Chondrogenic differentiation of bone marrow mesenchymal stem cells

Chen Liang; He Shao-ru; Zhuang Jian; Zheng Man-li; Sun Yun-xia; Liang Hui-xin; Liu Yu-mei; Sun Xin; Chen Xiao-bo

doi:10.3969/j.issn.2095-4344.2013.27.002

Abstract： BACKGROUND: Bone marrow mesenchymal stem cells are important seeded cells for construction of tissue-engineered trachea, but there is no special surface marker. Therefore, identification of bone marrow mesenchymal stem cells is mostly based on morphology, phenotype antigen and the function of differentiation. OBJECTIVE: To explore the feasibility of the tracheal chondrogenic differentiation of bone marrow mesenchymal stem cells under a special condition through isolation, cultivation and identification of bone marrow mesenchymal stem cells. METHODS: Rabbit bone marrow was acquired in the sterile environment to isolate and culture bone marrow mesenchymal stem cells to passage 2 by bone marrow adherence and screening method. Flow cytometry identified the phenotype CD44, CD45 of bone marrow mesenchymal stem cells at passages 1 and 2. Rabbit tracheal samples were acquired in the sterile environment, the tracheal chondrocytes were isolated and cultured by enzyme digestion, and toluidine blue staining was used to detect aggrecan. Bone marrow mesenchymal stem cells were co-cultured with tracheal chondrocytes by Transwel and transforming growth factor β1. Cel morphology was detected under an inverted microscope. Real-time quantitative PCR and toluidine blue staining detected the extracel ular matrix components, such as type Ⅱ col agen and aggrecan.RESULTS AND CONCLUSION: After isolation and culture, cells were spindle and irregular in morphology, and passaged cells thrived that were gathered into a fish-like colony growth. For passage 1 bone marrow mesenchymal stem cells, the positive rates of phenotype antigen CD44 and CD45 were respectively 96.97% and 13.72%; for passage 2 cells, the positive rates of phenotype antigen CD44 and CD45 were 99.11% and 8.54%, respectively. Tracheal chondrocytes were positive for toluidine blue staining. The morphology of induced bone marrow mesenchymal stem cells changed from long fusiform to triangular or irregular shape, indicating the chondrocytes expressed type Ⅱ col agen and aggrecan, and toluidine blue staining was positive. These results showed bone marrow adherence and screening method could acquire bone marrow mesenchymal stem cells, and the purity of passage 2 cells is higher. Under a special condition, bone marrow mesenchymal stem cells have the potential of chondrogenic differentiation, and can be selected as seed cells for construction of tissue-engineered trachea.