中国组织工程研究

中国组织工程研究 ›› 2022, Vol. 26 ›› Issue (31): 4975-4981.doi: 10.12307/2022.709

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

圣草酚对过氧化氢所致SH-SY5Y细胞线粒体动力学失衡及凋亡的影响

李苏垚1，2，郭敏芳2，于婧文2，孟 涛2，穆秉桃2，李梦迪1，2，李 娜1，2，宋丽娟1，3，马存根1，2， 尉杰忠1，2，4   

  1. 1山西中医药大学神经生物学研究中心/国家中医药管理局益气活血法治疗多发性硬化重点研究室，山西省晋中市   030619；2山西大同大学脑科学研究所，山西省大同市   037009；3山西医科大学第一临床医学院神经内科，山西省太原市   030001；4大同市第四人民医院，山西省大同市   037009
  • 收稿日期:2021-08-24 接受日期:2021-10-11 出版日期:2022-11-08 发布日期:2022-04-25
  • 通讯作者: 尉杰忠，博士，教授，博士生导师，山西中医药大学神经生物学研究中心/国家中医药管理局益气活血法治疗多发性硬化重点研究室，山西省晋中市 030619；山西大同大学脑科学研究所，山西省大同市 037009；大同市第四人民医院，山西省大同市 037009 马存根，博士，教授，博士生导师，山西中医药大学神经生物学研究中心/国家中医药管理局益气活血法治疗多发性硬化重点研究室，山西省晋中市 030619；山西大同大学脑科学研究所，山西省大同市 037009
  • 作者简介:李苏垚，女，河南省平顶山市人，山西中医药大学在读硕士，主要从事神经退行性变疾病的基础和应用研究。 郭敏芳，女，山西省大同市人，2007年青岛大学毕业，硕士，副教授，主要从事神经变性疾病的发病机制和防治研究。
  • 基金资助:
    国家自然科学基金面上项目(81473577)，项目负责人：马存根；中国科学院分子发育生物学国家重点实验室开放课题(2020-MDB-KF-09)，项目负责人：宋丽娟；山西省应用基础研究计划项目(201901D211538)，项目负责人：宋丽娟；山西省高等学校科技创新项目(2019L0734)，项目负责人：宋丽娟；山西省高等学校科技创新项目(2020L0484)，项目负责人： 郭敏芳；山西省卫健委医学科技领军团队(2020TD05)，项目负责人：马存根；山西中医药大学青年科学家培育项目(2021-PY-QN-09)，项目负责人：宋丽娟

Effect of eriodictyol on the imbalance of mitochondrial dynamics and apoptosis in SH-SY5Y cells induced by hydrogen peroxide

Li Suyao1, 2, Guo Minfang2, Yu Jingwen2, Meng Tao2, Mu Bingtao2, Li Mengdi1, 2, Li Na1, 2, Song Lijuan1, 3, Ma Cungen1, 2, Yu Jiezhong1, 2, 4   

  1. 1Research Center of Neurobiology, Shanxi University of Chinese Medicine, Jinzhong 030619, Shanxi Province, China; 2Institute of Brain Science, Shanxi Datong University, Datong 037009, Shanxi Province, China; 3Department of Neurology, First Affiliated Hospital, Shanxi Medical University, Taiyuan 030001, Shanxi Province, China; 4Datong Fourth People’s Hospital, Datong 037009, Shanxi Province, China
  • Received:2021-08-24 Accepted:2021-10-11 Online:2022-11-08 Published:2022-04-25
  • Contact: Yu Jiezhong, MD, Professor, Doctoral supervisor, Research Center of Neurobiology, The Key Research Laboratory of Benefiting Qi for Acting Blood Circulation Method to Treat Multiple Sclerosis of State Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, Shanxi Province, China; Institute of Brain Science, Shanxi Datong University, Datong 037009, Shanxi Province, China; Datong Fourth People’s Hospital, Datong 037009, Shanxi Province, China Ma Cungen, MD, Professor, Doctoral supervisor, Research Center of Neurobiology, The Key Research Laboratory of Benefiting Qi for Acting Blood Circulation Method to Treat Multiple Sclerosis of State Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, Shanxi Province, China; Institute of Brain Science, Shanxi Datong University, Datong 037009, Shanxi Province, China
  • About author:Li Suyao, Master candidate, Research Center of Neurobiology, The Key Research Laboratory of Benefiting Qi for Acting Blood Circulation Method to Treat Multiple Sclerosis of State Administration of Traditional Chinese Medicine, Shanxi University of Chinese Medicine, Jinzhong 030619, Shanxi Province, China; Institute of Brain Science, Shanxi Datong University, Datong 037009, Shanxi Province, China Guo Minfang, Master, Associate professor, Institute of Brain Science, Shanxi Datong University, Datong 037009, Shanxi Province, China Li Suyao and Guo Minfang contributed equally to this article.
  • Supported by:
    General Program of the National Natural Science Foundation of China, No. 81473577 (to MCG); Open Project of the State Key Laboratory of Molecular Developmental Biology, Chinese Academy of Sciences, No. 2020-MDB-KF-09 (to SLJ); Shanxi Provincial Applied Basic Research Project, No. 201901D211538 (to SLJ); Shanxi Provincial Colleges and Universities Science and Technology Innovation Project, No. 2019L0734 (to SLJ); Shanxi Provincial Colleges and Universities Science and Technology Innovation Project, No. 2020L0484 (to GMF); the Leading Team of Medical Science and Technology of Shanxi Provincial Health Commission, No. 2020TD05 (to MCG); Young Scientist Training Program of Shanxi University of Chinese Medicine, No. 2021-PY-QN-09 (to SLJ)

