脱钙骨基质材料的生物学表现:降解性能、孔隙率及其黏附性能特征

Biological characteristics of demineralized bone matrix: Degradation capacity, interval porosity and adhesion rate

目的:体外观察脱钙骨基质的降解性能及孔隙率,同时采用兔骨髓间充质干细胞与脱钙骨基质在体外复合培养,了解兔骨髓间充质干细胞对这种天然材料的黏附能力。方法:实验于2005-01-08/03-15在兰州大学骨科研究所进行。取6月龄的青紫蓝兔1只,麻醉后处死,取四肢骨干骺端及椎体松质骨,采用Urist提供的方法制作脱钙骨基质,将脱钙骨基质置于磷酸盐缓冲溶液中12周测定其降解率;用液体置换法测其孔隙率;用细胞计量法测定生长良好的浓度为1×108L-1的第3代骨髓间充质干细胞与脱钙骨基质复合培养6h后的黏附率。结果:脱钙骨基质的降解随时间的延长逐渐加速,8周前,其降解速率较慢,8周后,其降解速度明显加快,完全降解需要10￣12周,与骨髓间充质干细胞的增殖规律近似;所测脱钙骨基质孔隙率为(77.15±3.44)%;1×108L-1的第3代骨髓间充质干细胞与脱钙骨基质的平均黏附率为71.25%。结论:脱钙骨基质的降解曲线与骨髓间充质干细胞的增殖曲线相一致,具备良好的孔隙率和黏附率,提示脱钙骨基质可能为软骨组织工程比较理想的生物支架材料。

AIM： To observe the degradation capacity and interval porosity of demineralized bone matrix （DBM）, and investigate the adhesion of rabbit mesenchymal stem cells （MSCs） onto DBM in vitro. METHODS： The experiment was conducted in Institute of Orthopaedics of Lanzhou University from August 1^st to March 15^th 2005. One Chinchilla Rabbit of 4-6 months old was selected and executed under anesthesia to harvest the bone metaphysis of four limbs and cancellous bone. Following the method described by Urist, allogenic DBM was made and soaked into phosphate buffered solution （PBS） for 12 weeks to determine its degradation capacity; liquid replacement method was used to test its interval porosity; the adhesion rate of MSCs on DBM was tested using cytometry after the 3^rd passage MSCs at concentration of 1×10^8L^-1 were co-cultured with DBM in vitro for 6 hours. RESULTS： The degradation rate of DBM was accelerated with the prolonging of degradation time. The degradation rate was slow within 8 weeks, and significantly accelerated afterward. The degrading time was about 10-12 weeks, which was similar to the proliferation rules of MSCs. The porosity rate of DBM was （77.15±3.44）%; the average adhesive rate of the 3^rd passage cells （1×10^8L^-1） onto DBM was 71.25%. CONCLUSION： The degrading curve of DBM is in accordance with the proliferation curve of MSCs, and DBM has satisfactory adhesion capacity and interval porosity, indicating that it is an ideal scaffold material for cartilage tissue engineedng.