| Objective: To investigate the method of inducing Bone marrow-derived mesenchymal stem cells (BMSCs) to diffrenciate into ligament fibroblast (LF) by basic fibroblast growth factor (bFGF) and bone morphogenetic protein 12 (BMP12) gene co-modification. Methods: (1) Isolate and culture the BMSCs and LF, and distinguish the two cell types in cytology and molecular biology; (2) Construct, identify and proliferate the recombinant adenovirus vectors containing the bFGF and BMP12 genes separately. Enhanced green fluorescent protein (eGFP) is used for the report gene in each adenovirus vector. (3) Observe the secretion of the protein of bFGF and BMP12 after transfection, and the cell proliferation, differentiation, and production of specific ligament fibroblast extracellular matrix proteins of the BMSCs after Ad.bFGF-eGFP and Ad.BMP12-eGFP co-transfection. Results: (1) We successfully isolated and cultured the BMSCs and LF. The two kinds of cells were similar on the general cell morphology, but significantly different in the cell proliferation, ultrastructure, cell phenotype and cell molecular biology. (2) We successfully constructed and proliferated the recombinant adenovirus vectors Ad.bFGF-eGFP and Ad.BMP12-eGFP. (3) After co-transfection by Ad.bFGF-eGFP and Ad.BMP12-eGFP, bFGF and BMP12 proteins expressed significantly in the BMSCs and greatly promoted the cell proliferation and induced the BMSCs to differentiate into LF obviously. After co-transfection, secretion of the specific ligament fibroblast extracellular matrix proteins by the BMSCs was significantly promoted. Conclusions: After co-modification by Ad.bFGF-eGFP and Ad.BMP12-eGFP, the BMSCs could significantly differentiate into LF phenotypes and show significantly enhanced ability of secreting the specific ligament fibroblast extracellular matrix proteins. We concluded that the BMSCs co-modified by bFGF and BMP12 genes could become a kind of new better seeding cells in ligament tissue engineering.
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