112 A new 3D skin model colonized with bacteria for a better understanding of Atopic Dermatitis

Abstract

Atopic dermatitis (AD) is a multifactorial disease associated with epidermal barrier disruption and inflammatory type 2 immune reaction. A microbial dysbiosis characterized by a dominance of Staphylococcus aureus (S. aureus) is also well documented. In earlier studies, in vitro models reproducing some features of AD have been developed by challenging epidermis with either interleukin (IL) cocktails or S. aureus extracts, or by silencing the expression of pivotal epidermal barrier genes. But none of them reproduced the whole pathophysiological AD features. In order to develop a model exposed to both inflammatory cytokines and S. aureus, reconstructed human epidermis (RHE) were treated with a cocktail of IL-4, IL-13, IL-31 and TNF-a from day 5 to day 11, and then exposed to an AD clinical isolate of S. aureus for 24h. Keratinocyte differentiation marker expression, lamellar body trafficking and filaggrin processing were analyzed by immunohistology, transmission electron microscopy and western blotting. IL-8 was measured by immunoassay (AlphaLISA) and S. aureus adherence and growth were evaluated. Treated RHE exhibited alterations observed in AD, including spongiosis, dysregulated expression of differentiation markers (filaggrin, loricrin, claudin-1, corneodesmosin), caspase 14 decrease and strong release of IL-8. We evidenced an abnormal lamellar body distribution by electron microscopy imaging. Moreover, we showed that S. aureus adhered and grew better in the presence of the inflammatory cocktail. Our S. aureus- colonized inflammatory AD skin model recapitulates the impairments of skin barrier observed in AD, at the ultrastructural, microbial and immune levels; it is therefore suitable for mimicking a mild to moderate AD skin profile in order to select active compounds.