ZIF-67-anchored halloysite nanotubes infused in chitosan hydrogel for enhanced bone tissue engineering

Despite the intrinsic regenerative capacity of bone, there remains a clinical need for advanced biomaterials that can simultaneously accelerate osteogenesis and promote vascularization. To address this, a chitosan (CS)-based hydrogel film was developed by incorporating zeolitic imidazolate framework-67 (ZIF-67) anchored functionalized halloysite nanotubes (fHNTs) using a simple, low-cost, solution casting technique. Structural analyses using X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) validated the successful synthesis of the CS/ZIF-67/fHNT hydrogel films. Moreover, morphological evaluations revealed a uniform dispersion of ZIF-67/fHNT nanocomposites within the matrix. Increasing the fHNT content markedly enhanced the mechanical strength and hydrophilicity of the hydrogel films, while also providing controlled degradation behaviour, highlighting the tunability and robustness of the hydrogel film. The hydrogel film also facilitated controlled cobalt ion release, bioactive apatite formation, and exhibited excellent hemocompatibility, collectively demonstrating its functional effectiveness. In vitro studies exhibited robust biological performance, showing support for osteoblast viability, along with enhanced osteogenesis evidenced by elevated alkaline phosphatase (ALP) activity, significant upregulation of osteogenic genes (RUNX2, IBSP, OPN, COL1A1, OSX, and BGLAP), and increased calcium and collagen deposition. Importantly, the hydrogel film effectively promoted angiogenic activity, as demonstrated by enhanced scratch migration, improved tube formation in HUVECs, and notable effects in chorioallantoic membrane (CAM) assay. By integrating both osteogenic and angiogenic stimulation within a single construct, the CS/ZIF-67/fHNT hydrogel film demonstrates a synergistic advantage, positioning it as an effective biomaterial for advanced bone tissue engineering applications.