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摘要：

文题释义：
圣草酚：作为食源性药物，广泛分布于柑橘类水果和花生中，是具有抗炎、抗氧化和抗癌活性的天然黄酮类化合物。有研究表明黄酮类化合物能够缓解糖尿病大鼠氧化应激，随着研究进一步深入，圣草酚在神经退行性疾病的防治中发挥巨大潜力，具有减轻神经炎症反应、改善神经系统氧化应激损伤等作用。
氧化应激和线粒体动力学：氧化应激是由于活性氧的产生和抗氧化系统两者之间不平衡而形成的氧化还原状态。线粒体是活性氧产生的主要来源，其动力学障碍在阿尔茨海默病的病理过程中发挥关键作用。目前研究表明，氧化应激广泛存在于神经退行性疾病中，通过调控线粒体动力学改善氧化应激可能成为防治阿尔茨海默病的新方法。
SY5Y细胞：为人神经母细胞瘤细胞，属于小梭形的贴壁细胞。

背景：线粒体动力学异常与神经退行性疾病中氧化应激现象密切相关。课题组前期研究发现圣草酚可以减轻阿尔茨海默病神经损伤，但其是否对线粒体动力学有调控作用尚未明确。
目的：探讨圣草酚抑制过氧化氢诱导的SH-SY5Y细胞凋亡的具体机制。
方法：将SH-SY5Y细胞分为如下3组：PBS对照组、过氧化氢(250 μmol/L)模型组、过氧化氢(250 μmol/L)+圣草酚治疗组(10 μmol/L)。干预24 h，应用试剂盒检测丙二醛水平和超氧化物歧化酶活性，显微镜下观察细胞形态，TUNEL染色观察细胞凋亡情况，JC-1染色观察线粒体膜电位，免疫荧光染色法和Western blot检测细胞凋亡蛋白以及线粒体融合和线粒体分裂蛋白的表达。
结果与结论：①与过氧化氢组相比，圣草酚治疗组丙二醛水平显著减少，超氧化物歧化酶活性显著增加；②与PBS对照组相比，过氧化氢组细胞密度下降，胞体肥大，细胞凋亡率增加，线粒体模电位下降，Bcl-2表达减少，Bax表达增加；与过氧化氢组相比，圣草酚治疗组上述指标得到明显改善；③与PBS对照组相比，过氧化氢组细胞线粒体分裂相关蛋白p-DRP1和FIS1的表达量升高，而线粒体融合相关蛋白OPA1和Mfn2的表达量降低；与过氧化氢组相比，圣草酚治疗可以降低p-DRP1和FIS1的表达，增加OPA1和Mfn2的表达；④结果说明，圣草酚可通过调控线粒体动力学抑制过氧化氢诱导的SH-SY5Y细胞凋亡。

https://orcid.org/0000-0003-0049-1658 (马存根)

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

关键词: SH-SY5Y细胞, 阿尔茨海默病, 圣草酚, 过氧化氢, 氧化应激, 凋亡, 线粒体动力学

Abstract: BACKGROUND: Abnormal mitochondrial dynamics is closely related to oxidative stress after affecting neurodegenerative diseases. Our previous studies have shown that eriodictyol can reduce nerve injury in Alzheimer’s disease, but whether it has a regulatory effect on mitochondrial dynamics is not clear.  
OBJECTIVE: To explore the mechanism of eriodictyol on SH-SY5Y cell apoptosis induced by hydrogen peroxide (H2O2).
METHODS:  SH-SY5Y cells were divided into three groups: PBS control group, H2O2 (250 μmol/L) group, and H2O2 (250 μmol/L) plus eriodictyol (10 μmol/L) treatment group (eriodictyol group). The intervention was performed for 24 hours. The kits were utilized to observe the level of malondialdehyde and the activity of superoxide dismutase. Cell morphology was observed under the microscope. TUNEL staining was used to observe the apoptosis. JC-1 staining was used to observe mitochondrial membrane potential. The expression levels of apoptotic proteins and mitochondrial fusion and fission protein-related proteins were detected by immunofluorescence staining and western blot assay.  
RESULTS AND CONCLUSION: (1) A significant decrease of malondialdehyde levels and a significant increase of superoxide dismutase activity were found in eriodictyol treated group, when compared with the H2O2 group. (2) Compared with the PBS control group, the H2O2 group showed decreased cell density, cytosolic hypertrophy, increased apoptosis, decreased mitochondrial membrane potential, decreased Bcl-2 expression, and increased Bax expression. Compared with the H2O2 group, above indexes were significantly improved in the eriodictyol group. (3) Compared with PBS control group, the expression levels of mitochondrial fission-related proteins dynamin-related protein 1 (p-DRP1) and mitochondrial fission protein 1 (FIS1) were elevated and the expression levels of mitochondrial fusion-related proteins optic atrophic protein 1 (OPA1) and mitofusin 2 (Mfn2) were decreased in the H2O2 group. Compared with the H2O2 group, eriodictyol treatment decreased the expression levels of p-DRP1 and FIS1 and increased the expression of OPA1 and Mfn2. (4) The results indicate that eriodictyol can inhibit H2O2-induced apoptosis in SH-SY5Y cells by regulating mitochondrial dynamics.

Key words: SH-SY5Y cells, Alzheimer’s disease, eriodictyol, H2O2, oxidative stress, apoptosis, mitochondrial dynamics

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