黄芪诱导大鼠骨髓间充质干细胞分化为神经样细胞

背景:体外培养扩增的骨髓间充质干细胞可诱导分化为神经细胞,但是有些诱导剂有一定的毒性,不能应用于人体内.目的:以黄芪为诱导剂,诱导大鼠骨髓间充质干细胞分化为神经样细胞.设计、时间及地点:随机对照实验,于2002-01/2005-01 在大理学院基础医学院的云南省重点实验室和四川大学基础医学院与法医学院的干细胞研究中心完成.材料:6周龄健康SD大鼠,体质量120～130g.方法:以密度梯度离心法从大鼠骨髓中分离培养骨髓间充质干细胞,并传代扩增.采含体积分数为0.125的黄芪无血清L-DMEM培养液诱导分化为神经样细胞,观察细胞形态变化,以免疫组织化学染色方法检测分化细胞中巢蛋白、神经元特异性烯醇化酶、胶窒宋嵝缘鞍椎谋泶?主要观察指标:①诱导分化细胞的免疫组织化学鉴定.②RT-PCR方法半定量分析相关基因Ngn-1和Wnt-1基因在诱导过程中的表达.结果:经传代后,骨髓间充质干细胞呈纤维细胞样,有较强的增殖能力.经黄芪诱导后,细胞形态发生改变,巢蛋白、神经元特异性烯醇化酶及胶质纤维酸性蛋白均阳性.Ngn-1和Wnt-1基因在黄芪诱导骨髓间充质干细胞过程中表达量升高.结论:骨髓间充质干细胞在黄芪诱导下可向神经样细胞分化.Ngn-1和Wnt-1基因在其分化过程中起正调控作用.

Differentiation of rat bone marrow mesenchymal stem cells into neuron-like cells induced by astragalus mongholicus

BACKGROUND: In vitro cultured bone marrow mesenchymal stem cells (BMSCs) can differentiate into neural cells. Someinductors are poisonous and cannot be used in human. OBJECTIVE: To induce the differentiation of rat BMSCs into neuron-like cells with astragalus mongholiens. DESIGN, TIME AND SETTING: The randomized control experiment was performed at the Yunnan Key Laboratory of Faculty of Basic Medicine of Dali University and Stem Cell Research Center of College of Preclinical Medicine and Forensic.Medicine of Sichuan University from January 2002 to January 2005. MATERIALS: Healthy SD rats aged 6-weeks old and weighing 120-130 g were used in this study. METHODS: BMSCs were collected from rat bone marrow by density gradient centrifugation, and then cultured. BMSCs were incubated in serum-free L-DMEM medium containing 0.125 volume fraction of astragalus mongholicus to observe morphologic changes in differentiated cells. Expression of the nestin, neuron specific enolase (NSE) and glial fibrillary acidic protein (GFAP) was detected by immunohistochemistry. MAIN OUTCOME MEASURES: Immunohistochemistry of differentiated cells; The expression of Wnt-1 gene and Ngn-1 gene were detected by semi-quantitative reverse transcdption-polymerase chain reaction during cell differentiation. RESULTS: After passage, BMSCs were fibroblast-like, and had high reproductive activity. After cultured with astragalus mongholicus, BMSC morphous changed. Nestin, NSE and GFAP were positive. The expression of Wnt-1 gene and Ngn-1 gene increased after astragalus mongholicus treatment. CONCLUSION: BMSCs can differentiate into neuron-like cells induced by astragalus mongholicus. Wnt-1 gene and Ngn-1 gene play a positive regulatory effect during the differentiation.