Development and Characterization of a Hydrogel Incorporating Protamine–Hyaluronic Acid Nanoparticles Co-Loaded with Disulfiram and Coumarin for Diabetic Wound Healing

Background: Diabetes-associated wound healing is frequently impaired due to persistent hyperglycemia, which causes vascular dysfunction and neuropathy, thereby delaying tissue repair. Objectives: In this study, hyaluronic acid and protamine nanoparticles (HA-PRO-NPs) were developed and optimized to encapsulate disulfiram (DSF) and coumarin (COM) for potential application in diabetic wound healing. Methods: The nanoparticles were prepared using a simple ionic interaction method and characterized for their particle size, zeta potential, morphology and encapsulation efficiency. Then, HA-PRO-NPs were incorporated into a hydrogel that contains carboxymethyl cellulose (CMC) and propylene glycol (PG). The hydrogel formulations were evaluated for swelling behavior, spreadability and DSF and COM release kinetics. Results: The prepared formulations were safe against human dermal fibroblasts (HDFs), with cell viability remaining above 70% over the concentration ranges tested. In the scratch assay, HA-PROT-DSF-COM-NPs improved HDF migration, achieving approximately 81% wound closure. An in vivo study was conducted using streptozotocin-induced diabetic mice. Topical application of Gel-HA-PROT-DSF-COM-NPs promoted wound closure. The study monitored wound contraction on Days 0, 3, 7, and 12. By Day 12, topical administration of the dual-cargo system (Gel-HA-PROT-DSF-COM NPs, n = 6) achieved 78.2 ± 4.1% wound closure. This was significantly higher than the single-drug formulation (Gel-HA-PROT-DSF-NPs: 59.5 ± 4.8%) and the untreated Control (Diabetic) group (55.4 ± 5.2%, p < 0.01), while the non-diabetic Control group (non diabetic) reached 97.6 ± 3.5% closure. Semi-quantitative histopathological evaluation showed that the Gel-HA-PROT-DSF-COM NPs achieved a healing score of 4/5, characterized by optimized epidermal regeneration with preserved adnexal structures and moderate dermal remodeling. Conclusions: In conclusion, Gel-HA-PROT-DSF-COM-NPs demonstrated promising potential as a topical formulation for improving wound healing under diabetic conditions.