兔骨髓间充质干细胞的分离、体外培养、成骨诱导与鉴定  

Experimental Study on Culture, Osteogenic Induction and Identification of Rabbit Bone Marrow-derived Mesenchymal Stem Cells in Vitro

作　　者：李伟岸[1] 韦庆军[1] 肖仕辉[1] 韦积华[1] 

机构地区：[1]广西医科大学第一附属医院创伤骨科手外科,广西南宁530021

出　　处：《医学信息》2013年第3期125-127,共3页Journal of Medical Information

摘　　要：目的建立一种兔骨髓间充质干细胞（Mesenchymalstemcell，MSC）体外分离培养、扩增及鉴定的方法。方法采用全骨髓贴壁培养法分离rBMSCs并进行体外扩增及纯化，倒置显微镜下观察原代及传代细胞的形态、生长情况，绘制细胞生长曲线，取第3代细胞分别向成骨细胞和成脂肪细胞定向诱导．通过染色和电镜检测成骨成脂诱导后变化情况。结果采用全骨髓贴壁培养法可获得纯度较高的兔骨髓间充质干细胞，原代和传代培养的rBMSCs具有活跃的增殖能力，成骨成脂诱导后，碱性磷酸酶染色、茜素红染色、vonKossa法染色、I型胶原免疫细胞化学检测和油红。染色均呈阳性。成骨诱导电镜检测呈阳性。结论全骨髓贴壁培养法操作相对简单，可大量分离、纯化、扩增骨髓间充质干细胞，所获细胞具有间充质干细胞的一般生物学特性。经诱导培养后具有多向分化潜能。本实验通过全骨髓贴壁法可以为组织工程提供充足的种子细胞来源。Objective To explore the approaches of isolation culture and identify the rabbit bone marrow-derived mesenchymal stem cells in vitro. Methods BMSCs from rabbit were isolated, cultured and purified by the whole bone marrow adherence method, The cell growth was observed under Phase-contrast microscope and the growth curve of cell proliferation was drawn accordingly . BMSCs were induced to differentiate into osteoblasts and adipoeytes and detected Alkaline phosphatase activities of different time points. Results BMSCs from rabbits were spindle eell-based, showing radial colony arrangement. The growth curve were consistent with the growth characteristics and good activity of normal cells. Following induction, oil red 0 staining, alkaline phosphatase staining, you Kossa staining and alizarin red staining produced a strong reaction in cells. The osteogenesis induced electron microseopy tested positive. Conclusion Whole bone marrow adherence method is simple and can isolate, purify and amplify BMSCs in vitro. The obtained ceils have general biological characteristics of mesenchymal stem ceils, and also have potentiality of multi-directional differentiation. This experimental method has important practical significance to provide adequate source of seed cells for tissue engineering.

关 键 词：骨髓间充质干细胞 全骨髓贴壁法 细胞培养 兔 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]