电针“水沟”“百会”穴对脑缺血再灌注损伤大鼠海马神经元自噬的影响

Effect of electroacupuncture at“Shuigou”(GV26)and“Baihui”(GV20)on autophagy of hippocampal neurons in rats with cerebral ischemia-reperfusion injury

目的:探讨电针“水沟”“百会”穴对脑缺血再灌注(I/R)损伤大鼠海马神经元自噬的影响。方法:SD大鼠随机分为假手术组、模型组、电针组,每组16只。采用一侧大脑中动脉栓塞(MCAO)的方法复制I/R损伤大鼠模型。电针组予“水沟”“百会”电针治疗,20min/次,1次/d,共5d。以Longa神经功能评分法评价神经功能;TTC染色法测定大鼠脑梗死体积百分比;ELISA法检测脑脊液白细胞介素(IL)-6、IL-18、肿瘤坏死因子-α(TNF-α)含量;荧光定量PCR和Western blot法分别检测缺血区海马组织自噬相关蛋白腺苷酸活化蛋白激酶(AMPK)、Beclin-1、VPS34、LC3BmRNA和蛋白的表达。结果:与假手术组比较,模型组神经功能评分明显升高(P<0.01),大鼠脑梗死体积百分比明显增大(P<0.01),脑脊液IL-6、IL-18、TNF-α含量明显升高(P<0.01,P<0.05),海马组织AMPK、Beclin-1、VPS34蛋白和mRNA表达水平,以及LC3B mRNA表达水平和LC3B-Ⅱ/LC3B-Ⅰ比值均升高(P<0.01,P<0.05)。与模型组比较,电针组神经功能评分降低(P<0.05),脑梗死体积百分比缩小(P<0.05),脑脊液IL-6、IL-18、TNF-α含量降低(P<0.05),海马组织AMPK、Beclin-1、VPS34蛋白和mRNA表达水平,以及LC3BmRNA表达水平和LC3B-Ⅱ/LC3B-Ⅰ比值均降低(P<0.01,P<0.05)。结论:电针治疗可能通过抑制海马神经元自噬水平,缓解MCAO大鼠I/R损伤,减轻神经受损程度。

Objective To explore the effect of electroacupuncture(EA)at“Shuigou”(GV6)and“Baihui”(GV20)on autophagy of hippocampal neurons in cerebral ischemia-reperfusion(I/R)injury rats.Methods Forty-eight healthy male SD rats were randomly divided into sham operation,model and EA groups,with 16rats in each group.The rat model of cerebral I/R injury was established by occlusion of the middle cerebral artery(MCAO).Rats of the EA group received EA at GV26and GV20for 20min,once daily for 5days.The neurological function of rats in each group was evaluated by Longa neurological function score.The cerebral infarction volume was measured by TTC staining.The levels of IL-6,IL-18and TNF-αin cerebrospinal fluid were detected by ELISA.Real-time PCR and Western blot were respectively used to detect the expressions of autophagy-related proteins AMPK,Beclin-1,VPS34and LC3B.Results Compared with the sham operation group,neurological function scores of rats in the model group were significantly increased(P<0.01);the volume of cerebral infarction was significantly increased(P<0.01);the contents of IL-6,IL-18and TNF-αin cerebrospinal fluid were increased(P<0.01,P<0.05);the mRNA expression levels of AMPK,Beclin-1,VPS34and LC3Bwere significantly increased(P<0.01);the protein expressions of AMPK,Beclin-1,VPS34and the ratio of LC3B-Ⅱ/LC3B-Ⅰwere increased(P<0.01,P<0.05).After intervention and in comparison with the model group,the neurological function scores were decreased(P<0.05);the cerebral infarct volume were decreased(P<0.05);the contents of IL-6,IL-18and TNF-αin cerebrospinal fluid were decreased(P<0.05);the mRNA expressions of AMPK,Beclin-1,VPS34and LC3Bwere significantly decreased(P<0.01);the protein expressions of AMPK,Beclin-1,VPS34and the ratio of LC3B-Ⅱ/LC3B-Ⅰwere decreased(P<0.05,P<0.01).Conclusion EA can improve the neurological function and alleviate the degree of nerve injury in rats with cerebral I/R injury,which may be related to inhibiting the autophagy level of hippocampal neurons.