Synergistic effect of RhBMP-2 and bFGF on ectopic osteogenesis in mice

Objective:To investigate the synergistic effect and mechanism of the combined application of recombinant human bone morphogenetic protein-2(rhBMP-2) and basic fibroblast growth factor(bFGF).Methods:24 KM male mice were randomly divided into 6 groups with 4 mice in each group,namely,Group A(control group),Group B(only treated with collagen),Group C(treated with2 ng bFGF+collagen).Group D(treated with 4μg rhBMP-2+collagen),Group E(treated with 4 μg rhBMP-2+2 ng bFGF+collagen) and Group F(treated with 4 μg rhBMP-2+4 ng bFGF+collagem.The composites were implanted into the intermuscular septum of hind legs mice:whereas in control group,intermuscular septum of mice was separated and no implantation was performed.General observation,detection of concentration of calcium content,micro computed tomography(Micro-CT).three-dimensional reconstruction scan.measurement of bone mineral density(BMD).bone volume fraction(BVF) and trabecular thickness(Tb.Th).as well as histological observation with HE staining and ALP and CD34 immumohistochemical staining were performed.Results:Ectopic osteogenesis was found in Groups D.F and F mice.The difference in concentration of calcium contents was statistically significant between Groups D and E(P<0.05),but insignificant between Groups E and F(P>0.05).Micro-CT and three-dimensional reconstruction revealed continuous newborn bone substance in external surface of ectopic bone formation,and the center of bone formation did not show obvious substantial filling by bone substance.The differences in BMD,BVF and Tb.Th were statistically significant between Groups D and E or F(P<0.01 or <0.05).HE staining showed that in Groups D.E and F.newborn bone substance was mainly located at the edge of ectopic bone formation,and the bone formation in Groups E and F was better than that in Group D.ALP and CD34 immumohistochemical staining revealed the positive expression mainly at the edge of ectopic bone formation,and area of positive expression in Groups E and F was larger than that in Groups D.Conclusions:rhBMP-2 possesses the capacity to induce ectopic osteogenesis independently,but bFGF does not have this ability;the combined application of rhBMP-2 and bFGF can enhance the synergetic effect on inducing ectopic osteogenesis.