骨髓间充质干细胞在软骨细胞培养条件下生物学特点的观察

Biological characteristics of rabbit marrow-derived mesenchymal stem cells cultured in condition for chondrocytes

目的 观察骨髓间充质干细胞 (Mesenchymalstemcells ,MSCs)在软骨细胞培养条件下传代培养的主要生物学特性。方法 梯度离心分离幼兔骨髓有核细胞 ,培养分离MSCs并进行传代培养 ,观测在体外软骨细胞培养条件下MSCs形态、生长特点及Ⅰ型和Ⅱ型胶原、蛋白多糖聚集体合成分泌以及碱性磷酸酶活性等方面的变化。结果 ①MSCs原代细胞呈可见的均匀分布的簇状生长 ,呈长梭形 ,在传代培养中形态特性无明显变化、均质性明显提高 ;②各代Ⅰ型胶原免疫组化均为阳性 ,Ⅱ型胶原免疫组化均为阴性。③在各代培养中碱性磷酸酶染色均为阴性 ,对甲苯胺蓝存在极弱的异染性反应 ;④细胞外基质中硫酸糖胺多糖含量亦很低。结论 传代培养中骨髓MSCs保持了常规低糖培养条件下的基本生物学特点 ,说明常用的软骨细胞基础培养条件同样适合骨髓MSCs的培养 .

Objective To observe the main biological characteristics of marrow-derived mesenchymal stem cells (MSCs) during the passage cultivation under the condition for chondrocyte culture. Methods The marrow-derived MSCs were isolated from young rabbits and cultivated in the condition for culturing chondrocytes. The changes of morphology, growth and proliferation, synthesis of collagen typeⅠ and Ⅱ and aggrecan, and activity of alkaline phosphatase (ALP) were observed during the passage cultivation of MSCs. Results 1) Primary MSCs proliferated in visible symmetric colonies with long-spindle shape. The morphological characteristics of marrow-derived MSCs had no change during passaging, and its homogenous rose with passaging. 2) Every passage cells showed positive staining of collagen typeⅠ, and negative of collagen typeⅡ. 3) The cells had negative staining of ALP, and weak heterochromatic to toluidine blue. The content of sulfate glycosaminoglycans (GAG) was very little in extracellular matrix. Conclusion The biological characteristics of MSCs in the basic condition for culturing chondrocytes are the same with in the regular low-glucose condition for culturing MSCs.