1.湖北医药学院附属襄阳市第一人民医院麻醉科，湖北 襄阳 441000;2.湖北医药学院附属襄阳市第一人民医院中心实验室，湖北 襄阳 441000;3.湖北医药学院附属襄阳市第一人民医院康复科，湖北 襄阳 441000;4.湖北省帕金森病临床研究中心，湖北 襄阳 441000
李傲涵，男，硕士，住院医师。 Email：446210721@qq.com。
罗辉宇 ，男，博士，主任医师。Email：603983267@qq.com。
湖北省科技计划项目（2021CFB582）；襄阳市科技局项目（2021ZD13）；襄阳市第一人民医院院级科研平台（XYY2022P02）。
1.Department of Anesthesiology， Xiangyang No.1 People's Hospital， Hubei University of Medicine， Xiangyang， Hubei 441000， China;2.Central Liaboratory， Xiangyang No.1 People's Hospital， Hubei University of Medicine， Xiangyang， Hubei 441000， China;3.Department of Rehabilitation Medicine， Xiangyang No.1 People's Hospital， Hubei University of Medicine， Xiangyang， Hubei 441000， China;4.Hubei Clinical Research Center of Parkinson's Disease， Xiangyang， Hubei 441000， China
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目的 探讨褪黑素（melatonin，MT）在1-甲基-4-苯基-吡啶离子（1-methyl-4-phenylpyridinium ion，MPP+）诱导的帕金森病体外模型中的作用及分子机制。方法 将MN9D细胞分为对照组、MPP+组、MT组、治疗组。采用细胞计数试剂盒8检测MT和MPP+对细胞活力的影响；采用线粒体膜电位检测试剂盒（JC-1）评价线粒体功能；Hoechst/PI双染法检测细胞凋亡；通过免疫荧光蛋白质印迹法（Western blotting）检测凋亡相关蛋白［Cleaved-Caspase 3和细胞色素C（cytochrome C，CytC）］以及泛醌―细胞色素C还原酶核心蛋白1（ubiquinol-cytochrome C reductase core protein 1，UQCRC1）蛋白的表达；采用干扰RNA技术沉默MN9D细胞中UQCRC1的表达，然后采用Western blotting检测凋亡相关蛋白表达。结果 MT可以减轻MPP+诱导的细胞活力下降（P<0.05）；恢复MPP+造成的线粒体膜电位下降（P<0.05）；减少MPP+诱导的凋亡细胞数量（P<0.05）；抑制凋亡蛋白（CytC和Cleaved-Caspase3）的表达（P<0.05）；上调UQCRC1的表达（P<0.05）。沉默UQCRC1后，MT组和治疗组的UQCRC1表达均下降（P<0.05）；MT对MPP+诱导细胞凋亡的保护作用下降（P<0.05）；凋亡蛋白（CytC和Cleaved-Caspase3）的表达增加（P<0.05）。结论 MT对 MPP+诱导的多巴胺能神经元损伤具有保护作用，其机制可能是通过上调UQCRC1抑制神经元凋亡。 [国际神经病学神经外科学杂志, 2023, 50(3): 26-31]
Objective To investigate the effects of melatonin in an in vitro model of Parkinson's disease induced by 1-methyl-4-phenylpyridinium ion （MPP+） and the molecular mechanisms.Methods MN9D cells were divided into control group， MPP+ group， melatonin group， and treatment group. The effects of melatonin and MPP+ on cell viability were determined using cell counting kit-8. Mitochondrial function was evaluated using the mitochondrial membrane potential assay kit JC-1. Cell apoptosis was determined with Hoechst/propidium iodide double staining. Immunofluorescence assay and Western blotting were used to measure the expression of apoptotic proteins ［cleaved caspase-3 and cytochrome C （CytC）］ and ubiquinol-cytochrome c reductase core protein 1 （UQCRC1）. RNA interference technology was used to silence UQCRC1 expression in MN9D cells to measure the expression of the apoptotic proteins by Western blotting.Results Melatonin significantly inhibited MPP+-induced decreases in cell viability and mitochondrial membrane potential， significantly reduced the number of MPP+-induced apoptotic cells， significantly down-regulated the expression of CytC and cleaved-caspase 3， and significantly up-regulated the expression of UQCRC1 （all P<0.05）. After silencing UQCRC1， the expression of UQCRC1 in the melatonin group and the treatment group was significantly decreased； the protective effect of melatonin against MPP+-induced apoptosis was significantly decreased； and the expression of CytC and cleaved-caspase 3 was significantly increased （all P<0.05）.Conclusions Melatonin has a protective effect against MPP+-induced damage to dopaminergic neurons， possibly by upregulating UQCRC1 to inhibit neuronal apoptosis. ［Journal of International Neurology and Neurosurgery, 2023, 50(3): 26-31］
引用本文
李傲涵,曾炼,刘颖,张振,胡鹏超,丁旭东,罗辉宇456.褪黑素通过上调泛醌―细胞色素C还原酶核心蛋白1抑制MPP+诱导的MN9D细胞凋亡和线粒体损伤[J].国际神经病学神经外科学杂志,2023,50(3):26-31111LI Aohan, ZENG Lian, LIU Ying, ZHANG Zhen, HU Pengchao, DING Xudong, LUO Huiyu222. Melatonin inhibits MPP+-induced apoptosis and mitochondrial damage in MN9D cells through upregulating ubiquinol-cytochrome c reductase core protein 1[J]. Journal of International Neurology and Neurosurgery,2023,50(3):26-31
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- 收稿日期:2022-10-26
- 最后修改日期:2023-02-24
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- 在线发布日期: 2023-08-16
