三维支架中骨髓间充质干细胞及软骨细胞和脂肪源性成熟基质细胞的成软骨效应

背景：软骨修复的关键是种子细胞在三维支架中的定向分化，但细胞放入三维支架中很难有透明软骨表现。 目的：考察3种种子骨髓间充质干细胞，软骨细胞和脂肪源性成熟基质细胞在支架中的成软骨特性。 方法：抽取羊骨髓，胰酶消化后获得原代骨髓间充质干细胞细胞、软骨细胞和脂肪源性成熟基质细胞，单层培养扩增。取P1骨髓间充质干细胞，P3软骨细胞和P3脂肪源性成熟基质细胞种植入不同比例(10%，20%，50%，80%，100%)的胶原/透明质酸支架中, 在无血清培养液中三维培养。2周后，用SO染色和免疫组织化学染色分析，观察硫酸软骨素和Ⅱ型胶原合成情况。 结果与结论：种子细胞在三维支架中特定条件下有成软骨效应，骨髓间充质干细胞在含20%透明质酸的胶原/透明质酸支架扩增少于3代有成软骨效应，软骨细胞传代数更高仍然有成软骨效应，但脂肪源性成熟基质细胞成软骨效应能力较弱。结果提示，在同样条件下骨髓间充质干细胞、软骨细胞较脂肪源性成熟基质细胞有更好的成软骨性能。 关键词：骨髓间充质干细胞；软骨细胞；组织工程；软骨；脂肪干细胞 doi:10.3969/j.issn.1673-8225.2010.16.004

Chondrogenesis of bone marrow mesenchymal stem cells, chondrocytes and adipose-derived adult stromal cells in three-dimensional scaffolds

BACKGROUND: The key factor for cartilage repair is an orient differentiation of seed cells in three-dimensional scaffold; however, clear cartilage is hardly observed in the three-dimensional scaffold. OBJECTIVE: To investigate the chondrogenesis of bone marrow mesenchymal stem cells (BMSCs), chondrocytes (CCs) and adipose-derived adult stromal cells (ADAS) in three-dimensional scaffolds METHODS: BMSCs, CCs, and ADAS were primary-cultured from goat bone marrow following trypsin digestion and amplified in monolayer. P1 BMSCs, P3 CCs, and P3 ADAS were implanted into collagen/hyaluronic acid scaffold at different concentrations of 10%, 20%, 50%, 80%, and 100%, and the cells were then three-dimensionally cultured in serum-free culture media. Two weeks later, SO staining and immunohistochemical staining were employed to observe the synthesis between of chondroitin sulfate and type II collagen. RESULTS AND CONCLUSION: The seeded cells had chondrogenesis in scaffolds under certain conditions. In details, BMSCs in 20% (W/W) hyaluronan/collagen I scaffolds could perform a hyaline-cartilaginous phenotype when they were expanded with less than 3 passages and cultured in a chondrogenic medium, and CCs did better than BMSCs in chondrogenesis with high passage number; however, ADAS just showed a minor chondrogenesis even in a pellet culture. BMSCs and CCs were better than ADAS as seeded cells for cartilage tissue engineering and regenerative medicine.