Improvement of a Three-Layered in vitro Skin Model for Topical Application of Irritating Substances

Abstract

In the field of skin tissue engineering, the development of physiologically relevant in vitro skin models comprising all skin layers, namely epidermis, dermis, and subcutis, is a great challenge. Increasing regulatory requirements and the ban on animal experiments for substance testing demand the development of reliable and in vivo-like test systems, which enable high-throughput screening of substances. However, the reproducibility and applicability of in vitro testing has so far been insufficient due to fibroblastmediated contraction. To overcome this pitfall, an advanced 3-layered skin model was developed. While the epidermis of standard skin models showed an 80% contraction, the initial epidermal area of our advanced skin models was maintained. The improved barrier function of the advanced models was quantified by an indirect barrier function test and a permeability assay. Histochemical and immunofluorescence staining of the advanced model showed well-defined epidermal layers, a dermal part with distributed human dermal fibroblasts and a subcutis with round-shaped adipocytes. The successful response of these advanced 3-layered models for skin irritation testing demonstrated the suitability as an in vitro model for these clinical tests: only the advanced model classified irritative and non-irritative substances correctly. These results indicate that the advanced set up of the 3-layered in vitro skin model maintains skin barrier function and therefore makes them more suitable for irritation testing.

A suspended layer additive manufacturing approach to the bioprinting of tri-layered skin equivalents

“… 24 as well as skin-on-a-chip technologies to screen native functions: permeability and cell response to actives. 25,26 Various models have been proposed ranging upward from mono-cell cultures, with three-layered organoids providing the most reliable platforms for studying tissue function and response. 27,28 However, achieving physiologically relevant thicknesses is often problematic, particularly in self-assembled models, where cells are seeded in an overlapping manner.…”

Implementing organ-on-chip in a next-generation risk assessment of chemicals: a review

“…Additional to metabolism studies, HSE are also used for skin permeation studies of topically applied substances due to the increased barrier function compared to RHE (Alberti et al 2017 ; Sriram et al 2018 ; Schimek et al 2018 ). The barrier properties of HSE models can be further improved by adding a hypodermis (subcutis) to advance the barrier function, as demonstrated by Schmidt et al ( 2020 ). This thicker three-layered skin model reduced the permeation, exhibited suitability as an in vitro test system for irritating substances.…”

“…This thicker three-layered skin model reduced the permeation, exhibited suitability as an in vitro test system for irritating substances. Moreover, the model was proposed to exploit dermal deposition as a possible new endpoint for chemicals in the lipid-rich hypodermis as there is a fundamental lack of studies for investigating the impact and effect on the pharmacokinetics (Turner and Balu-Iyer 2018 ; Schmidt et al 2020 ). To study sun-associated adverse effects or vitiligo pathogenesis, HSE models are complemented with melanocytes (to treat pigmentation).…”

Recent Advances in Regenerative Tissue Fabrication: Tools, Materials, and Microenvironment in Hierarchical Aspects

“…[ 21 ] Tightly connected to the epidermal basement membrane, the dermis consists of papillary and reticular regions that comprise an interconnected network of collagen and elastin fibers produced by fibroblast cells. [ 22 ] Deep in the dermis, the hypodermis, as the deepest layer of skin, connects the skin to the bones and the muscles’ underlying fascia. [ 23 ] Mimicking the hierarchical and unique structure of an organ is essential if an artificial organ with functions and characteristics similar to those of a real cell is to be constructed while considering the biological tissue requirements and the technical strategies ( Figure 1 ).…”