人恶性胶质瘤细胞株U251三种克隆的分化特性和胶质瘤干细胞的初步鉴定

Differentiation property of three types of colonies derived from malignant human glioma cell line U251 and identification of glioma stem cells

目的观测U251细胞的克隆在形态和分化上的差异，鉴定含胶质瘤干细胞的克隆。方法克隆形成实验观察U251细胞克隆的形态并分类，免疫细胞化学染色观测不同克隆GFAP和nestin的表达差异。分离低分化克隆，检测其自我更新能力、多向分化潜能及CD133表达情况。结果U251细胞可形成紧密型、中间型和松散型三种克隆类型。紧密型克隆分化程度最低，此克隆在无血清培养基内形成神经干细胞样细胞球，具有自我更新能力、多向分化潜能，并表达CD133。结论U251细胞的克隆在形态和分化程度上存在差异，紧密型克隆可能富含胶质瘤干细胞。

Objective To investigate morphological and differential heterogeneities of U251 glioma cell colonies, and identifying the eolonies comprised of tumor stem cells. Method Colony formation assay was employed to observe and classify the colonies derived from U251 glioma cells, then glial fibrillary acidic protein （GFAP） and nestin were labeled in all colonies to determine differentiation status. Furthermore, we isolated and cultured the colonies with the poorest differentiation, and the properties of tumor stem cells： self-renewal, multiple lineage differentiation potential, and expression of stem cell marker CD133 were assayed. Results Colonies derived from U251 glioma cells had three types： tight, transitional and loose colonies. Among these colonies, the tight colonies showed the poorest differentiation. U251 cells of tight colonies possessed the properties of self-renewal and multiple lineage differentiation. Moreover, they were found to be able to express CD133 and nestin. Conclusions U251 cell colonies varied in morphology and differentiation, and the tight colonies might be comprised of tumor stem cells. Our study provided a convenient and efficient method for cloning glioma stem cells.