Evaluation of in vitro coculture of keratinocytes derived from foreskin and adipose-derived mesenchymal stem cells (AMSCs) on a multilayer oxygen-releasing electrospun scaffold based on PU/PCL.Sodium percarbonate (SPC)-gelatine/PU.

Abstract

Despite significant advancements in tissue engineering and regenerative medicine during the last two decades, the fabrication of proper scaffolds with appropriate cells can still be considered a critical achievement in this field. Hypoxia is a major stumbling block to chronic wound healing, which restrains tissue engineering plans because a lack of oxygen may cause cell death. This study evaluated the cocultured human keratinocytes and human adipose-derived mesenchymal stem cells (AMSCs) on a multilayer oxygen-releasing electrospun scaffold based on PU/PCL.Sodium percarbonate (SPC)-gelatin/PU. The scaffold was characterized using Fourier transform infrared (FTIR) and scanning electron microscopy (SEM) methods. Flow cytometry confirmed mesenchymal stem cells, and then the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay and DAPI staining were used to assess the in vitro biocompatibility of the scaffold. The experimental results showed that the multilayer electrospun scaffold containing 2.5% SPC could efficiently produce oxygen. Furthermore, according to cell viability results, this structure makes a suitable substrate for the coculture of keratinocytes and AMSCs. Gene expression analysis of various markers such as Involucrin, Cytokeratin 10, and Cytokeratin 14 after 14 days confirmed that keratinocytes and AMSCs coculture on PU/PCL.SPC-gelatin/PUelectrospun scaffold promotes dermal differentiation and epithelial proliferation compared to keratinocytes single-cell culture. Therefore, our study supports using oxygen-releasing scaffolds as a potential strategy to hasten skin tissue regeneration. Based on the results, this structure is suggested as a promising candidate for cell-based skin tissue engineering. Given that the developed oxygen-generating polymeric electrospun scaffolds could be used as part of a future strategy for skin tissue engineering, the PU/PCL.SPC-gelatin/PUhybrid electrospun multilayer scaffold in combination with keratinocyte/AMSC coculture is proposed as an effective substrate for skin tissue engineering and regenerative medicine platforms.