Bone regeneration remains a major clinical challenge, requiring advanced biomaterials capable of guiding tissue healing. This study investigates fibrin-based composites incorporating peptide amphiphiles (PAs) engineered to co-assemble with fibrin and present binding epitopes for Bone Morphogenetic Protein-2 (BMP-2) and Transforming Growth Factor Beta-1 (TGF-β1). These materials are designed to locally sequester growth factors, thereby modulating cellular responses involved in bone formation. Their performance was evaluated in vivo using a rat drill-hole defect model, alongside in vitro studies aimed at assessing their influence on osteogenic differentiation and exploring their potential role in different ossification pathways.
Fibrinogen solutions were combined with negatively charged PAs, including unmodified, BMP-2-binding, TGF-β1-binding, and dual-binding variants, followed by thrombin-induced polymerisation. In vivo experiments (approved by the Cantonal Ethics Committee of the Canton of Grisons) involved the creation of a 1.5 mm diameter, 3 mm deep unicortical defect in the proximal tibia of female Wistar rats. Defects were treated with PA-based materials, while controls included empty defects and fibrin-only constructs. Healing was assessed after six weeks using micro-computed tomography and histological analysis. In vitro, human bone marrow stromal cells (BMSCs) were used to evaluate osteogenic differentiation. Early osteogenic activity was assessed through alkaline phosphatase (ALP) quantification, while chondrogenic matrix production was evaluated via glycosaminoglycan (GAG) quantification. Gene expression analysis of osteogenic and hypertrophic markers, including COL10A1 and RUNX2, was performed, and matrix deposition and mineralisation were analysed at later time points.
All materials supported bone healing in vivo. While cortical healing was comparable across groups, BMP-2-binding composites demonstrated a slight improvement in trabecular bone regeneration compared to controls. In contrast, TGF-β1-binding formulations showed reduced healing scores, suggesting that sequestration of this growth factor may limit its bioavailability in vivo. In vitro results showed that all fibrin-based materials supported osteogenic differentiation of BMSCs under intramembranous conditions, with no significant differences observed between groups. Competitive ELISA confirmed that PAs functionalised with TGF-β1-binding epitopes effectively interact with the active form of the growth factor. Ongoing work is further investigating the role of these materials in endochondral ossification pathways.
Overall, fibrin–PA composites represent a versatile platform for localised growth factor delivery and modulation of bone regeneration. The findings suggest that BMP-2 sequestration may enhance regenerative outcomes, while TGF-β1 binding may reduce effective signalling and delay trabecular repair. These results highlight the importance of controlled growth factor presentation and provide a foundation for future studies aimed at tailoring biomaterials to specific bone healing mechanisms and clinical applications.