Abstract
Bone tissue engineering (BTE) aims to regenerate the damaged or diseased bone by combining cells, growth factors, and biomaterials. Synthetic bone tissue, which is widely available, is a promising alternative to autografts, allografts, and natural fillers. Biomaterials like 3D scaffolds mimic the extracellular matrix, supporting cell growth and tissue regeneration. However, clinical applications face challenges such as limited osteogenic and angiogenic capabilities, weak mechanical strength, and risks of infection and immune reactions. The current work introduces a mechanically robust cryogel-hydrogel hybrid scaffold, loaded with extracellular vesicles (EVs) and cell secretomes as osteoinductive cues. The scaffold’s mechanical strength was assessed against its individual components. Its physical characteristics, including swelling capacity and porous structure, were analyzed using field emission scanning electron microscopy (FESEM), and the incorporation of nano-hydroxyapatite (nHAp) was confirmed through Fourier transform infrared (FTIR) spectroscopy. Three scaffolds were tested: a cryogel scaffold of gelatin and nano-hydroxyapatite (nHAp), a hydrogel scaffold of poly(ethylene glycol) diacrylate (PEGDA), and a hybrid cryogel scaffold of gelatin and nano-hydroxyapatite with a PEGDA interpenetrating network (IPN). The results indicated that the hybrid cryogel scaffold with IPN had the greatest normalized ALP content and lowest cytotoxicity, making it the best choice for further study. To explore the osteoinductive potential of Mesenchymal Stem Cell-derived EVs, three scaffold variations were tested: a hybrid IPN scaffold, IPN + nHAp scaffold, and IPN + nHAp + EVs scaffold. These were evaluated for viable cell count, ALP activity, DNA content, and calcium accumulation. The IPN + nHAp + EVs scaffold demonstrated the greatest potential for BTE, with the highest mechanical strength—20.82-fold greater than the single-network cryogel and 1.75-fold stronger than the PEGDA hydrogel. It also showed the highest normalized ALP content, 3.39-fold and 1.47-fold greater than the IPN and IPN + nHAp scaffolds, respectively, and the highest calcium deposition, 1.30-fold higher than the IPN + nHAp scaffold. In conclusion, this study successfully developed a hybrid scaffold system with improved mechanical characteristics and osteogenesis potential, advancing the field of bone tissue engineering.
Published Version
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