P17 - Scar-Reducing Ionically Conductive Chitosan Bandage: Combining Ionic and Electrical Stimulation for Optimal Wound Healing

Saturday, October 11, 2025

10:00 AM - 11:00 AM PDT

Location: Exhibit Hall F, G & H

Presenting Author(s)

LV

Laxmi Chinmaya Vobbineni

Undergraduate Student
University of Connecticut, United States

Co-Author(s)

Sama Abdulmalik, PhD

Assistant Professor
Department of Growth and Development, College of Dentistry, Omaha, NE, USA, United States

Primary Investigator(s)

SK

Sangamesh G. Kumbar, PhD

Professor
Department of Growth and Development, College of Dentistry, Omaha, NE, USA, United States

Introduction: : Chronic and complex skin wounds present a major clinical challenge, often healing poorly and resulting in excessive scarring, reduced mobility, and impaired tissue function. Current treatments, including dressings, pressure therapies, and growth factors, are limited by high costs, invasive procedures, and inconsistent outcomes. There is a critical need for therapies that not only accelerate healing but also reduce fibrosis and restore skin function.

Electrical stimulation (ES) has emerged as a promising strategy by enhancing fibroblast migration, angiogenesis, and extracellular matrix (ECM) remodeling. Separately, potassium channel blockers like 4-aminopyridine (4AP) support tissue regeneration by modulating ion gradients and cellular signaling. While each therapy shows promise, their combined application may offer greater benefits than either alone. Delivering these stimuli in a sustained and controlled manner remains a significant barrier to translation.

To address this, we developed an ionically conductive (IC) chitosan-based polymer platform designed for dual delivery of 4AP and ES. This study investigates the synergistic effects of combined ES and 4AP treatment on human dermal fibroblasts (hDFBs) in vitro using this IC platform. We hypothesize that ES softens fibrotic ECM, improving drug penetration, while 4AP enhances mitochondrial activity and gene expression critical to healing. Together, this dual approach may promote collagen organization, angiogenesis, and regenerative repair.

This integrated approach aims to overcome current limitations in wound care by combining chemical and electrical cues to accelerate healing and minimize scarring. These findings will lay the groundwork for future studies on chronic wounds and clinical translation.

Materials and

Methods: : To fabricate the ionically conductive scaffold, a 3% (w/v) chitosan solution in 2% glacial acetic acid was mixed with 5% (w/w) halloysite nanotubes (HNTs) preloaded with 4AP. HNTs were loaded by dispersing in saturated 4AP solution (50 mg/mL), followed by ultrasonication and cyclic vacuum to enhance encapsulation. The HNT/4AP powder was washed, centrifuged, and freeze-dried. The composite was cast into molds, frozen in liquid nitrogen, and lyophilized. Scaffolds were crosslinked in 1 N NaOH and incubated in 2% phytic acid to introduce phytate ions. Conductivity was measured in PBS using a bipotentiostat. Drug release was quantified by UV-Vis spectroscopy at 262 nm, and mechanical properties were assessed by uniaxial tensile testing.

For in vitro studies, human dermal fibroblasts (hDFBs) were cultured on tissue culture polystyrene plates. Dose-response assays were conducted using logarithmic concentrations of 4AP (0–1000 ng/mL) with or without ES (1 V, 20 Hz, 20 min every other day) to determine optimal 4AP concentration. Scratch assays assessed cell migration, and inflammation was induced with LPS (1 µg/mL) to evaluate cytokine expression. Mitochondrial protection was assessed under oxidative stress using H₂O₂ (0.3 mM) and MitoTracker staining. Gene expression of healing markers was analyzed by qPCR at days 7 and 14.

In vivo, full-thickness dorsal wounds (1 × 2 cm) were created in Wistar rats (n = 6/group). Groups included no scaffold, scaffold, scaffold+4AP, scaffold+ES, and scaffold+4AP+ES. Electrical stimulation was applied via TENS unit (2 mA, 20 Hz, 20 min every other day). Healing was assessed through imaging, histology, and immunohistochemistry.

Results, Conclusions, and Discussions:: The ionically conductive scaffold exhibited sustained release of 4AP over 21 days, with only 6 to 8 percent of the drug released from the crosslinked matrix. SEM imaging confirmed a porous microarchitecture, and both conductivity and mechanical properties remained stable throughout the time period.

In vitro, 100 ng/mL of 4AP significantly enhanced hDFB viability under low-serum conditions and was identified as the optimal dose. Scratch assays showed that the combination of 4AP and ES promoted the fastest wound closure, achieving approximately 99 percent closure by day 3 compared to 78 percent in untreated controls (Figure 1). Under oxidative stress, this dual treatment preserved mitochondrial activity and reduced expression of IL-1β, IL-6, and MMP-2, indicating anti-inflammatory and cytoprotective effects.

Gene expression analysis showed upregulation of key wound healing genes. Cells treated with 4AP+ES displayed higher mRNA levels of extracellular matrix proteins (collagen type I and III), remodeling enzymes (matrix metalloproteinase-2), contractile markers (alpha-smooth muscle actin), and pro-regenerative growth factors (fibroblast growth factor-2 and transforming growth factor-beta) (Figure 2). This indicates a coordinated transition into the proliferative and early remodeling phases of wound repair, supporting matrix deposition, angiogenesis, and contraction.

In vivo, a full-thickness dorsal wound model in Wistar rats confirmed the benefits of this dual treatment. By day 14, wounds treated with 4AP+ES scaffolds showed the greatest closure (93.47 ± 4.48%), outperforming scaffold-only (71.64 ± 15.20%), scaffold+4AP (78.60 ± 8.64%), and scaffold+ES (85.95 ± 14.84%) groups (Figure 3). Histological analysis showed reduced scar elevation, improved re-epithelialization, and increased hair follicle regeneration. Masson’s trichrome staining confirmed dense, aligned collagen deposition, and immunostaining revealed increased Col-I and reduced Col-III, consistent with favorable remodeling. CD31 staining also showed enhanced angiogenesis. Keratin markers CK-10, CK-14, and CK-15 indicated epidermal recovery, and elevated Ki-67 marked increased cell proliferation. Neurotrophic markers NGF, SOX10, and NF-H were also elevated, indicating improved nerve regeneration.

The combined use of 4AP and ES promotes cellular energy maintenance, reduces inflammation, supports extracellular matrix remodeling, and enhances vascular and neural integration. These findings support a promising therapeutic approach for improving chronic wound healing through simultaneous biochemical and bioelectrical stimulation.

Acknowledgements and/or References (Optional): :