新生小鼠海马神经干细胞体外培养条件下的增殖与分化

背景：神经干细胞移植可用于中枢神经损伤的临床治疗。 目的：观察新生小鼠海马神经干细胞在体外培养条件下的增殖和分化。 方法：从新生昆明小鼠海马取材,采用机械分离法对原代细胞进行无血清培养,采用机械法复合酶消化法对原代细胞进行传代,以体积分数为10%胎牛血清诱导分化。免疫荧光行巢蛋白、β-微管蛋白Ⅲ和胶质纤维酸性蛋白染色,对培养的细胞进行鉴定。CCK-8法检测神经干细胞增殖能力。 结果与结论：从新生小鼠海马分离得到的细胞具有连续传代形成克隆球的能力,克隆球呈巢蛋白免疫反应阳性;加入胎牛血清可诱导分化为β-微管蛋白Ⅲ和胶质纤维酸性蛋白阳性细胞。提示实验成功建立了体外培养新生小鼠海马组织分离和培养神经干细胞的方法,培养的神经干细胞在体外培养条件下保持着自我增殖和分化的潜能。

Proliferation and differentiation of neural stem cells from newborn mouse hippocampi in vitro

BACKGROUND:Neural stem cells transplantation can be used for clinical therapy of central nervous system injury. OBJECTIVE:To observe the characteristics of proliferation and differentiation of neural stem cells cultured in vitro. METHODS:Neural stem cells were isolated mechanically from newborn Kunming mouse hippocampus and cultured with serum-free cell culture medium. They were obtained by mechanical dissociation and enzymatic digestion. The neurospheres were identified by immunofluorescent staining of Nestin. The differentiation of neural stem cells was induced by 10% fetal bovine serum and identified by immunohistochemistry of β-Ⅲ-tubulin and glial fibrillary acidic protein. CCK-8 assay was used to ascertain the capacity for neural stem cell proliferation. RESULTS AND CONCLUSION:The neural stem cells obtained from newborn mouse hippocampus had the capacity for forming the neurospheres which were positive for Nestin. The differentiated cells induced by 10% fetal bovine serum expressed β-Ⅲ-tubulin or glial fibrillary acidic protein. The method of isolation  and culture of neural stem cells from newborn mouse hippocampus in vitro was successfully established. Neural stem cells have the capacity for self-renewal and the potential for differentiation.