蒙古羊骨髓间充质干细胞的分离培养及多向诱导分化

Isolation,Culture and Multiple Differentiation Induction of Mongolia Sheep Bone Marrow Mesenchymal Stem Cell

本研究旨在建立蒙古羊骨髓间充质干细胞(bone marrow mesenchymal stem cell,BMSC)体外分离、纯化、增殖方法,诱导其向脂肪细胞、成骨细胞及软骨细胞分化。穿刺法抽取蒙古羊骨髓组织,采用密度梯度离心法结合贴壁培养法分离纯化BMSC,增殖,并测定其生长曲线及细胞倍增时间。对第3代蒙古羊BMSC进行成脂、成骨及成软骨诱导,观察诱导后的细胞形态。采用油红O、茜素红、HE等染色法在组织学水平对BMSC进行成脂、成骨和成软骨分化鉴定。结果显示,蒙古羊BMSC呈现均一梭形成纤维细胞样生长,生长曲线呈S型,生长增殖能力良好。在不同分化诱导之后,细胞呈现脂肪细胞、成骨细胞及软骨细胞的表型特征。表明获得的蒙古羊BMSC具有多向分化潜能,所分离细胞确为骨髓间充质干细胞。

To explore the optimal methods of isolating, puring, proliferating of Mongolia sheep bone marrow mesenchymal stem cells （BMSC）, and their multiple differentiation potentiality. Bone marrow （BM） was collected from one 30-day-old sheep by puncture method. BMSC were harvested by gradient centrifuge and adhesive culture methods. Growth curves and cells double time of passaged BMSC were determined. The P3 BMSC were cultured in vitro under adipogenic, osteogenie and chondrogenic inductive environments and induced into adipocytes, osteoblates, chrondrocytes. Their adipogenic, osteogenie and chondrogenic differentiation properties were confirmed by histological staining such as oil red,Alizarin Red, HE. The Mongolia sheep BMSC were uniform spindle-shaped in appearance, showed active proliferative capacity and had S shape of growth curve. They could he induced to differentiate into adipocytes,osteoblasts and chondrocytes under different induction medium. The Mongolia sheep BMSC showed strong proliferation and multiple differentiation potential. It proved that the cells isolated were indeed the bone marrow mesenchymal stem cells.