In Vitro Cytocompatibility Assessment of Novel 3D Chitin/Glucan- and Cellulose-Based Decellularized Scaffolds for Skin Tissue Engineering

Abstract

Background: Naturally derived sustainable biomaterials with high flexibility, mechanical properties, biocompatibility, and the ability to manipulate surface chemistry, providing a natural cellular environment, can be used for tissue engineering applications. However, only a few researchers have demonstrated the exploitation of natural architectures for constructing three-dimensional scaffolds. The chemical decellularization technique for fabricating natural scaffolds and their cytocompatibility assessment for tissue engineering applications need to be thoroughly explored and evaluated. Methods: Decellularization of natural scaffolds has been performed via a chemical method using anionic detergent sodium dodecyl sulfate (SDS) which was used for the in vitro culturing of murine embryonic NIH/3T3 fibroblasts. Techniques such as field-emission scanning electron microscopy (FE-SEM), compressive testing and swelling ratio, and biodegradation were performed to characterize the properties of fabricated decellularized natural scaffolds. Nucleic acid quantification, DAPI, and H&E staining were performed to confirm the removal of nuclear components. In vitro cytocompatibility and live/dead staining assays were performed to evaluate cultured fibroblasts’ metabolic activity and qualitative visualization. Results: 3D chitin/glucan- and cellulose-based scaffolds from edible mushroom (stem) (DMS) and unripe jujube fruit tissue (DUJF) were fabricated using the chemical decellularization technique. FE-SEM shows anisotropic microchannels of highly microporous structures for DMS and isotropic and uniformly arranged microporous structures with shallow cell cavities for DUJF. Both scaffolds exhibited good mechanical properties for skin tissue engineering and DUJF showed a higher compressive strength (200 kPa) than DMS (88.3 kPa). It was shown that the DUJF scaffold had a greater swelling capacity than the DMS scaffold under physiological conditions. At 28 days of incubation, DUJF and DMS displayed approximately 14.97 and 15.06% biodegradation, respectively. In addition, DUJF had greater compressive strength than DMS. Compared to DMS scaffolds, which had a compressive stress of 0.088 MPa at a 74.2% strain, the DUJF scaffolds had a greater compressive strength of 0.203 MPa at a 73.6% strain. The removal of nuclear DNA in the decellularized scaffolds was confirmed via nucleic acid quantification, DAPI, and H&E staining. Furthermore, both of these scaffolds showed good adherence, proliferation, and migration of fibroblasts. DMS showed better biocompatibility and high viability of cells than DUJF. Conclusions: This sustainable scaffold fabrication strategy is an alternative to conventional synthetic approaches for the in vitro 3D culture of mammalian cells for various tissue engineering and cultured meat applications.