Vasculo-osteogenic keratin-based nanofibers containing merwinite nanoparticles and sildenafil for bone tissue regeneration

Vascularization of bone tissue constructs plays a pivotal role in facilitating nutrient transport and metabolic waste removal during the processes of osteogenesis and bone regeneration in vivo. In this study, a sildenafil (Sil)-loaded nanofibrous scaffold of keratin/Soluplus/merwinite (KS.Me.Sil) was fabricated through electrospinning and the effectiveness of the scaffold was assessed for bone tissue engineering applications. The KS.Me.Sil nanofibrous scaffold exhibited notably enhanced ultimate tensile strength (3.38 vs 2.61 MPa) and elastic modulus (69.83 vs 46.27 MPa) compared to the KS scaffold. The in vitro release of Ca²⁺, Si⁴⁺ and Mg²⁺ ions and the release of Sil from the nanofibers as well as biodegradability and bioactivity were evaluated for 14 days. Protein adsorption capability and cytocompatibility of the scaffolds were tested. Alkaline phosphatase activity test, Alizarin red staining and qRT-PCR analysis demonstrated that the KS.Me.Sil nanofibers had the best osteogenic activity among other samples. Also, the results of the chorioallantoic membrane assay showed an almost threefold increase in blood vessel density in the group treated with the KS.Me.Sil nanofibers extract compared to the KS. In conclusion, our findings suggest that the electrospun KS.Me.Sil nanofibrous scaffold offers a robust structure with exceptional osteogenic and angiogenic characteristics, making it a promising candidate for bone tissue engineering applications.