Multimodal Evaluation of Collagen-Based Biomimetic Scaffolds Enriched With Keratin and Hydroxyapatite: Integration of Serological, Immunological, and Histological Analysis in a Bilateral Osteochondral Rat Model

This study evaluates the regenerative potential of three types of collagen-based biomimetic scaffolds in osteochondral defects in Wistar rats. Formulations tested included: F1 (pure collagen - control), F5 (collagen with hydroxyapatite - for osteoconductivity assessment), and F6 (collagen with hydroxyapatite and keratin - for osteogenic and immunomodulatory potential). Using a bilateral osteochondral defect model, the research integrated serologic, immunologic, and histologic analysis over 60 days. All scaffolds showed excellent hemocompatibility ( 5% hemolysis). Cytokine profiles revealed that F6 reduced the pro-inflammatory cytokines (IL-6, IL-8, TNF-α) and increased the anti-inflammatory IL-10, indicating an environment favorable for regeneration. C-reactive protein levels were decreased in the F6 group, but the serum alkaline phosphatase was significantly increased, suggesting intense osteoblastic activity. Histological evaluation confirmed the superiority of the F6 scaffold, with high scores for cell density, collagen matrix organization, mineralization, and vascularization. The results demonstrate that the addition of keratin to the collagen-hydroxyapatite composite creates a superior scaffold that modulates the systemic immune response and promotes bone regeneration. These findings consolidate the further development of keratin-enriched biomimetic composites to act as scaffolds for regenerative strategies in orthopedic surgery.