A Proinflammatory Psoriatic Microenvironment Has Early Effects on Keratinocyte Proliferation/Differentiation and Induces Ferroptosis in HaCaT Cells

<i>Background</i>: The interaction between keratinocytes and proinflammatory cytokines is essential in the development of psoriatic lesions. The synergism among these cytokines and their involvement in ferroptosis are not yet elucidated. This study aimed at evaluating the early impact of a complete proinflammatory microenvironment on keratinocyte differentiation, intercellular adhesion, proliferation, and induction of ferroptosis. <i>Methods</i>: HaCaT cells were differentiated with 1.8 mM CaCl₂ and treated with a cytokine combination (MIX) containing IL-17A, IL-22, IL-23, and TNF-alpha for 24 and 48 h. Claudin 1 (CLDN-1), Zonula Occludens 1 (ZO-1), and keratins (K)10/K14 expression was analyzed by immunofluorescence and immunoblot analysis, paralleled by proliferation and ultrastructural analysis. Ferroptosis was induced with erastin and RSL3 and evaluated by testing glutathione (GSH)/glutathione peroxidase 4 (GPX4) protein expression, GSH levels, cell availability/toxicity, intracellular iron and ATP levels. <i>Results</i>: After MIX incubation at T48, CLDN-1 and ZO-1 immunofluorescences were reduced in HaCaT cells, while K10 and K14 were unaffected. The proliferative activity was reduced. Psoriatic-like MIX triggered the ferroptotic pathway, as shown by the increase in intracellular iron levels as well as by the reduction in GPX4 protein expression, the decrease in GSH levels, cell availability, and ATP levels. <i>Conclusions</i>: This experimental model mimics the early pathogenetic processes underlying psoriatic plaque formation/progression paving the way for new therapeutic strategies.