可注射型生物蛋白胶包埋骨髓基质细胞工程化组织的体外实验

Tissue-Engineered Bone Marrow Stromal Cells Embedded with Injectable Fibrin Glue in Vitro

目的构建可注射型生物蛋白胶包埋骨髓基质细胞的工程化组织,体外培养并研究其生物学特性,探讨将可注射型生物蛋白胶作为组织工程支架用于临床的实验基础。方法体外培养浇铸有骨髓基质细胞的生物蛋白胶,通过倒置相差显微镜、激光共聚焦显微镜观察载体内细胞生长及载体降解情况,5-溴脱氧尿苷(5-Bromodeocyuridine,BrdU)掺入标记后免疫组化等方法研究可注射型载体内包埋细胞的增殖情况。结果骨髓基质细胞包埋于生物蛋白胶内能很好地存活并增殖,2d后细胞呈典型的成纤维细胞形态;6d后生物蛋白胶边缘部分开始降解,细胞脱落至培养板;体外培养14d,细胞生长良好,大部分生物胶降解,脱落的细胞增多,贴壁生长的细胞形态正常;3周后生物蛋白胶完全降解。结论生物蛋白胶聚合后包埋的种子细胞能够正常增殖,生物蛋白胶是一种理想的适用于微创方法修复组织的可注射型组织工程培养和移植的支架。

Objective To construct tissue-engineered graft with bone marrow stromal cells （BMSCs） imbedded in the polymerized fibrin biopolymer scaffold in vitro, observe its biological characteristics and degradation in vitro, and explore the biological basis of using injectable fibrin glue as a scaffold in tissue engineering by minimal invasive surgery. Methods BMSCs embedded with injectable fibrin glue were polymerized and cultured in vitro. Its biological characteristics, including gel formation, cell growth, histological features, and degeneration time of the fibrin glue were observed under an inverted phase contrast microscope and laser confocal microscope. 5-Bromodeocyuridine （Brdu） incorporation, frozen section and immunohistochemical staining were employed to study the growth and proliferation of the BMSCs, and the degradation of the fibrin glue. Results The BMSCs embedded in the fibrin glue grew well, homogeneously distributed in the polymerized fibrin glue. Brdu incorporation and immunohistochemical staining showed that the BMSCs embedded in the fibrin glue could also proliferate well. Two days after the embedment, the cells with typical fibroblast-shaped morphology could be observed. The edge of the fibrin glue began to degrade six days later and the cells fell off to the culture template. After fourteen days, most of the fibrin glue degraded and the adherent cells grew well. The polymerized fibrin glue degraded completely after 3 weeks. Conclusions Seeding cells embedded in polymerized fibrin glue can grow and proliferate well. Fibrin glue is an ideal injectable scaffold for embedding BMSCs for culture and transplantation by minimally invasive surgery method.