解剖学报

解剖学报 ›› 2022, Vol. 53 ›› Issue (1): 11-18.doi: 10.16098/j.issn.0529-1356.2022.01.002

• 神经生物学 • 上一篇    下一篇

18α-甘草次酸通过抗氧化作用促进成年小鼠室管膜下区神经干细胞增殖

刘雪芹1 牛晓洁1 王必慧1 王婷玉2 杨静1 陆利1*   

  1. 1.山西医科大学人体解剖学教研室; 2.山西医科大学第二临床医学系，太原 030001
  • 收稿日期:2020-06-11 修回日期:2020-08-03 出版日期:2022-02-06 发布日期:2022-02-06
  • 通讯作者: 陆利 E-mail:luli@sxmu.edu.cn
  • 基金资助:
    国家自然科学基金面上项目;山西医科大学卓越医生教育培养计划2.0指令性项目

18α-glycyrrhetinic acid promoting neural stem cells proliferation  through antioxidant in the subventricular zone of adult mice

LIU Xue-qin1  NIU Xiao-jie1  WANG Bi-hui1  WANG Ting-yu2  YANG Jing1  LU Li1*   

  1. 1. Department of Human Anatomy, Shanxi Medical University, Taiyuan 030001, China; 2. The Second Department of Clinical Medicine, Shanxi Medical University, Taiyuan 030001, China
  • Received:2020-06-11 Revised:2020-08-03 Online:2022-02-06 Published:2022-02-06
  • Contact: LU Li E-mail:luli@sxmu.edu.cn

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摘要：

目的  探讨18α-甘草次酸（18α-GA）对成年小鼠室管膜下区（SVZ）神经干细胞（NSCs）增殖的影响及其机制。  方法  100只6月龄BALB/c小鼠随机平均分为18α-GA组（腹腔注射18α-GA 40 mg/kg两个月，以DMSO为溶解介质）和DMSO对照组（腹腔注射含等体积DMSO溶解介质的PBS溶液），每组50只，其中15只用于在体实验，其余35只小鼠断头取脑，分离培养SVZ区NSCs。采用Ki-67染色、成球实验、BrdU掺入实验及CCK-8实验检测SVZ区NSCs增殖活力，二氢乙啶（DHE）染色等方法检测活性氧簇(ROS）及超氧化物歧化酶1（SOD1）水平， Real-time PCR及Western blotting检测SVZ区NSCs 的核因子E2相关因子2 (Nrf2)表达。   结果  在体实验结果显示，腹腔注射18α-GA后，小鼠SVZ区Ki-67阳性细胞数较DMSO对照组明显增多，ROS水平较DMSO对照组明显下降，而SOD1在mRNA和蛋白水平均明显升高，分别是对照组的（3.17±0.073）倍和（2.12±0.02）倍（P<0.05和P<0.001）。细胞实验结果表明，18α-GA组小鼠SVZ区的NSCs成球速度快，数量多，其中直径在40～60 μm和≥60μm的神经球数量分别是DMSO对照组的2.1倍和4倍（P<0.001）；18α-GA组小鼠SVZ区的NSCs活力明显提高（P<0.001），BrdU阳性率是DMSO对照组的（1.75±0.17）倍（P<0.01）；18α-GA组小鼠SVZ区NSCs的ROS水平较DMSO对照组下降18.91%±4.33%（P<0.05），而SOD1在mRNA和蛋白水平均明显升高（P<0.05和 P<0.01）。此外，腹腔注射18α-GA后小鼠SVZ区及其NSCs中Nrf2的mRNA和蛋白水平均高于对照组（P<0.01和 P<0.05）。  结论  在体给予18α-GA可提高成年NSCs的SOD1活性，减少细胞内ROS堆积，维持和改善成年小鼠SVZ区NSCs的增殖潜能。 

关键词: 18α-甘草次酸；神经干细胞；抗氧化；免疫印迹法；小鼠 

Abstract:

Objective  To explore the effect of 18α-glycyrrhetinic acid (18α-GA) on the proliferation of adult mice neural stem cells (NSCs) and its underlying mechanism.    Methods  One hundred 6-month BALB/c mice were randomly divided into DMSO control group and 18α-GA group (mice were intraperitoneally injected with 40 mg / kg 18α-GA every day for 2 months). The proliferation capability, oxidative status and  nuclear factor E2-related factor 2 (Nrf2) level of NSCs in the adult mice subventricular zone (SVZ) were measured through both in vivoand  in vitro experiments, including Ki-67 staining, neurosphere formation assay, BrdU incorporation, CCK-8 assay, reactive oxygen species (ROS) detection, superoxide dismutase 1 (SOD1) determination, Real-time PCR and Western blotting.    Results  Elevated Ki-67 positive cells were observed in SVZ of mice with 18α-GA application. Meanwhile, ROS level attenuated but SOD1 mRNA and protein level increased significantly in the SVZ of 18α-GA group mice, the latter of which were (3.17±0.073) and (2.12±0.02) times respectively than that of the control group (P<0.05 and P<0.001). Likewise, the similar changes were exhibited in vitro data. NSCs of 18α-GA group mice displayed higher proliferation potency confirmed by accelerated neurosphere formation and increased neurosphere number (P<0.001), as well as higher BrdU positive ratio (P<0.01) and NSCs vitality (P<0.001). NSCs of mice with 18α-GA injection exhibited decreased ROS level by 18.91%±4.33% (P<0.05) and enhanced SOD1 level, compared with those in NSCs of DMSO group mice. Furtherly, the Nrf2 expression in SVZ and NSCs of 18α-GA group mice was higher than that of the control group.  Conclution  18α-GA administration plays a vital role in the maintainence and amelioration of adult mice NSCs proliferation through activating SOD1 and diminishing ROS aggregations. 

Key words: 18α-glycyrrhetinic acid, Neural stem cell, Antioxidant, Western blotting, Mouse

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