摘要
背景:选择合适的诱导方法诱导人胎盘间充质干细胞分化为神经元样细胞是临床治疗神经损伤的关键。目的:在人胎盘间充质干细胞中加入商品化的人胚胎脑组织蛋白提取物,观察其诱导分化为神经元样细胞的可行性。方法:采用酶消化法分离培养人胎盘间充质干细胞,传至第3代,将人早期胚胎脑组织蛋白提取物加入诱导培养基培养6 d,加入浓度为20 nmol/L全反式维甲酸和500μg/L音猬因子培养3 d,换新的培养液,包含体积分数为10%胎牛血清,10μg/L脑源性神经营养因子,20μg/L神经胶质细胞源性神经营养因子,50μg/L胰岛素样生长因子Ⅱ型继续培养3 d。结果与结论:胎盘组织经酶消化后获得贴壁细胞,传至第2代细胞形态为梭形,成漩涡样生长,传至第3代细胞形态较均一。诱导后细胞经免疫细胞化学法检测均表达神经元特异性烯醇化酶、巢蛋白、胶质纤维酸性蛋白、神经丝蛋白、神经生长相关蛋白43;ELISA法检测细胞培养上清脑源性神经营养因子、神经营养因子3、神经营养因子4为阳性表达;RT-PCR检测细胞微管相关蛋白、神经元特异性烯醇化酶、神经丝蛋白、神经生长相关蛋白43均为阳性表达。结果表明人胚胎脑组织蛋白提取物使人胎盘间充质干细胞诱导分化为神经元样细胞。
BACKGROUND： It is a key to choose an appropriate method to trans-differentiate mesenchymal stem cells from human placenta into neuron-like cells for clinical treatment of neural system injury. OBJECTIVE： To observe the feasibility of the neuronal differentiation of mesenchymal stem cells from human placenta with protein extracts of human embryonic brain tissue. METHODS： Mesenchymal stem cells from human placenta were isolated and cultured using enzyme digestion method. The third passage of cells were incubated in induction medium containing protein extracts of human embryonic brain tissue for 6 days, and then cultured in culture medium containing 20 nmol/L all-trans retinoic acid and 500 μg/L sonic hedgehog for 3 days, followed by 3-day continuous culture in 10 μg/L brain-derived neuretrophic factor, 20 μg/L glial cell line-derived neurotrephic factor, 50 μg/L insulin-like growth factor type II RESULTS AND CONCLUSION： The adherent cells were obtained after enzyme digestion of placenta tissue. The passage 2 cells were fusiform-shaped and exhibited a whirlpool-like growth. The passage 3 cells were uniform inmorphology. After induction, immunocytochemistry assay showed that the cells expressed neuron-specific enolase, nestin, glial fibrillary acidic protein, neurofilament protein, and nerve growth associated protein 43; ELISA results showed that cells were positive for brain-derived neurotrophic factor, neurotrophin 3, neurotrophin 4; RT-PCR results showed that cells were positive for microtubule-associated protein, neuron-specific enolase, neurofllament protein, nerve growth associated protein 43. Results indicate that mesenchymal stem cells from human placenta can differentiate into neuron-like cells under induction of protein extracts of human embryonic brain tissue.
出处
《中国组织工程研究》
CAS
CSCD
2014年第50期8168-8173,共6页
Chinese Journal of Tissue Engineering Research
基金
宁夏自然科学基金项目(NZ1229)
宁夏医科大学校级课题(XM201135)~~
