AMPK介导线粒体融合与裂变在抑制P2X7受体抗神经元缺氧/复氧损伤中的作用  

Role of AMPK-mediated mitochondrial fusion and fission in inhibiting P2X7 receptor anti-hypoxia/reoxygenation injury of neurons

作　　者：王雪如 门运政 胡淼 洪玉洁 刘心怡 董淑英 WANG Xue-ru;MEN Yun-zheng;HU Miao;HONG Yu-jie;LIU Xin-yi;DONG Shu-ying(School of Pharmacy,Bengbu Medical College,Bengbu Anhui 233030,China)

机构地区：[1]蚌埠医学院药学院,安徽蚌埠233030

出　　处：《蚌埠医学院学报》2020年第6期714-719,共6页Journal of Bengbu Medical College

基　　金：国家自然科学基金项目(81402930,81673791);蚌埠医学院研究生创新科学研究项目(Byycx1813);蚌埠医学院大学生创新科研项目(201810367055)。

摘　　要：目的:探讨AMPK介导下线粒体融合与裂变在抑制P2X7受体抗神经元缺氧/复氧(hypoxia/reoxygenation,H/R)损伤中的作用。方法:培养原代神经元细胞,并随机分为正常对照(Control)组、缺氧/复氧(H/R)组、H/R+Bright Blue G(BBG)组、H/R+BBG+Dorsomorphin组。采用Calcein-AM/PI双染法检测细胞存活率;采用活性氧(ROS)试剂盒检测ROS的产生;采用Mito Tracker TM Green FM检测线粒体形态;采用Western blotting法检测p-AMPK、p-Drp1、Mfn2蛋白的表达。结果:与Control组相比,当神经元细胞发生H/R损伤时,其细胞死亡率、线粒体裂变、ROS的含量都明显增加,BBG预处理后,H/R损伤神经元细胞的死亡率、线粒体裂变、ROS的含量都明显减少,而AMPK抑制剂Dorsomorphin则减弱了BBG对H/R损伤神经元细胞的保护作用(P<0.01)。Western blotting结果显示,与Control组相比,H/R组神经元细胞p-AMPK与Mfn2的表达显著降低,而p-Drp1的表达显著增高(P<0.01);BBG预处理后,H/R损伤神经元细胞p-AMPK与Mfn2的表达显著增高,而p-Drp1的表达则显著降低(P<0.01);AMPK抑制剂Dorsomorphin则减弱了BBG增加H/R损伤神经元p-AMPK与Mfn2的表达而降低p-Drp1的表达的作用(P<0.01)。结论:抑制P2X7受体可通过抑制线粒体裂变促进线粒体融合减轻神经元缺氧/复氧损伤,其机制可能与激活AMPK有关。Objective:To investigate the role of mitochondrial fusion and fission regulated by AMPK in the protective effects of inhibiting P2X7 receptor against hypoxia/reoxygenation injury in neurons.Methods:The primary neurons were cultured,and randomly divided into the Control group,hypoxia/reoxygenation(H/R)group,H/R+Bright Blue G(BBG)group,H/R+BBG+Dorsomorphin group.The viability of neurons was assessed using Calcein-AM/PI kit,the ROS level was detected using ROS kit,the morphology of mitochondrial morphology was detected using Mito Tracker TM Green FM kit,and the expression levels of p-AMPK,p-Drp1 and Mfn2 were detected using Western blotting.Results:Compared with Control group,the death rate,mitochondrial fission and level of ROS in neurons increased significantly in H/R group.After pretreated with BBG,the death rate,fission of mitochondrial and level of ROS in neurons with H/R injury decreased significantly,while the protective effects of inhibitor of AMPK Dorsomorphin on neurons with H/R injury were attenuated(P<0.01).Compared with the Control group,the results of Western blotting showed that the expression levles of p-AMPK and Mfn2 decreased significantly,while the expression level of p-Drp1 increased markedly in H/R group(P<0.01).After pretreated with BBG,the expression levels of p-AMPK and Mfn2 increased significantly,while the expression level of p-Drp1 decreased markedly.The inhibitor of AMPK Dorsomorphin weakened the BBG increasing the expression levels of p-PMAK and Mfn2 in H/R injured neurons and decreas the expression level of p-Drpl(P<0.01).Conclusions:The inhibition of P2X7 receptor can reduce the damage of neurons induced by H/R injury through inhibiting mitochondrial fission to promote mitochondrial fusion,which may be involved in the activation of AMPK.

关 键 词：缺氧/复氧损伤 神经元细胞 活性氧 线粒体融合与裂变 

分 类 号：R743.3[医药卫生—神经病学与精神病学]