An innovative three-dimensional model of normal human skin to study the proinflammatory psoriatic effects of tumor necrosis factor-alpha and interleukin-17

Abstract

Background: Among all cytokines involved in the pathogenesis and in the progression of psoriasis, Tumor Necrosis Factor (TNF)-alpha and interleukin (IL)-17 play a pivotal role.

Objective: The aim of the present study was to mimic a psoriatic microenvironment and to investigate the early effects of TNF-alpha and IL-17 in a three-dimensional model of organotypic normal human skin.

Methods: Human skin explants were obtained from plastic aesthetic surgery of healthy young women 20-40years old (n=7). The study was approved by the Institutional Review Board and written informed consent was obtained from all subjects. Bioptic fragments were cultured at the air-liquid interface overnight in a Transwell system and further divided before adding either 50ng/ml IL-17 or 100ng/ml TNF-alpha or a combination of both cytokines. For each subject, a control sample was cultured without any cytokine. Samples were harvested 24 or 48h after cytokine incubation. At both time points and for all cytokine treatments a bioptic fragment obtained from each patient was processed. Epidermal proliferation, expressions of terminal differentiation (keratin 10, K10, and 14, K14) and of intercellular adhesion (occludin for tight junctions and E-cadherin for adherens junctions) biomarkers were investigated by indirect immunofluorescence.

Results: IL-17 and TNF-alpha induced an early and statistically significant inhibition of keratinocyte proliferation (more than 80% compared with their respective controls). At 24h, the combination of both cytokines did not further reduce cell proliferation. Starting from 24h of incubation, a non-continuous occludin expression in the granular layer was observed after both IL-17 and TNF-alpha exposure. Immunolabelling for E-cadherin in adherens junctions, for K10 in the suprabasal layers, and for K14 in the basal layer was similar in all experimental groups and unaffected after cytokine treatment.

Conclusions: These results suggest that in this experimental model IL-17 and TNF-alpha induced an early alteration of the homeostasis of the inner proliferative layer and of the upper granular layer, as shown by cell proliferation inhibition and occludin expression.

Keywords: Cell junctions; Keratin 10; Keratin 14; Keratinocyte proliferation; Psoriasis.