Hydrogel-controlled release of BMAL1-enriched exosomes accelerates diabetic wound healing by promoting angiogenesis and regulating macrophage polarization through activation of the MAPK signaling pathway

Diabetic wounds are characterized by delayed healing due to impaired angiogenesis and dysregulated immune-inflammatory responses. In this study, we developed a novel therapeutic approach utilizing a 4ArmPEG-NHS/4ArmPEG-NH 2 cross-linked hydrogel for the sustained delivery of exosomes (Exos) derived from genetically engineered human adipose-derived mesenchymal stem cells (ADSCs) overexpressing Brain and Muscle Arnt-Like 1 (BMAL1). The efficacy of this system was evaluated both in vitro and in vivo. The hydrogel exhibited excellent injectability, tissue adhesiveness, and rapid hemostatic properties, enabling prolonged release of BMAL1-overexpressing exosomes (BMAL1-Exos). In a diabetic mouse wound model, treatment with BMAL1-Exos@Gel significantly accelerated wound closure, enhanced re-epithelialization, increased microvascular density (as indicated by CD31 and α-SMA immunostaining), improved blood perfusion, and promoted deposition of type I and type III collagen. Mechanistic investigations demonstrated that BMAL1-Exos effectively ameliorated high glucose-induced dysfunction in HUVECs, NIH/3T3 fibroblasts, and HaCaT keratinocytes, while promoting macrophage polarization toward the pro-reparative M2 phenotype and stimulating angiogenesis. These effects were mediated through modulation of the MAPK signaling pathway—specifically activation of JNK, ERK, and p38 kinases—along with upregulation of VEGFb and downregulation of TNF-α. In summary, our research findings indicate that hydrogel loaded with BMAL1-Exos hold promise as a diabetic wound repair strategy by enhancing angiogenesis, modulating immune responses, and remodeling the extracellular matrix. [Graphical abstract] F igure 1. Construction Process of BMAL1 Gene-Modified Exosome-Composite Hydrogel (BMAL1-Exos@Gel) and Its Mechanism for Wound Repair (A) After preparing exosomes (Exos) from adipose-derived stem cells (ADSCs), the BMAL1 gene was transfected to achieve high expression (BMAL1), ultimately obtaining BMAL1-Exos. (B) Chemical synthesis of the hydrogel: N-hydroxysuccinimide ester (-NHS) reacts efficiently with amine (-NH 2 ) to form an amide bond, rapidly curing in situ to seal the wound. (C) Therapeutic mechanism of BMAL1-Exos@Gel in a diabetic mouse model: After local injection of BMAL1-Exos@Gel, it downregulates the expression of M1 macrophage marker iNOS and pro-inflammatory factors (TNF-α, IL-6) by modulating the MAPK signaling pathway, while significantly upregulating M2 macrophage marker CD206 and pro-angiogenic factors (VEGFb), as well as vascular maturation markers (α-SMA, CD31), thereby synergistically promoting wound re-epithelialization and angiogenesis. BMAL1: Brain and Muscle Arnt-Like 1; This figure was drawn by Figdraw.