文章摘要
陈 伟,赵 麟,刘 宁,郑 平,任大斌,段 剑,冯九庚.星形胶质细胞来源的GJA1⁃20k在氧化应激后参与神经元保护作用的机制[J].南京医科大学学报,2020,(8):1098~1104
星形胶质细胞来源的GJA1⁃20k在氧化应激后参与神经元保护作用的机制
Protective effects and mechanism of astrocytic GJA1⁃20k on neurons in oxidative stress injury
投稿时间:2019-04-13  
DOI:10.7655/NYDXBNS20200804
中文关键词: GJA1⁃20k  星形胶质细胞  神经元  线粒体  氧化应激
英文关键词: GJA1⁃20k  astrocyte  neuron  mitochondria  oxidative stress
基金项目:国家自然科学基金(81701231);上海市自然科学基金(16ZR1431500)
作者单位
陈 伟 南昌大学第一附属医院神经外科江西 南昌 330008南京医科大学第一附属医院神经外科江苏 南京 210029 
赵 麟 南京医科大学第一附属医院神经外科江苏 南京 210029 
刘 宁 南京医科大学第一附属医院神经外科江苏 南京 210029 
郑 平 上海健康医学院附属上海浦东新区人民医院神经外科上海 201299 
任大斌 上海健康医学院附属上海浦东新区人民医院神经外科上海 201299 
段 剑 南昌大学第一附属医院神经外科江西 南昌 330008 
冯九庚 南昌大学第一附属医院神经外科江西 南昌 330008 
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中文摘要:
      目的:观察星形胶质细胞(astrocyte)通过上调神经元(neuron)内源性缝隙连接蛋白α1截短单体?20k(gap junction protein alpha 1 truncated monomer? 20k,GJA1?20k)参与氧化应激损伤后神经保护作用的机制。方法:采用C57BL/6胎鼠以原代培养法获取神经元及星形胶质细胞,建立共培养模型,给予神经元过氧化氢(H2O2)损伤,及星形胶质细胞胰岛素样生长因子1(insulin?like growth factor?1,IGF?1)受体阻滞剂AG1024,分别设立Neuron组、Neuron+Stress组、Neuron+Astrocyte +Stress组及Neuron+Astrocyte+Stress+AG1024组,通过Western blot测定神经元GJA1?20k、去磷酸化(non?phosphorylated,NP)?Cx43的表达,谷氨酸转运酶(glutamate transporter?1,GLT?1)、线粒体功能相关蛋白(PGC?1α、mtTFA、Tom20、CoxⅣ)、凋亡相关蛋白(Bcl?2、Bax、Caspases?9)的变化,采用酶联免疫荧光分析(ELFA)及ELISA法分别检测氧化应激因子NAPDH 氧化酶活性和白介素(interleukin,IL)?1β、IL?6、肿瘤坏死因子?α(tumor necrosis factor?α,TNF?α)等炎性因子含量的变化,采用Annexin V?FITC/PI测定神经元凋亡。结果:星形胶质细胞共培养可明显上调在神经元氧化损伤后内源性GJA1?20k和NP?Cx43表达,抑制PGC?1α、mtTFA、Tom20、CoxⅣ的下调,上调凋亡抑制因子Bcl?2表达,下调凋亡促进因子Bax、Caspase?9表达,降低NAPDH氧化酶活性,降低炎性产物IL?1β、IL?6和TNF?α的水平及抑制神经元的凋亡(P < 0.05)。在给予AG1024后,可明显抑制与以上因素相关的星形胶质细胞对神经元的保护作用(P < 0.05)。结论:与IGF?1相关的星形胶质细胞对神经元的保护机制可能与增加神经元内源性GJA1?20k的含量及线粒体功能的保护有关。
英文摘要:
      Objective:To observe the neuroprotective mechanism of astrocytes after oxidative stress injury by up?regulating endogenous gap junction protein alpha 1 truncated monomer? 20k(GJA1?20k)of neurons. Methods:Neurons and astrocytes were obtained from C57BL/6 fetal mice by primary culture method,and co?culture model was established. Neurons were injured by hydrogen peroxide(H2O2),insulin?like growth factor?1(IGF?1)receptor blocker AG1024 was given to astrocytes,respectively. Thus,Neuron+Astrocyte+Stress group and Neuron+Astrocyte+Stress+AG1024 group were established. Meanwhile,Neuron(Neuron alone without treatment)group and Neuron+Stress group(separately cultured Neuron given with H2O2)were also set up as control groups. The changes of GJA1?20k,non?phosphorylated(NP)?Cx43,glutamate transporter?1(GLT?1),mitochondrial function?related proteins(PGC?1α,mtTFA,Tom20,CoxⅣ),apoptosis?related proteins(Bcl?2,Bax,Caspases?9)were measured by Western blot. The oxidative stress factor NAPDH oxidase activity,inflammatory factors interleukin(IL)?1β,IL?6 and tumor necrosis factor?α(TNF?α)were detected by ELFA and ELISA,apoptosis of neurons was measured by Annexin V?FITC/PI assay,respectively. Results:Astrocytes co?culture significantly up?regulated the expression of endogenous GJA1?20k and NP?Cx43,reversed the down?regulation of PGC?1α,mtTFA,Tom20 and CoxⅣ,up?regulated apoptotic inhibitor Bcl?2,down?regulated apoptotic promoter Bax,Caspase?9,and reduced the expression of NAPDH oxidase activity and inflammatory products IL?1β,IL?6 and TNF?α(P < 0.05). The protective effect of astrocytes on neurons was significantly inhibited by AG1024(P < 0.05). Conclusion:The protective mechanism of astrocyte on neuron associated with IGF?1 may be related to the increase of endogenous GJA1?20k and protection of mitochondria function in neurons.
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