Abstract
Background:
Diabetic wounds (DW) represent one of the most severe complications of diabetes, leading to significant morbidity and mortality. Conventional treatment strategies often fail to address the complex, multifactorial nature of diabetic wound healing, highlighting the need for an integrated therapeutic approach.

Purpose:
This study aimed to develop and evaluate a novel therapeutic system comprising human beta defensin-2 (HBD-2) loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles incorporated into a collagen–chitosan (COL–CS) composite scaffold for enhanced diabetic wound healing.

Materials and Methods:
Biodegradable, crosslinked COL–CS composite scaffolds impregnated with HBD-2-loaded PLGA nanoparticles were formulated and characterized for structural and functional properties. Parameters such as matrix degradation, porosity, and drug release profile were evaluated. In vitro studies assessed biocompatibility, cell migration, and angiogenic potential. Antimicrobial activity was tested against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. In vivo wound healing efficacy was evaluated and compared with control and non-treated groups. Biochemical markers, including MMP-9, TNF-α, MPO, NAG, nitric oxide (NO), and IL-10, were quantified using ELISA.

Results:
The developed crosslinked scaffold demonstrated reduced matrix degradation, optimal porosity, and sustained drug release compared to non-crosslinked scaffolds. In vitro findings confirmed excellent biocompatibility, enhanced cell migration, and increased angiogenesis. The scaffold exhibited significant antimicrobial activity against tested pathogens. In vivo results showed accelerated wound healing in the HBD-2 COL–CS treated group compared to control groups. ELISA analysis revealed a significant reduction in pro-inflammatory markers (MMP-9, TNF-α, MPO, NAG, and NO) along with an increase in anti-inflammatory IL-10 levels.

Conclusions:
The HBD-2-loaded PLGA nanoparticle impregnated COL–CS scaffold significantly enhanced diabetic wound healing through synergistic effects, including antimicrobial action, anti-inflammatory response, angiogenesis, and tissue regeneration. This multifunctional scaffold presents a promising therapeutic strategy for effective management of diabetic wounds.