脑络欣通和左归丸药物血清体外培养神经干细胞的增殖与分化  

Effects of drug-contained sera of Naoluo Xintong versus Zuogui pill on the proliferation and differentiation of in vitro cultured neural stem cells

作　　者：陈立铭[1] 裴磊[2] 

机构地区：[1]长江大学附属第一医院荆州市第一人民医院,湖北省荆州市434000 [2]华中科技大学同济医学院神经生物系,湖北省武汉市430030

出　　处：《中国组织工程研究》2016年第32期4845-4851,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81571078)~~

摘　　要：背景:神经干细胞种子来源数量相对有限,增殖分化量相对较少,如何采取有效的措施促进神经干细胞增殖、分化成为研究的焦点。目的:比较脑络欣通和左归丸药物血清对体外培养的大鼠神经干细胞增殖分化的作用。方法:体外分离培养SD大鼠胚胎神经干细胞,加入含10%的脑络欣通以及10%的左归丸药物血清培养基中共同培养。采用倒置显微镜及免疫荧光染色观察脑络欣通和左归丸药物血清对体外培养的大鼠神经干细胞增殖分化的作用,采用倒置显微镜、免疫荧光染色方法进行观察比较。结果与结论:(1)细胞培养24 h后倒置显微镜下观察细胞开始团生长,并聚集成球,但是细胞直径相对较小,48 h后神经球进一步增大,形态相对规则,但是未见神经球分化;(2)左归丸组细胞培养5 d后细胞球突起平均长度,显著大于脑络欣通组(P<0.05);(3)左归丸药物血清培养下大鼠神经干细胞分化为MAP-2阳性细胞率显著低于脑络欣通组(P<0.05),分化为胶质纤维酸性蛋白阳性细胞率显著高于脑络欣通组(P<0.05);(4)将培养48 h的神经球分别放2种入药物血清中培养,12 h后发现神经球贴壁生长,少量细胞迁移。随着时间的延长,迁移的细胞数量开始增多;(5)免疫细胞荧光染色下显示2种药物血清培养下神经元突出均相对较多,且突起的长度较长,神经干细胞神经元以及星形胶质细胞比例差异不显著(P>0.05)。(6)结果提示脑络欣通和左归丸药物血清对体外培养的大鼠神经干细胞增殖分化均具备明显的促进作用,均能营造更加适宜神经细胞增殖等微环境,但是2种药物存在明显的差异,通过脑络欣通和左归丸药物诱导神经细胞增殖分化是可行的。BACKGROUND： Because of limited source and a relatively weak ability of differentiation and proliferation,how to take positive and effective measures to promote neural stem cell proliferation, differentiation has become the focus of research.OBJECTIVE： To investigate the effects of drug-contained sera of Naoluo Xintong versus Zuogui pill on the proliferation and differentiation of in vitro cultured rat neural stem cells.METHODS： Embryonic neural stem cells of Sprague-Dawley rats were isolated and cultured in vitro, and then were co-cultured with the serum medium containing 10% Naoluo Xintong and 10% Zuogui pill,respectively. Comparative observations were performed between two groups by inverted microscope and immunofluorescence staining.RESULTS AND CONCLUSION： Under the inverted microscope, the cells began to grow in cluster and gather into a ball, but the diameter was relatively small after 24-hour culture; the neurospheres expended further, with relatively regular shape, but no neurosphere differentiation appeared after 48 hours of culture.The average prominent length of the neurospheres in the Zuogui pill group was significantly greater than that in the Naoluo Xintong group after 5 days of culture（P〈0.05）. The rate of rat neural stem cells differentiating into MAP-2-positive cells in the Zuogui pill group was significantly lower than that in the Naoluo Xintong group（P〈0.05）, but the rate of differentiated cells positive for glial fibrillary acidic protein in the Zuogui pill group was significantly higher than that in the Naoluo Xintong group（P〈0.05）. After 48 hours of culture, neurospheres were cultured in the different drug-contained media, and 12 hours later, the neurospheres adhered to the wall, and a small amount of cell migration occurred. Then, cell migration began to increase with time. Under the immunofluorescence staining： prominent neurons with long protrusions were increased in both two groups, but there were no significant differences in the proportion of neurons and astr

关 键 词：干细胞 细胞增殖 细胞分化 神经干细胞 细胞运动 分化 脑络欣通 左归丸 药物血清 体外培养 增殖分化 大脑皮层 倒置显微镜 免疫荧光染色 微环境 国家自然科学基金 

分 类 号：R394.2[医药卫生—医学遗传学]