BFGF And BMP-2Gene Transfection Of Osteoblast Function In Vitro Study Of Sheep BMSCs

Objective:To analyze bFGF and BMP-2gene alone and co-transfection of inter-vitro sheep BMSCs proliferation and bone effects of differentiation, more Collagen-Ⅰ, OC, OPN and other bone related genes in turn after transfection of mesenchymal stem cells expressing the relative difference between the amount for the construction of seed cells for bone tissue engineering to make a theoretical basis.Methods:The whole bone marrow culture method sheep BMSCs, build bFGF, BMP-2lentiviral vectors were transfected sheep BMSCs, BMSCs by flow cytometry transfection efficiency, will get bFGF group, BMP-2group, the combined group, control cells, the RNA was extracted using real-time quantitative PCR and mRNA and protein levels detected by Western-blot Collagen-Ⅰ, OC, OPN and other bone related genes relative expression changes.Results:(1) lentiviral vector transfected BMSCs transfection rate of95.4%, and bFGF and BMP-2were stable expression.(2) the level of mRNA, in the Collagen-Ⅰ and OC genes, bFGF main effects, BMP-2main effects, interaction effects were statistically significant (P<0.05). bFGF and BMP-2can promote BMSCs factor of two gene expression, and bFGF between the factor and the presence of BMP-2synergy. When combined gene transfection Collagen-Ⅰ and OC gene expression is higher than single gene transfection. But in the OPN gene, bFGF main effect was not statistically significant (P>0.05). Multivariate analysis showed, bFGF and BMP-2two factors could increase osteoblast-related gene mRNA expression, and there is a synergistic interaction (P<0.05).(3)the protein level, in the Collagen-Ⅰ OC and OPN protein, bFGF main effects, BMP-2main effects, interaction effects were statistically significant (P<0.05). bFGF and BMP-2two factors can promote BMSCs osteogenic protein-related gene proteins expression, the bFGF and BMP-2was synergistic factor respectively. When combined gene transfection Collagen-Ⅰ and OC protein expression is higher than single gene transfection. Multivariate analysis showed, bFGF and BMP-2two factors could increase the expression of osteoblast-related proteins and there is a synergistic interaction (P<0.05).Conclusion:In vitro experiments showed that bFGF and BMP-2has on osteoblast-related gene expression synergistic interactions and co-transfected group Collagen-Ⅰ, OC, OPN and other bone-related genes into mRNA and protein levels relative expression were the highest in the group of cells into bone powerful, suitable for use as seed cells for bone tissue engineering.