Ultrastructural features in organotypic cultures of normal human skin in an in vitro microenvironment mimicking atopic dermatitis

Atopic dermatitis (AD) is a common inflammatory skin disease charactided by chronic, systemic inflammation, early age of onset, persistent itch, marked redness, cracking, and dryness of the skin. Skin infections occur frequently in AD, contribute to the disordered immune activity and are probably related to disruption of skin barrier by inhibiting lipids, tight junctions, and antimicrobial peptide formation. Among the several cytokines involved in the AD pathogenesis interleukin (IL) -4, IL- 1 3 and IL22 play a key role. The present study is focused on the early effects of pro-inflammatory cytokines on ultrastructural changes using transmission electron microscopy (TEM) in a well standardized 3D model of normal human skin organotypic culture. Skin explants obtained from plastic surgery of healthy 20-40 yearold women (n = 5) after informed consent were cultured ovemight in Dulbecco's modified Eagle's medium and treated with 50 nglml IL-4, 50 nglml IL-13 and 100 nglml IL-22 alone or in combination (TRIS). Samples were then harvested 24 and 48 hours after the cytokine incubation. In all samples exposed to cytokines, desmosomes appeared well preserved, comparable to controls. TEM analysis revealed that, starting from24 hours of culture, the exposure toIL-4, but not to IL13, caused a marked enlargement of intercellular spaces and chromatin condensation. In TRIS treated samples, these alterations were less evident, however, we have observed condensation of cytoskeletal filaments. Altogether, this present study strongly suggest that this experimental approach useful for studying the early, direct, and specific effects of pro-inflammatory AD cytokines and may be useful for assessing new biological drugs directed against a specific cytokine.