雷公藤红素后处理对大鼠局灶性脑缺血再灌注损伤后脑组织NF-κB及TNF-α、IL-1β的影响

Effect of celastrol postconditioning on NF-κB，TNF-α，IL-1β in brain after focal cerebral ischemia-reperfusion injury in rats

摘要:

目的：探讨雷公藤红素后处理对大鼠局灶性脑缺血再灌注（ischemia-reperfusion，I/R）损伤的影响及其作用机制。方法：健康成年SD大鼠64只，雌雄各半，随机分为4组（n=16）：假手术组（S组）、雷公藤红素对照组（S+C组）、局灶性脑缺血再灌注组（I/R组）、雷公藤红素后处理组（I/R+C组）。线栓法制作大鼠大脑中动脉闭塞（middle cerebral artery occlusion，MCAO）2 h-再灌注24 h模型。S组在假手术后经腹腔注射二甲基亚砜（dimethyl sulfoxide，DMSO）0.3 ml/kg，S+C组在假手术后经腹腔注射雷公藤红素3 mg/kg，I/R组在再灌注5 min经腹腔注射DMSO 0.3 ml/kg，I/R+C组在再灌注5 min经腹腔注射雷公藤红素3 mg/kg。在再灌注前5 min和再灌注后24 h进行神经功能缺失评分；在再灌注后24 h进行氯化三苯四唑氮（2，3，5-triphenyl tetrazolium chloride，TTC）染色观察脑梗死体积及梗死体积百分比，HE染色观察缺血侧海马CA1区病理改变，ELISA方法测定缺血侧脑组织肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）及白细胞介素-1β（interleukin-1β，IL-1β）含量，Western blot方法测定缺血侧脑组织核因子-κB （nuclear factor-κB，NF-κB）p65及其抑制蛋白（inhibitor of κB，IκB）α蛋白表达水平。结果：与S组、S+C组分别比较，I/R组和I/R+C组神经功能缺失评分、脑梗死体积及梗死体积百分比、TNF-α及IL-1β含量、NF-κB p65蛋白表达水平均升高，IκBα蛋白表达水平降低（P<0.01），缺血侧海马CA1区有明显病理损伤。与I/R组比较，I/R+C组神经功能缺失评分、脑梗死体积及梗死体积百分比、TNF-α及IL-1β含量、NF-κB p65蛋白表达水平均降低，IκBα蛋白表达水平升高（P<0.01），缺血侧海马CA1区病理损伤较轻。S组与S+C组比较差异无统计学意义（P >0.05）。结论：雷公藤红素后处理可以减轻大鼠局灶性脑I/R损伤，其作用机制可能与抑制NF-κB的活化，减少TNF-α及IL-1β产生，从而减轻炎症反应有关。

Abstract:

Objective：To investigate the effects and the mechanism of celastrol postconditioning against focal cerebral ischemia-reper－fusion（I/R） injury in rats. Methods：Sixty-four SD rats with half males and half females were randomly divided into 4 groups（n=16）：sham operation group（group S），celastrol control group（group S+C），focal cerebral I/R group（group I/R），celastrol postconditioning group（group I/R+C）. Focal cerebral I/R models were produced by occlusion of middle cerebral artery for 2 h followed by 24 h of reper－fusion. Dimethyl sulfoxide（DMSO） 0.3 ml/kg was injected intraperitoneally after the sham operation in group S；celastrol 3 mg/kg was injected intraperitoneally after the sham operation in group S+C；DMSO 0.3 ml/kg was injected intraperitoneally at 5 min after the reperfusion in group I/R and celastrol 3 mg/kg was injected intraperitoneally at 5 min after the reperfusion in group I/R+C. The neu－rologic deficit scores were measured at 5 min before the reperfusion and at 24 h after the reperfusion. The infarct volume and the in－farct volume fraction were detected by TTC staining. The pathological changes in CA1 region of ischemic hippocampus were detected by HE staining. The content of tumor necrosis factor-α（TNF-α） and interleukin-1β（IL-1β） in ischemic brain were detected by ELISA. The protein expression of nuclear factor-κB（NF-κB） p65 and inhibitor of κB（IκB）α in ischemic brain were detect－ed by Western blot. Results：The neurologic deficit scores，the infarct volume and the infarct volume fraction，the content of TNF-α and IL-1β，the protein expression of NF-κB p65 were all significantly higher in group I/R and I/R+C than in group S and S+C respectively. The protein expression of IκBα was significantly lower in group I/R and I/R+C than in group S and S+C respectively（P<0.01）. The neurologic deficit scores，the infarct volume and the infarct volume fraction，the content of TNF-α and IL-1β，the protein expression of NF-κB p65 were all significantly lower in group I/R+C than in group I/R and the protein expression of IκBα was significantly higher in group I/R+C than in group I/R（P<0.01）. The pathological changes in CA1 region of ischemic hippocampus were significantly attenuated in group I/R+C than in group I/R. The dif－ference between group S and group S+C was not statistical significant（P >0.05）. Conclusion：Postconditioning with celastrol attenuates focal cerebral I/R injury in rats；which might be related to inhibit the activaty of NF-κB，reduce the production of TNF-α and IL-1β，attenuate the inflammatory response in brain.