Design of Hyaluronic Acid-Based Glutathione-Gelatin-Collagen-Laminin Loaded 3D Bioactive Transdermal Tissue Scaffold: Determination of Biocompatibility, Antimicrobial and ROS

Abstract

This study reports the development of a 3D-bioprinted hyaluronic acid (HyA)–based transdermal scaffold incorporating glutathione (GSH), gelatin, collagen (COL), and laminin to enhance wound healing through combined antioxidant, antibacterial, and regenerative functionalities. The scaffold was fabricated using a multi-layer bioprinting strategy, yielding a structurally stable matrix with tunable mechanical performance across Control, TDM_1, TDM_2, and TDM_3 formulations. Mechanical characterization revealed a progressive increase in stiffness with higher matrix density, with elastic modulus values ranging from 9.8 MPa (Control) to 14.3 MPa (TDM_3), and corresponding maximum stress values increasing from 1.32 to 2.19 MPa. Texture Profile Analysis (TPA) further demonstrated formulation-dependent improvements in functional mechanical behavior: hardness increased from 0.42 to 0.79 N, compressibility from 0.35 to 0.70 N, and adhesiveness from −0.18 to −0.37 N s across the scaffold series. These results indicate that optimized structures possess superior deformation resistance, energy absorption capacity, and tissue-adhesive characteristics required for transdermal stability. The GSH encapsulation efficiency reached 81.71%, with a loading capacity of 13.62%, and Franz diffusion studies revealed a sustained drug release of 63.17% over 24 h. Dynamic vapor sorption (DVS) showed <5% moisture uptake, confirming hydrolytic stability, while thermogravimetric analysis (TGA) demonstrated minimal degradation below 200 °C. FTIR spectroscopy verified successful integration of HyA and COL through characteristic amide and carboxyl peaks. NIH-3T3 fibroblast assays indicated >90% viability, and reactive oxygen species (ROS) levels were reduced by up to 45%, confirming biocompatibility and antioxidant efficacy. Antibacterial tests showed activity against E. coli and S. aureus at concentrations >200 μg/mL. SEM imaging revealed a porous microarchitecture supportive of fibroblast attachment and proliferation. Collectively, the updated mechanical and TPA findings confirm that the optimized bioprinted scaffold, particularly TDM_3, offers robust structural integrity, strong bioadhesive performance, sustained drug delivery, and enhanced bioactivity, positioning it as a promising multifunctional platform for chronic wound management.