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 fibroblast-mediated 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.
Highlights
The assessment of skin toxicity is an essential part of the analyzing the overall effect of chemicals and pharmaceutical products
To understand what prevents dermal-mediated contraction of the epidermal layer of skin models, the new construction approach was compared to skin models cultured with a more common method (Figure 1)
Isolated adipocytes and fibroblasts were encapsulated into a collagen hydrogel and seeded into a silicone tube attached beneath the cell culture insert
Summary
The assessment of skin toxicity is an essential part of the analyzing the overall effect of chemicals and pharmaceutical products. As an ethical alternative for skin toxicity testing, and due to the limited transferability of results from animal assays to human reactions, in vitro-generated organ-like reconstructed human skin models have been developed using tissue engineering techniques. Global legislation has been committed to the development of alternative test methods, in accordance to the 3Rs (reduction, refinement, and replacement of animal experimentation) established by Russell et al (1959). Several skin models are currently commercially available from various producers and academic institutes. While, such models are used for skin corrosion, skin irritation, skin barrier formation, and skin absorption assays, amongst others, the only approved in vitro tests use epidermal models (OECD, 2019a,b). The absence of dermal and subcutaneous layers limits the application of the
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