Injectable graphene oxide-PEG/collagen hydrogel enables controlled Atsttrin/TNFR2 signaling and 14-3-3ε-dependent bone regeneration under diabetic conditions

Atsttrin, a therapeutic derivative of progranulin, promotes tissue repair via TNFR2-mediated anti-inflammatory and regenerative signaling. Our group developed an injectable, thermoresponsive graphene oxide-polyethylene glycol/collagen type I hydrogel for sustained and localized delivery of Atsttrin and systematically evaluated its therapeutic efficacy in vitro and in vivo models. The hydrogel demonstrated favorable rheological properties, controlled degradation, high porosity, and sustained Atsttrin release over seven weeks. In vitro cytocompatibility assays confirmed its non-cytotoxic nature, which was further corroborated by long-term in vivo safety assessments indicating the absence of organ toxicity up to three months post-administration. Functional studies using wild-type and 14-3-3ε knockout bone marrow-derived mesenchymal stem cells revealed that Atsttrin significantly enhanced chondrogenic and osteogenic differentiation in WT cells, effects that were markedly reduced in KO cells. Hydrogel-encapsulated Atsttrin elicited more robust regenerative responses compared to administration of free Atsttrin, underscoring the critical role of controlled release in enhancing therapeutic efficacy. In vivo, Atsttrin-loaded hydrogels significantly accelerated fracture healing in both WT and diabetic mice, with diminished efficacy in 14-3-3ε-deficient models. Collectively, these results support an important role for 14-3-3ε in mediating the regenerative effects of Atsttrin and demonstrate the potential of the GO-PEG/Collagen I hydrogel as a localized delivery platform for bone repair.