nNOS参与OGD介导的促神经干细胞增殖效应

中国临床药理学与治疗学 ›› 2014, Vol. 19 ›› Issue (10): 1099-1106.

nNOS参与OGD介导的促神经干细胞增殖效应

钱晓丹, 罗春霞, 朱东亚

南京医科大学药学院药理实验室,南京 21000,江苏

收稿日期:2014-01-23 修回日期:2014-04-25 出版日期:2014-10-26 发布日期:2014-10-29
通讯作者: 朱东亚,男,教授,研究方向:神经药理学,脑缺血、抑郁症与神经再生,疼痛。Tel: 13016934689 E-mail: dyzhu@njmu.edu.cn
作者简介:钱晓丹,女,硕士,研究方向:脑缺血与神经再生。 Tel: 15850688027 E-mail: qxd@njmu.edu.cn。罗春霞,女,副教授,研究方向:神经药理学,脑缺血与神经再生。 Tel: 13913904160 E-mail: chunxialuo@njmu.edu.cn

nNOS is required for enhancing proliferation of NSCs induced by OGD

QIAN Xiao-dan, LUO Chun-xia, ZHU Dong-ya

Department of Pharmacology, Pharmacy School, Nanjing Medical University, Nanjing 21000, Jiangsu, China

Received:2014-01-23 Revised:2014-04-25 Online:2014-10-26 Published:2014-10-29

摘要/Abstract

摘要： 目的体外研究脑缺血促神经发生的机制。方法体外培养神经干细胞(neural stem cells,NSCs)与海马神经元,并采用氧糖剥夺模型(oxygen and glucose deprivation, OGD)模拟在体缺血,通过RT-PCR、Western blot、免疫细胞化学、酶活性测定、亚硝酸盐/硝酸盐(nitrite/nitrate,NOx) 含量测定等多种方法研究神经干细胞的生物学行为以及介导这种效应的分子机制。结果 (1)OGD通过直接和间接(神经元)作用增加神经干细胞中BrdU⁺ 细胞比例。(2)OGD上调神经干细胞中神经元型一氧化氮合酶(neuronal nitric oxide synthase,nNOS)并下调神经元中nNOS,且都有利于神经干细胞增殖。(3)采用nNOS基因敲除小鼠来源的神经干细胞以及神经元(用于共培养)进行实验,发现OGD并不能升高神经干细胞中BrdU⁺ 细胞比例。结论神经干细胞和神经元中nNOS水平变化共同参与了OGD诱导的促神经干细胞增殖效应。

关键词: 神经元型一氧化氮合酶, 神经干细胞, 氧糖剥夺, 增殖

Abstract: AIM: To explore the mechanism of ischemia-induced neurogenesis in vitro. METHODS: Monolayer-cultured neural stem cells (NSCs) or neurons (before co-culture) were exposed to oxygen and glucose deprivation (OGD) for 2 h or 2.5 h. Cell proliferation was assessed by BrdU incorporation. We also performed RT-PCR, western blot, immunofluorescence, NOS activity assay and NOx content measurement to detect alteration of neuronal nitric oxide synthase (nNOS) in NSCs and neurons. RESULTS: That NSCs exposed to OGD alone or co-cultured with the OGD-injured neurons displayed a remarkably active proliferation rate. It was demonstrated that OGD exposure up-regulated nNOS in NSCs and down-regulated nNOS in neurons. nNOS knockout in NSCs or neurons( for co-culture experiment) reversed the enhanced proliferation rate of NSCs induced by OGD.CONCLUSION: nNOS alterations both in NSCs and neurons accounts for the OGD-induced NSCs proliferation.

Key words: neuronal nitric oxide synthase, neural stem cells, OGD, proliferation

中图分类号:

R965.2

钱晓丹, 罗春霞, 朱东亚. nNOS参与OGD介导的促神经干细胞增殖效应[J]. 中国临床药理学与治疗学, 2014, 19(10): 1099-1106.

QIAN Xiao-dan, LUO Chun-xia, ZHU Dong-ya. nNOS is required for enhancing proliferation of NSCs induced by OGD[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2014, 19(10): 1099-1106.

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