR group were statistically higher （P〈0.05）. As compared with IR group, the escape latency and swimming distance of rats in the esmolol group were significantly shortened one, 3 and 7 d after surgery （P〈0.05）, the W/D ratio of the hippocampus and content of EB in brain tissues of esmolol group were significantly decreased （P〈0.05）, the levels of TNF-α, IL-6 and IL-1β in the hippocampus of esmolol group were signficantly lower（P〈0.05）, and the NF-κB p65 or TLR4 mRNA and protein expressions in hippocampus of esmolol group were statistically lower in the esmolol group （P〈0.05）.ConclusionEsmolol preconditioning can alleviate cerebral injury and improve learning and memory abilities of rats with repeated cerebral IR injury, which may be involved in alleviating inflammation and suppressing TLR4/NF-κB pathway.