构建组织工程骨与生物蛋白胶双相接种法的实验研究

目的观察采用生物蛋白胶（fibrin glue,FG）双相接种技术促进组织工程骨体外构建的可行性。方法以生物蛋白胶为介质,用双相接种技术将羊骨髓间充质干细胞（mesenchymal stem cells,MSCs）与同种异体冻干脱钙骨基质（freeze-dried demineralized bone matrix,FDBM）复合,在体外构建组织工程骨,以静置接种法作为对照。通过细胞计数、光学和电子显微镜及四甲基偶氮唑盐比色法（methyl thiazolyl tetrazolium,MTT）检测观察细胞在支架材料上的粘附、生长情况。结果双相接种法在不同接种密度时都可保持细胞上架率维持在80%左右,上架细胞的数量随接种细胞密度增加成比例增加;FG对细胞的增殖没有明显影响,提高接种密度使细胞生长曲线提前达到平台期,各组的细胞数量峰值接近。结论双相接种技术可显著提高种子细胞的上架效率,可以作为一种体外构建组织工程骨的好方法。

Enhancing Tissue Engineering Bone Formation in vitro by Biphasic Seeding Technique with Fibrin Glue

Objective To study the feasibility of enhancing tissue engineering bone formation in vitro with biphasic seeding technique.Methods Different densities of goat mesenchymal stem cells （MSCs） were seeded onto allogenic freeze-dried demineralized bone matrix （FDBM） introduced by fibrin glue and cocultured in vitro.FDBM was soaked in cell suspension for 4 hours without fibrin glue in the control group.The adhesion and proliferation of MSCs were evaluated by cell counting,microscope,electronic microscope and methyl thiazolyl tetrazolium （MTT） test.Results With the biphasic seeding technique,the adhesive rates of different seeding densities were kept around 80%,the number of cells upon the scaffolds increased following the increase of seeding densities.Fibrin glue had no obvious impact on the cell proliferation.Increasing the seeding densities made growth curves arrived at the platform phase earlier.The peak number of cells in all groups were close.Conclusion Biphasic seeding technique can significantly increase the adhesive rates of MSCs seeding on the FDBM scaffolds,and may be a suitable method to form tissue engineering bone in vitro.