A bilayer skin-mimicking 3D-printed/electrospun wound dressing incorporating deferoxamine and propolis for angiogenesis-associated wound healing: an in vitro, in ovo, and in vivo evaluation

Abstract

Bilayer wound dressings can integrate an outer protective barrier with a bioactive wound-contact layer, offering functional stratification that more closely reflects selected epidermal and dermal roles than single-layer dressings. In this study, a bilayer wound dressing was fabricated by combining a propolis-coated 3D-printed (3DP) polycaprolactone (PCL) top layer with a deferoxamine-loaded electrospun PCL/polyvinyl alcohol (PVA) base layer (NF.DFO). The ethanolic propolis (Pr) extract used for dip coating was chemically standardized and showed a total phenolic content of 477 +/- 2.53 mg GAE/g extract and a total flavonoid content of 134 +/- 5.43 mg QE/g extract. The 3DP layer exhibited a strut diameter of 445.33 +/- 16.5 microm, a pore size of 737.53 +/- 77.05 microm, and a water contact angle of 98.75 degrees , indicating a hydrophobic barrier surface. The bilayer scaffold showed moderate water uptake, reduced water vapor permeability, a tensile strength of 15.69 +/- 2.27 MPa, and an interlayer peel strength of 19.4 +/- 2.6 mN/mm, supporting its handling stability and interfacial integrity. DFO release reached 84.5 over seven days under in vitro conditions. In vitro, in ovo, and in vivo evaluations showed cytocompatibility, cell adhesion, antibacterial activity associated with propolis, enhanced early angiogenic-marker response, reduced IL-6 and TNF-alpha immunopositive densities, improved collagen organization, and faster macroscopic wound closure in the BL group compared with the other tested groups. Overall, the BL scaffold represents a promising preclinical 3D-printed/electrospun wound-dressing platform containing 1 wt DFO for dry to low-to-moderately exuding wounds; however, further studies are required to confirm vascular maturation, cytokine regulation, and clinical relevance.