| Objective: Deerhorn glue to Lentiviral vectors carrying h BMP7 gene infection BMSCs of rat of the proliferation and osteogenesis of sample transformation, thereby providing theoretical implications for seed cells in the construction of tissue-engineered bone.Methods: ① Extract and cultivate rat BMSCs in vitro, identified by cell surface morphology and specific markers of cells.②The expression lentivirus vectors carrying BMP7 gene were constructed and transfection in BMSCs. BMP7 protein and m RNA levels was detected by RT-PCR and Western blotting;③Cells were divided into four groups: control group, deerhorn glue group, BMP7 group and combined group. Comparison groups of cell proliferation, alkaline phosphatase, Osteocalcin, calcification in nodule and real-time quantitative PCR detection BMSCs into osteoblasts- related gene expression level.Results: We successfully purified and proliferated BMSCs with high cell viability and differentiation ability. Fibroblast-like cells were harvested;Successfully builded h BMP7 gene with Lentiviral vectors PLV-sf GFP(2A)-BMP7, and through the infection virus particles containing h BMP7 genes get high BMP7 expression of BMSCs;Added deer horn glue to medium and overexpression of BMP7 BMSCs cells promotes proliferation of BMSCs and enhance the activity of alkaline phosphatase in the cell, improves the content of osteocalcin, enhance the collagen typeⅠ, integrinα2, integrinβ1 levels of m RNA.Conclusion: Deerhorn glue joint rat BMSCs transfected by h BMP 7 gene has dramatically strengthen their ability to differentiate into osteoblasts..Compared with single gene transfection induced and antler glue induced alone, the combined group cells of osteogenesis is strongest and is suitable for use as seed cells of tissue engineered bone. |
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