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.