Human epidermal keratinocytes and human dermal fibroblasts interactions seeded on gelatin hydrogel for future application in skin in vitro 3-dimensional model

Abstract

Introduction: Plenty of biomaterials have been studied for their application in skin tissue engineering. Currently, gelatin-hydrogel is used to support three-dimensional (3D) skin in vitro models. However, mimicking the human body conditions and properties remains a challenge and gelatin-hydrogels have low mechanical properties and undergo rapid degradation rendering them not suitable for 3D in vitro cell culture. Nevertheless, changing the concentration of hydrogels could overcome this issue. Thus, we aim to investigate the potential of gelatin hydrogel with different concentrations crosslinked with genipin to promote human epidermal keratinocytes and human dermal fibroblasts culture to develop a 3D-in vitro skin model replacing animal models.Methods: Briefly, the composite gelatin hydrogels were fabricated using different concentrations as follows 3%, 5%, 8%, and 10% crosslinked with 0.1% genipin or non-crosslinked. Both physical and chemical properties were evaluated.Results and discussion: The crosslinked scaffolds showed better properties, including porosity and hydrophilicity, and genipin was found to enhance the physical properties. Furthermore, no alteration was prominent in both formulations of CL_GEL 5% and CL_GEL8% after genipin modification. The biocompatibility assays showed that all groups promoted cell attachment, cell viability, and cell migration except for the CL_GEL10% group. The CL_GEL5% and CL_GEL8% groups were selected to develop a bi-layer 3D-in vitro skin model. The immunohistochemistry (IHC) and hematoxylin and eosin staining (H&E) were performed on day 7, 14, and 21 to evaluate the reepithelization of the skin constructs. However, despite satisfactory biocompatibility properties, neither of the selected formulations, CL_GEL 5% and CL_GEL 8%, proved adequate for creating a bi-layer 3D in-vitro skin model. While this study provides valuable insights into the potential of gelatin hydrogels, further research is needed to address the challenges associated with their use in developing 3D skin models for testing and biomedical applications.

Contour Analysis of Three-Dimensional Peri-Implant Mucosal Model as an Endpoint Analysis of Photofunctionalization Effects on Implant Abutment Materials

“…On this account, the ascertainment of the specific type of cells that remain attached to various abutment surfaces after conducting the pull test would be the focus to further explore the effect of photofunctionalization on abutment surfaces, and the nature of the soft tissue attachment should be conducted. To accomplish this, the method involves conducting double immunocytochemistry staining for both human gingival fibroblast and oral epithelium in one specimen; for example, vimentin or α-SMA can be used to stain human gingival fibroblasts, and pancytokeratin or cytokeratin 14 for epithelial cells and gene markers [ 52 , 53 ]. In the same manner, double immunohistochemical staining can also be performed on the histological ground section of the soft tissue–implant interface in addition to the hematoxylin and eosin stain.…”

The determination of fibroblast and keratinocyte death types after their transplantation into γ-irradiated porous scaffold in vitro

“…In a series of studies, it has been demonstrated that culturing keratinocytes under unfavorable conditions without the support of fibroblasts or mesenchymal cells leads to the complete death of keratinocytes through apoptosis after 14 days of cultivation (Tanaka et al, 2022, Xu et al, 2023. However, when keratinocytes are cultured on a collagen gel with fibroblasts, they do not undergo apoptosis and actively proliferate (Tahri et al, 2023;Zhang et al, 2023). This result can be explained by the fact that fibroblasts synthesize keratinocyte growth factor (KGF), epidermal growth factor (EGF), colony-stimulating growth factor/interleukin (IL6), fibroblast growth factor (FGF10), which stimulate keratinocytes to synthesize interleukin IL1, which stimulates fibroblasts to the synthesis of KGF (Yen et al, 2014;Bártolo et al, 2022).…”

“…#2800005) and cultured for 168 hours (37.0 ± 0.1 °C, 5.0 ± 0.1% CO 2 , 99.5 ± 0.1% RH, mixing 5 revolutions/hour) in the CERO BioLevitator bioreactor (Omni Life Science GmbH & Co). The conditions for the production of media, immobilization and cultivation of fibroblasts and keratinocytes were in accordance with the current practice of dynamic cultivation of this type of cells (Hofmann et al, 2023;Tahri et al, 2023).…”