Bone regeneration and natural repair are long-standing processes that can lead to uneven new tissue growth. By introducing scaffolds that can be autografts and/or allografts, tissue engineering provides new approaches to manage the major burdens involved in this process. Polymeric scaffolds allow the incorporation of bioactive agents that improve their biological and mechanical performance, making them suitable materials for bone regeneration solutions. The present work aimed to create chitosan/beta-tricalcium phosphate-based scaffolds coated with silk fibroin and evaluate their potential for bone tissue engineering. Results showed that the obtained scaffolds have porosities up to 86%, interconnectivity up to 96%, pore sizes in the range of 60–170 μm, and a stiffness ranging from 1 to 2 MPa. Furthermore, when cultured with MC3T3 cells, the scaffolds were able to form apatite crystals after 21 d; and they were able to support cell growth and proliferation up to 14 d of culture. Besides, cellular proliferation was higher on the scaffolds coated with silk. These outcomes further demonstrate that the developed structures are suitable candidates to enhance bone tissue engineering.

The eggshell membrane (ESM), resembling the extracellular matrix (ECM), acts as a protective barrier against bacterial invasion and offers various biofunctions due to its porous structure and protein‐rich composition, such as ovalbumin, ovotransferrin, collagen, soluble protein, and antimicrobial proteins. However, the structure of ESM primarily comprises disulfide bonds and heterochains, which poses a challenge for protein solubilization/extraction. Therefore, the method of dissolving and extracting bioactive protein components from ESM has significant potential value and importance for exploring the reuse of egg waste and environmental protection. In this study, soluble ESM proteins (SEPs) were extracted from conventional (industrial‐fed) and organic (free‐grounded) using an acidic 3‐mercaptopropionic acid (3‐MPA) extraction strategy. FTIR was employed to monitor the chemical changes in the ESM, while LC–MS/MS was used to conduct the proteomic analysis. The biocompatibility and effects of SEP cocktails on ECM synthesis were also investigated. The results indicated that the acidic 3‐MPA strategy effectively altered the ESM chemical composition, thereby facilitating SEPs extraction. The SEPs from conventional and organic eggs have different protein profiles but with partial overlapping. SEPs from both sources showed similar desirable biosafety profiles and dose‐dependent promotion of osteoblastic (ECM) component synthesis, suggesting that different egg sources may contribute to consistent core biological functions of protein products, they may also introduce different functional priorities.

Osteochondral defects present significant challenges for effective tissue regeneration due to the complex composition of bone and cartilage. To address this challenge, this study presents the fabrication of hierarchical scaffolds combining chitosan/β-tricalcium phosphate (β-TCP) to simulate a bone-like layer, interconnected with a silk fibroin layer to mimic cartilage, thus replicating the cartilage-like layer to mimic the native osteochondral tissue architecture. The scaffolds were produced by freeze-drying and then crosslinking with genipin. They have a crosslinking degree of up to 24%, which promotes a structural rearrangement and improved connection between the different layers. Micro-CT analysis demonstrated that the structures have distinct porosity values on their top layer (up to 84%), interface (up to 65%), and bottom layer (up to 77%) and are dependent on the concentration of β-tricalcium phosphate used. Both layers were confirmed to be clearly defined by the distribution of the components throughout the constructs, showing adequate mechanical properties for biomedical use. The scaffolds exhibited lower weight loss (up to 7%, 15 days) after enzymatic degradation due to the combined effects of genipin crosslinking and β-TCP incorporation. In vitro studies showed that the constructs supported ATDC5 chondrocyte-like cells and MC3T3 osteoblast-like cells in duo culture conditions, providing a suitable environment for cell adhesion and proliferation for up to 14 days. Overall, the physicochemical properties and biological results of the developed chitosan/β-tricalcium phosphate/silk fibroin bilayered scaffolds suggest that they may be potential candidates for osteochondral tissue strategies.