中国神经再生研究(英文版)

中国神经再生研究(英文版) ›› 2019, Vol. 14 ›› Issue (5): 826-833.doi: 10.4103/1673-5374.249230

• 原著:脑损伤修复保护与再生 • 上一篇    下一篇

1,10邻二氮杂菲双过氧钒酸钾(BpV)通过增加DNA甲基转移表达酶抑制海马神经元细胞系增殖

  

  • 出版日期:2019-05-15 发布日期:2019-05-15
  • 基金资助:

    国家自然科学基金(81160244，81360316，81460283，81660307)；中国内蒙古科学基金(2018LH08078，2016MS(LH)0307)；包头市卫生基金(2018LH08078，2016MS(LH)0307，WSJJ 2016008；中国内蒙古教育研究基金(NJZY207、NJZY 17243、NJZY 17250、NJZY 17251)；包头医学院基金(BYJJ-DF201602，BYJJ-YF201615，BSJJ 201617，BYJJ-QM201633，BYJJ-QM201656，BYJJ201502)；包头市科技规划项目(CX2017-5);国家关键项目(2017YFC1308405)

Potassium bisperoxo (1,10-phenanthroline) oxovanadate suppresses proliferation of hippocampal neuronal cell lines by increasing DNA methyltransferases

Xiao-Li Tian 1, 2, 3 , Shu-Yuan Jiang 1, 2 , Xiao-Lu Zhang 1, 2, 3 , Jie Yang 1, 2 , Jun-He Cui 1, 2 , Xiao-Lei Liu 1, 2 , Ke-Rui Gong 4 , Shao-Chun Yan 1, 2 , Chun-Yang Zhang 5 , Guo Shao 1, 2, 3   

  1. 1 Biomedicine Research Center, Basic Medical College and Baotou Medical College of Neuroscience Institute, Baotou Medical College, Baotou, Inner Mongolia Autonomous Region, China
    2 Inner Mongolia Key Laboratory of Hypoxic Translational Medicine, Baotou Medical College, Baotou, Inner Mongolia Autonomous Region, China
    3 Beijing Key Laboratory of Hypoxic Conditioning Translational Medicine, Xuanwu Hospital, Capital Medical University, Beijing, China
    4 Department of Oral and Maxillofacial Surgery, University of California San Francsico, San Francisco, CA, USA.
    5 Department of Neurosurgery, the First Affiliated Hospital of Baotou Medical College, Baotou, Inner Mongolia Autonomous Region, China
  • Online:2019-05-15 Published:2019-05-15
  • Contact: Guo Shao, MD, shao_guo_china@163.com; Chun-Yang Zhang, zhangchunyangbyyfy@163.com.
  • Supported by:

    This study was supported by the National Natural Science Foundation of China, No. 81160244, 81360316, 81460283, 81660307 (all to GS); the Inner Mongolia Science Foundation of China, No. 2018LH08078 (to GS), 2016MS(LH)0307 (to SYJ); the Baotou Health Foundation, China, No. WSJJ2016008 (to SYJ); the Inner Mongolia Educational Research Foundation of China, No. NJZY207 (to GS), NJZY17243 (to SCY), NJZY17250 (to XLL), NJZY17251 (to SYJ); the Baotou Medical College Foundation of China, No. BYJJ-DF201602, BYJJ-YF201615, BSJJ201617, BYJJ-QM201633, BYJJ-QM201656, BYJJ201502 (to GS); the Science and Technology Planning Project of Baotou of China, No.CX2017-5 (to GS); the National Key R&D Program of China, No. 2017YFC1308405 (to GS).

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摘要：

有报道显示，1,10邻二氮杂菲双过氧钒酸钾(BpV (phen))能够阻滞细胞周期，为了验证BpV (phen)是否通过影响DNA甲基转移酶来调节基因的表达，进而影响细胞周期的变化，实验将来源于中国医学科学院肿瘤研究所、北京协和医学院分子肿瘤学国家实验室的HT22小鼠海马神经元细胞系，分别加入0.3和3 μM的BpV(Alexis, San Diego, USA)进行培养。采用incuCyte实时视频图像系统、倒置显微镜、MTS方法细胞增殖情况、观察形态学变化、检测细胞活力的变化，并应用Q-PCR和免疫荧光染色检测DNA甲基转移酶和细胞周期调节因子P21的mRNA和蛋白水平上的变化，应用ELISA进行了DNA甲基转移酶活力的检测。应用流式细胞术检测BpV (phen)对细胞周期的影响。结果发现：(1) 0.3 μM BpV (phen)对HT22细胞的增殖、形态、细胞活力均无影响；(2)然而当浓度增加到3 μM BpV (phen)时，HT22细胞活力下降，细胞周期明显阻滞于S期，而DNA甲基转移酶3B的mRNA和蛋白表达量增加；(3)从而得出，BpV (phen) 3 μM可能通过增加小鼠海马神经元DNA甲基转移酶的表达抑制HT22细胞的增殖和阻滞细胞周期，并增加周期调节因子P21表达量。

orcid: 0000-0003-3766-5076 (Guo Shao)
           0000-0002-0310-0205 (Chun-Yang Zhang)

关键词: 海马神经元, 1,10邻二氮杂菲双过氧钒酸钾, DNA甲基转移酶, p21, HT22细胞, 细胞周期, 免疫印迹, DNA甲基化, 神经再生

Abstract:

Bisperoxo (1,10-phenanthroline) oxovanadate (BpV) can reportedly block the cell cycle. The present study examined whether BpV alters gene expression by affecting DNA methyltransferases (DNMTs), which would impact the cell cycle. Immortalized mouse hippocampal neu¬ronal precursor cells (HT22) were treated with 0.3 or 3 μM BpV. Proliferation, morphology, and viability of HT22 cells were detected with an IncuCyte real-time video imaging system or inverted microscope and 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethonyphenol)-2-(4-sul¬fophenyl)-2H-tetrazolium, respectively. mRNA and protein expression of DNMTs and p21 in HT22 cells was detected by real-time polymerase chain reaction and immunoblotting, respectively. In addition, DNMT activity was measured with an enzyme-linked immuno¬sorbent assay. Effects of BpV on the cell cycle were analyzed using flow cytometry. Results demonstrated that treatment with 0.3 μM BpV did not affect cell proliferation, morphology, or viability; however, treatment with 3 μM BpV decreased cell viability, increased expression of both DNMT3B mRNA and protein, and inhibited the proliferation of HT22 cells; and 3 μM BpV also blocked the cell cycle and increased expres¬sion of the regulatory factor p21 by increasing DNMT expression in mouse hippocampal neurons.

Key words: nerve regeneration, hippocampal neurons, potassium bisperoxo (1,10-phenanthroline) oxovanadate, DNA methyltransferase, p21, HT22 cell, cell cycle, immunoblotting, DNA methylation, neural regeneration