转化生长因子β1诱导骨髓间充质干细胞向软骨细胞分化

Differentiation of bone marrow mesenchymal stem cells into chondrocytes induced by transforming growth factor beta 1

背景:转化生长因子β1是关节软骨组织工程研究的首选生长因子,适当浓度能刺激关节软骨细胞增殖、分裂和分化。目的:建立含有转化生长因子β1的特殊诱导体系培养条件下骨髓间充质干细胞分化为软骨细胞的能力,观察诱导后的细胞形态及表型变化。方法:于兔胫骨结节内侧抽取骨髓,采用贴壁培养法分离纯化兔骨髓间充质干细胞,取第3代骨髓间充质干细胞行流式细胞仪检测鉴定其表面抗原,以含有转化生长因子β1的特殊软骨诱导体系的培养条件下对第3代骨髓间充质干细胞诱导培养21d,诱导后与人鼻中隔软骨细胞进行比较。采用免疫组织化学法对Ⅱ型胶原进行定性检测。结果与结论:贴壁培养法可分离并纯化兔骨髓间充质干细胞,所得第3代骨髓间充质干细胞表面抗原CD44阳性,CD34、CD45阴性。经诱导培养21d后细胞形态变为不规则,Ⅱ型胶原免疫组织化学染色显示可见阳性细胞。提示含有转化生长因子β1的特殊软骨诱导体系的培养条件下,骨髓间充质干细胞可以转化为软骨细胞,且与正常软骨细胞无明显差异。

BACKGROUND： Transforming growth factor beta 1 （TGF-β1） is a first selected growth factor in study of articular cartilage tissue engineering. Suitable concentration can stimulate proliferation, division and differentiation of articular chondrocytes. OBJECTIVE： To establish an special inducing system for differentiation of bone marrow mesenchymal stem cells （BMSCs） into chondrocytes in contained TGF-β1 culture in vivo,and to observe the changes in cell form and phenotype. METHODS： BMSCs were separated and purified from rabbit tibial tubercle using attachment culture method. Surface antigen of the third-generated BMSCs was determined by using flow cytometry. Subsequently,the third-generated BMSCs were induced with TGF-β1-contained special inducing system in 21-day culture, which was compared with human septal cartilage cells of nose following induction. Collagen type Ⅱ was qualitatively determined using immunohistochemical method. RESULTS AND CONCLUSION： Rabbit BMSCs were separated and purified using attachment culture method. The third-generated BMSCs were positive for surface antigen CD44,but negative for surface antigen CD34 and CD45. Cell form was irregular 21 days after induction. Immunohistochemical staining for type II collagen demonstrated positive cells. Results suggested that under culture system containing TGF-β1 BMSCs may directionally differentiate into chondrocytes,and there were not significant differences with normal chondrocytes.