白藜芦醇调节HMGB1/TLR4/NF-κB信号通路对急性脑梗死大鼠神经元损伤的影响

Effect of Resveratrol on Neuronal Damage in Rats with Acute Cerebral Infarction by Regulating the HMGB1/TLR4/NF-κB Signaling Pathway

目的探讨白藜芦醇调节高迁移率族蛋白B1(HMGB1)/Toll样受体4(TLR4)/核因子κB(NF-κB)信号通路对急性脑梗死(ACI)大鼠神经元损伤的影响。方法大鼠随机分为空白组、梗死组、白藜芦醇组、HMGB1激活剂(重组大鼠HMGB1蛋白,rRHMGB1)组、白藜芦醇+rRHMGB1组,每组18只。除空白组外,其它组大鼠均采用大脑中动脉闭塞法构建急性脑梗死模型,建模成功后1 h开始给药处理,每日给药1次,持续7 d。检测神经功能损伤评分、脑梗死体积比率变化;ELISA法检测梗死区脑组织中肿瘤坏死因子α(TNF-α)、白细胞介素(IL)-1β和IL-6水平;尼氏染色检测梗死区脑组织存活的神经元数量;TUNEL染色检测梗死区脑组织的神经元凋亡;Western Blot法检测脑组织中Bcl2相关X蛋白(Bax)、TLR4、裂解的天冬氨酸特异性半胱氨酸蛋白酶3(Cleaved Caspase-3)、HMGB1、p-NF-κB p65蛋白。结果与空白组比较,梗死组神经功能损伤评分、脑梗死体积比率,梗死区脑组织中TNF-α、IL-1β、IL-6水平,神经元凋亡率及Bax、TLR4、Cleaved Caspase-3、HMGB1、p-NF-κB p65蛋白均升高(P<0.05);梗死区脑组织中存活的神经元数量降低(P<0.05)。与梗死组比较,白藜芦醇组及白藜芦醇+rRHMGB1组神经功能损伤评分,脑梗死体积比率,梗死区脑组织中TNF-α、IL-1β、IL-6水平,神经元凋亡率及Bax、TLR4、Cleaved Caspase-3、HMGB1、p-NF-κB p65蛋白均降低(P<0.05);梗死区脑组织中存活的神经元数量升高(P<0.05)。rRHMGB1组上述对应指标变化趋势均比梗死组升高(P<0.05)。与白藜芦醇组比较,白藜芦醇+rRHMGB1组神经功能损伤评分,脑梗死体积比率,梗死区脑组织中TNF-α、IL-1β、IL-6水平,神经元凋亡率及Bax、TLR4、Cleaved Caspase-3、HMGB1、p-NF-κB p65蛋白均升高(P<0.05);梗死区脑组织中存活的神经元数量降低(P<0.05)。结论白藜芦醇改善急性脑梗死大鼠神经元损伤的机制可能与抑制HMGB1/TLR4/NF-κB信号通路有关。

Objective This study aims to investigate the effect of resveratrol on neuronal damage in rats with acute cerebral infarction(ACI)by regulating the high-mobility group protein B1(HMGB1)/Toll-like receptor 4(TLR4)/nuclear factor-κB(NF-κB)signaling pathway.Methods Rats were randomly separated into blank group,infarction group,resveratrol group,HMGB1 activator(recombinant rat HMGB1 protein,rRHMGB1)group,and resveratrol+rRHMGB1 group,with 18 rats in each group.Except for the blank group,rats in all other groups were used to construct ACI models using the method of middle cerebral artery occlusion.Drug treatment began at 1 hour after successful modeling,once a day,for 7 days.Changes in neurological impairment score and cerebral infarction volume coefficient were detected.ELISA was applied to detect levels of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),and IL-6 in infarction region of brain tissue.Nissl staining was applied to detect the number of surviving neurons in infarction region of brain tissues.TUNEL staining was applied to detect neuronal apoptosis in infarction region of brain tissue.Western Blot was applied to detect Bcl2 associated X protein(Bax),TLR4,Cleaved Caspase-3,HMGB1,and p-NF-κB p65 protein in brain tissue.Results Compared with the blank group,the neurological damage score,cerebral infarction volume coefficient,levels of TNF-α,IL-1β,IL-6 in the infarction region of brain tissue,neuronal apoptosis rate,Bax,TLR4,Cleaved Caspase-3,HMGB1,p-NF-κB p65 protein increased in the infarction group,while the number of surviving neurons in the infarction region of brain tissue decreased(P<0.05).Compared with the infarction group,the neurological damage score,cerebral infarction volume coefficient,levels of TNF-α,IL-1β,IL-6 in the infarction region of brain tissue,neuronal apoptosis rate,and protein expression of Bax,TLR4,Cleaved Caspase-3,HMGB1,p-NF-κB p65 decreased in resveratrol group and resveratrol+rRHMGB1 group,the number of surviving neurons in the infarction region of brain tissue increased(P<0.05).Change of the corresponding indicators in the rRHMGB1 group was opposite to that in the resveratrol group(P<0.05).Compared with the resveratrol group,the neurological damage score,cerebral infarction volume coefficient,levels of TNF-α,IL-1β,IL-6 in the infarction region of brain tissue,neuronal apoptosis rate,the protein expression of Bax,TLR4,Cleaved Caspase-3,HMGB1,p-NF-κB p65 increased in the resveratrol+rRHMGB1 group,the number of surviving neurons in the infarction region of brain tissue decreased(P<0.05).Conclusion The mechanism by which resveratrol improves neuronal damage in ACI rats may be related to the inhibition of the HMGB1/TLR4/NF-κB signaling pathway.