醒神通督针刺对脑缺血再灌注损伤大鼠自噬的作用研究  

Effect of Xingshen Tongdu Acupuncture on Autophagy in Rats with Cerebral Ischemia-Reperfusion Injury

作　　者：兰瑞[1] 邹旭欢 马云枝[1] 付雪琴 王漫漫[2] 王玮玮[2] 张勇[3] LAN Rui;ZOU Xuhuan;MA Yunzhi;FU Xueqin;WANG Manman;WANG Weiwei;ZHANG Yong(The First Af liated Hospital of Henan University of Chinese Medicine,Zhengzhou 450000,China;Henan University of Chinese Medicine,Zhengzhou 450000,China;The Third Af liated Hospital of Zhengzhou University,Zhengzhou 450000,China)

机构地区：[1]河南中医药大学第一附属医院,郑州450000 [2]河南中医药大学,郑州450000 [3]郑州大学第三附属医院,郑州450000

出　　处：《中国中医基础医学杂志》2024年第4期633-638,共6页JOURNAL OF BASIC CHINESE MEDICINE

基　　金：国家自然科学基金项目(81973618,81503422);河南省自然科学基金项目(202300410399)。

摘　　要：目的从自噬探讨醒神通督针刺抗脑缺血再灌注损伤的作用机制。方法雄性SD大鼠随机分为假手术组、脑缺血再灌注模型组(简称模型组)和脑缺血再灌注+醒神通督针刺组(简称针刺组)。采用大脑中动脉线栓法构建脑缺血再灌注损伤模型,针刺组大鼠造模前5 d开始干预,取穴“百会”“四神聪”“足三里”,每日针刺1次,直至再灌注24 h。另外2组大鼠只抓取不针刺。再灌注24 h后采用氯化三苯基四氮唑(triphenyltetrazolium chloride,TTC)染色观察脑梗死面积,透射电镜观察神经元及自噬的亚显微结构,Western blot、免疫荧光检测缺血皮层微管相关蛋白1轻链3(microtubule-associated protein light chain 3B,LC3)、肌球蛋白样BCL2结合蛋白(coiled coil moesin like BCL2 interacting protein,Beclin-1)、自噬相关蛋白(autophagy related protein,Atg)12、蛋白激酶B(protein kinase B,PKB,又称Akt)与磷酸化Akt(phosphorylated Akt,p-Akt)的表达。结果与假手术组比较,模型组大鼠大脑中动脉供血区梗死面积明显增加(P<0.05),且缺血皮层神经元结构严重破坏,自噬明显激活,自噬蛋白Beclin-1、Atg12表达及LC3Ⅱ/I比值显著升高(P<0.05),p-Akt表达下降(P<0.05)。与模型组比较,针刺组大鼠脑梗死面积减少(P<0.05),透射电镜显示神经元结构较为正常,自噬体较少。此外,针刺组自噬蛋白Beclin-1、Atg12表达及LC3Ⅱ/I比值明显降低(P<0.05),p-Akt表达显著上调(P<0.05)。结论针刺具有抗脑缺血再灌注损伤作用,可能通过抑制自噬的活化实现。Objective To investigate the effects of Xingshen Tongdu acupuncture on rats after cerebral ischemia and reperfusion based on autophagy.Methods Male SD rats were randomly divided into 3 groups:sham control(Sham),ischemia and reperfusion(IR),IR plus Xingshen Tongdu acupuncture(Acu).The model was induced by middle cerebral artery occlusion(MCAO)followed by reperfusion.The acupuncture intervention was started 5 days before modeling to the rats of the Acu group,and the acupoints“Baihui”“Sishencong”and“Zusanli”were selected for treatment,once a day until 24 h after reperfusion.Rats in the other 2 groups were only grabbed without acupuncture.After 24 h of reperfusion,cerebral infarct areas of rats were evaluated by triphenyltetrazolium chloride(TTC)staining in all groups.Ultrastructural morphologies of autophagy were observed by transmission electron microscope(TEM).Levels of microtubule-associated protein light chain 3B(LC3),coiled coil moesin like BCL2 interacting protein(Beclin-1),autophagy related protein(Atg)12,protein kinase B(Akt)and p-Akt were detected by Western blot and immunofluorescent staining.Results Compared with the Sham group,cerebral infarct areas of MCA territory were increased(P<0.05),and TEM indicated that neuronal autophagy was significantly activated in the rats of the IR group.Moreover,Beclin-1,Atg12 expression levels,and the ratio of LC3Ⅱ/I were significantly up-regulated(P<0.05),while the p-Akt level was decreased(P<0.05)in the rats of the IR group.Compared with the IR group,the cerebral infarct area decreased(P<0.05),while TEM revealed a relatively normal neuronal structure with fewer autophagosomes in the rats of Acu group.Furthermore,the expression of Beclin-1 and Atg12 as well as LC3Ⅱ/I ratio decreased obviously(P<0.05),while the expression of p-Akt protein was significantly up-regulated(P<0.05)in the rats of Acu group.Conclusion The protective effects of acupuncture on cerebral ischemia and reperfusion injury may be related to inhibiting autophagy.

关 键 词：醒神通督 针刺 脑缺血再灌注损伤 自噬 

分 类 号：R245[医药卫生—针灸推拿学]