BMSCs移植通过激活AMPK/mTOR信号通路介导的自噬抑制大鼠蛛网膜下腔出血后神经元凋亡  

BMSCs transplantation inhibits neuronal apoptosis after subarachnoid hemorrhage in rats through activation of AMPK/mTOR signaling pathway⁃mediated autophagy

作　　者：崔彩玲 许心语 张豫 崔博文 胡艾蕙 刘俊杰 李建民 CUI Cai-ling;XU Xin-yu;ZHANG Yu;CUI Bo-wen;HU Ai-hui;LIU Jun-jie;LI Jian-min(College of Clinical Medicine,North China University of Science and Technology,Tangshan 063000,China;Department of Neurosurgery,the Affiliated Hospital of North China University of Science and Technology,Tangshan 063000,China)

机构地区：[1]华北理工大学临床医学院,河北唐山063000 [2]华北理工大学附属医院神经外科,河北唐山063000

出　　处：《海南医学院学报》2023年第21期1607-1613,共7页Journal of Hainan Medical University

基　　金：唐山市科学技术与发展计划项目(21130224C);河北省2022年度医学科学研究课题计划(20221533);大学生创新训练计划(R2022166)。

摘　　要：目的:探究大鼠蛛网膜下腔出血(SAH)后骨髓间充质干细胞(BMSCs)的干预对神经细胞凋亡和自噬的作用及其过程。方法:将48只Sprague-Dawley(SD)大鼠(雄性)随机分为:假手术(Sham)组、模型(SAH)组、模型组+骨髓间充质干细胞(BMSCs)组、模型组+BMSCs+AMPK抑制剂(Compound C)组,共4组,每组12只。除Sham组外,其他组通过造模形成具有血管内点的SAH样本,Compound C组、BMSCs组分别将Compound C和10μL生理盐水在模型制作前30 min,用大鼠大脑立体定位器经左侧脑室注入大鼠,造模成功后将2μL浓度为1×108/mL的细胞悬液于大鼠造模后1 h经脑室注入。观察大鼠脑组织水肿程度;用Garcia评分量表测试大鼠的神经功能,TUNEL染色监测大鼠海马回的神经元凋亡情况;免疫组化反映大鼠海马回LC3-Ⅱ与Beclin-1两种蛋白表达;应用Western blot检测Beclin-1,LC3-Ⅱ以及AMPK通路中的p-AMPK/AMPK、p-mTOR/mTOR等蛋白的暴露量。结果:与Sham组相比,SAH组大鼠脑水肿加重,神经元凋亡率增高,神经功能减弱,蛋白质颗粒降低(P<0.05),p-AMPK及其相关蛋白的暴露率降低(P<0.05);与SAH组相比,BMSCs组脑水肿减轻,神经元凋亡率下降,且LC3-Ⅱ、Beclin-1及p-AMPK蛋白明显增加,p-mTOR蛋白明显减少(P<0.05);与BMSCs组相比,Compound C组大鼠脑组织水肿加重,神经凋亡率上升,神经功能减弱(P<0.05),LC3-Ⅱ、Beclin-1、p-AMPK蛋白表达含量降低,p-mTOR蛋白含量升高(P<0.05)。结论:BMSCs移植可减轻SAH后的神经元凋亡;其原理可能是激活AMPK/mTOR通路,加强神经元的自噬,从而抑制其向凋亡的转化。Objective:To explore the impacts of bone marrow mesenchymal stem cells(BMSCs)intervention on the neu⁃ronal withering and autophagy in rats after subarachnoid hemorrhage(SAH)in rats and its process.Methods:Forty⁃eight male SD rats,were randomly assigned to the followings groups:Sham surgery(Sham)group,modeling(SAH)group,modeling group+bone marrow mesenchymal stem cells(BMSCs)group,the modeling group+BMSCs+AMPK inhibitor(Compound C)group,with twelve rats in each group.Except Sham group,other groups formed SAH samples with intravascular points.Com⁃pound C and 10μL of normal saline were injected into the left lateral ventricle of rats using a stereoscope brain machine 30 minutes before modeling in compound C and BMSCs groups,respectively.And 2μL of cell suspension at a concentration of 1×108/mL was injected into the ventricles of the brain 1 hour after the model was established.The severity of brain swelling in rats was ob⁃served.Garcia were used to test the neural function of rats.TUNEL staining was used to monitor neuronal apoptosis in the rat hip⁃pocampal gyrus.Immunohistochemical fluorescence reflected the expression of two proteins,LC3⁃II and Beclin⁃1,in rat hippo⁃campal gyrus.Western blotting was applied to measure the expressions of autophagy⁃associated proteins in the AMPK pathway.Results:Compared with the group undergoing sham⁃surgery,brain edema worsened in the model group;neuronal apoptosis rate was increased;neural function was weakened.Protein granules decreased(P<0.05)and expression levels of p⁃AMPK and other proteins decreased(P<0.05);brain swelling and neuronal apoptosis were reduced in the BMSCs group by comparison with the modeling group’s,with substantial elevation in the expression of proteins comprising LC3⁃Ⅱ,Beclin⁃1,and p⁃AMPK.And the standard of p⁃mTOR protein expression was considerably lessened(P<0.05);rats belonging to group compound C showed in⁃creased brain swelling which is relative to that of BMSCs group,increased neuronal apoptosis,decreased neuro

关 键 词：骨髓间充质干细胞 蛛网膜下腔出血 AMPK/mTOR 自噬 

分 类 号：R743.35[医药卫生—神经病学与精神病学]