Injectable Thymosin β4-Modified Hyaluronic Acid Hydrogel with Exosomes for Stem Cell Homing and Neuronic-Angiogenic-Osteogenic Coupled Cranial Repair

Accelerating angiogenesis, neurogenesis, and in situ stem cell recruitment at the site of bone defects is critical for bone regenerative repair. Bone marrow mesenchymal stem cell (BMSC) exosomes are cell-free therapeutic agents with bone-enhancing effects. Thymosin β4 (Tβ4) is a short peptide known for its key role in tissue repair and angiogenesis. In this study, we successfully developed a multifunctional injectable Exo@Tβ4/HAMA hydrogel platform by grafting Tβ4 onto methylmalonic anhydride-modified hyaluronic acid (HAMA) via photo-cross-linking and then encapsulating BMSC-derived exosomes. In vitro results demonstrated that the Exo@Tβ4/HAMA hydrogel exhibited improved mechanical properties, favorable biocompatibility, and the ability to significantly recruit BMSCs. Additionally, it showed superior vasculogenic effects on HUVECs and osteogenic differentiation potentials on BMSCs. In vivo studies revealed that the hydrogel successfully promoted both neurogenesis, angiogenesis, and new bone formation. It also facilitated osteogenesis through the ERK1/2-dependent RUNX2 signaling pathway. Our results suggest that this hydrogel platform exerts a robust multisystemic regulatory effect, fostering rat bone repair through the synergistic promotion of in situ stem cell recruitment, angiogenesis, neurogenesis, and osteogenesis. As a simple-to-prepare and multifunctional integrated bone graft, this hydrogel platform holds a significant promise in establishing a conducive microenvironment for optimal bone healing.