3D scaffold with VEGF/FGF9 conjugated fibrin, Nano Calcium Sulfate and BMP2 genetically engineered mesenchymal stem cells promotes vascularized bone formation
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Introduction: Bone defects including bone fractures are increasing social and medical problems in the world, which dramatically decreased overall health and quality of life and caused huge medical costs of nearly $20 billion per year in United State. Therefore there is an urgent need for bone reconstruction. Bone tissue engineering has been extensively studied as an alternative to overcome clinical disadvantages of current bone grafts. In our previous study, we developed an injectable nano scale calcium sulfate combined with alginate paste (nCS) to support the delivery and growth of BMP2 genetically engineered mesenchymal stem cells (nCS+M/B2), which significantly repaired bone defects. Because effective bone formation, especially for critical size bone defects, also requires vascular intervention. Materials & Methods: In this study, we developed a new fibrin gel system, in which angiogenic factor VEGF, FGF9 or both were conjugated to fibrin to increase their stability meanwhile slow down the release. To investigate whether this system combined with nCS+M/B2 would synergistically promote vascularized bone regeneration, nCS+M/B2 was coated with VEGF- or/and FGF9- conjugated fibrin to form a sandwich-like 3D scaffold, and then subcutaneously implanted into mice to test the osteogenic and angiogenic activity. MicroCT, alkaline phosphatase activity assay and histological analysis were used to evaluate bone formation while immunostaining were employed to indicate newly formed vascular. Result: Coating with fibrin greatly preserved the nCS+M/B2 scaffold structure and also promoted de novo bone formation. Conjugated FGF9 alone did not significantly elevate ectopic bone formation, however, FGF9 combined with VEGF significantly promoted vascularized bone formation. We also compared different ways to load M/B2 and found that the group equally loaded M/B2 to fibrin gel and nCS showed best bone formation compared with the groups loaded a same amount of M/B2 to fibrin or nCS only, suggesting M/B2 may have different functions in nCS and fibrin. Conclusion: Overall, our results suggested that integration of nCS+M/B2 (support bone formation) with fibrin-based VEGF/FGF9 release system (support vascular formation) is an innovative strategy to improve bone regeneration.