Wound Tissue Regeneration by Microfluidic Generated Fibroblast Cell/CuO Nanosheet-Loaded Alginate Hydrogel on an Excisional Full-Thickness Rat Model

Chronic ulcers present numerous challenges in treatment such as prolonged inflammation, infections resistant to drugs, and the formation of scars. In this research, we developed a calcium ion (Ca²⁺) cross-linked alginate (Alg) hydrogel loaded with CuO nanosheet/fibroblast cells via a microfluidic system with substantial efficiency in accelerating healing and preventing infection. Initially, the soft lithography method was utilized to fabricate the microfluidic system, which was employed to produce alginate hydrogel incorporating nanosheets of copper oxide (CuO) and MEF cells. The properties of hydrogel and copper oxide nanosheets were analyzed by using FE-SEM, EDS/EDX, and elemental mapping to determine their physicochemical characteristics. The viability of mouse embryonic fibroblast cells (MEF) in alginate-CuO hydrogel was explored through cell viability assay, and the antibacterial properties were also studied using colony-forming assay. The healing abilities of the hydrogel were investigated using an excisional, full-thickness wound rat model. Our results revealed proper antimicrobial and angiogenic properties with slight cytotoxicity for CuO nanosheets at a concentration of 25 μg/mL. The alginate-CuO-cell-treated group exhibited a faster wound contraction and healing among all treatments. The results of the in vivo assay along with histology and gene expression indicate a synergistic cooperation between MEF and CuO, leading to enhanced re-epithelialization, angiogenesis, and matrix remodeling. In this research, a therapeutic hydrogel with qualities like microbicidal, angiogenic, immune system modulation, and promotion of ECM and epithelium regeneration, resulting in faster healing, was developed. Moreover, there was a synergic impact noticed between CuO nanosheets and MEF cells as well as improved formation of blood vessels and collagen accumulation. In conclusion, this biocompatible hydrogel offers a promising strategy for effective wound healing without the need for invasive procedures.