青蒿琥酯通过PI3K-Akt通路促进大鼠室管膜下区神经干细胞增殖的体外研究  

Artesunate promotes proliferation of rat neural stem cells from subventricular zone via PI3K-Akt pathway in vitro

作　　者：葛红飞[1] 左世伦[1] 李强[1] 谭亮[1] 吴江怡 蒋周阳[1] 喻安永[1] 兰川[1] 胡荣[1] 冯华[1] 

机构地区：[1]第三军医大学西南医院神经外科,全军神经外科研究所,全军神经创伤防治重点实验室,重庆400038 [2]南华大学附属第一医院骨科,湖南衡阳421000

出　　处：《第三军医大学学报》2015年第8期767-771,共5页Journal of Third Military Medical University

基　　金：国家自然科学基金(81371340);国家重点基础研究发展计划(973计划;2014CB541600)~~

摘　　要：目的探讨青蒿琥酯(artesunate,ART)对室管膜下区(subventricular zone,SVZ)神经干细胞(neural stem cells,NSCs)增殖的影响。方法不同浓度的ART(400 nmol/L、800 nmol/L、1.6μmol/L、3.2μmol/L)作用于NSCs 24 h后,采用CCK-8法检测其对NSCs增殖的影响。促增殖作用明显的800 nmol/L ART组加入10μmol/L PI3K-Akt通路阻断剂(LY294002),采用CCK-8法检测该通路对NSCs增殖的影响;分别设置空白对照组、LY294002组、800 nmol/L ART组、800 nmol/L ART+LY294002组,待细胞培养24 h时,行Ki-67免疫荧光染色观察ART对NSCs增殖的影响及PI3K-Akt通路在其中的作用,同时采用Western blot检测各组Nestin、Akt及p-Akt蛋白的表达水平。结果 1CCK-8法检测结果显示800 nmol/L ART组在波长450 nm处的光密度值较空白对照组明显升高,在加入LY294002后,其光密度值明显下降(P<0.05),与空白对照组水平相当;2Ki-67免疫荧光染色结果提示800 nmol/L ART可促进NSCs增殖,LY294002可抑制这一作用;3Western blot检测结果显示:神经干细胞表面标志蛋白Nestin及p-Akt蛋白表达水平在800 nmol/L ART组明显上调,在加入LY294002后两者表达水平均下调(P<0.05)。结论 ART可在体外促进SVZ区NSCs的增殖,其机制与PI3K-Akt信号通路相关。Objective To investigate the effect of artesunate（ ART） on proliferation of neural stem cells（ NSCs） from subventricular zone（ SVZ） in rats. Methods NSCs were treated with different concentrations of ART（ 0. 4,0. 8,1. 6,and 3. 2 μmol / L） for 24 h,and CCK8 assay was used to evaluate the effect of ART on proliferation of NSCs. The PI3K-Akt pathway inhibitor（ LY294002） was added in the0. 8 μmol / L ART group to detect its inhibitory effect on the proliferation of NSCs. The results of CCK8 assay were confirmed via Ki-67 fluorescence staining. Western blot assay was used to assess the expressions of Nestin,Akt and p-Akt after treatment for 24 h in different groups as follows： control group,LY294002 group,0. 8 μmol / L ART group,and 0. 8 μmol / L ART ＋ LY294002 group. Results（ 1） CCK8 assay showed that the absorbance values at wavelength of 450 nm were significantly higher in the 0. 8 μmol / L ART group than in the control group,and decreased to the level of the control group after addition of LY294002（ P〈0. 05）.（ 2） The Ki-67 positive cell number was higher in the 0. 8 μmol/L ART group as compared with other groups,and decreased to the level of the control group after addition of LY294002.（ 3） Western blot results showed that the expression of Nestin and p-Akt were significantly up-regulated in the 0. 8 μmol / L ART group,but after addition of LY294002,the expression was lowered（ P〈0. 05）. Conclusion ART could promote the proliferation of NSCs from the SVZ,and the PI3K-Akt signaling pathway might play an important role in this process.

关 键 词：青蒿琥酯 神经干细胞 PI3K-AKT信号通路 室管膜下区 

分 类 号：R285.5[医药卫生—中药学] R322.81[医药卫生—中医学]