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Atopic dermatitis (AD) is a chronic cutaneous inflammation driven by T helper 2 cells (Th2) and characterized by a persistent itch. This response relies on Thymic Stromal Lymphopoietin (TSLP) production by keratinocytes and T cell orientation by TSLP-stimulated skin dendritic cells (DCs). Here, we propose new strategies to model AD pathogenesis in human systems. We validated vitamin D3 as an initiation stimulus for human TSLP-dependent Th2 (IL-13+) responses in healthy skin explants. In skin DCs and monocyte-derived DCs (MonoDCs), we showed that vitamin D3 provokes marked phenotypic alterations, characterized by CD14 expression. These cells specifically lead to TSLP-independent Th2 (IL-4+) orientation. In parallel, we have constructed a 3D skin model harboring DCs, sensory neurons and keratinocytes, in which we will evaluate whether vitamin D3 and other stimuli may induce Th2 responses, and how this might be regulated by neurons. This approach will help the rational design of AD-specific therapies.; La dermatite atopique (DA) est une maladie cutanée inflammatoire chronique, caractérisée par un infiltrat de lymphocytes T helper 2 (Th2) et une sensation de prurit. Cette réponse est sous influence du Thymic Stromal Lymphopoietin (TSLP), produit par les kératinocytes, qui affecte la capacité des cellules dendritiques (DCs) de la peau à orienter des lymphocytes T. Nous proposons ici de nouvelles stratégies pour modéliser la DA in vitro chez l’homme. Nous avons validé la vitamine D3 pour initier des réponses Th2 TSLP-dépendantes (IL-13+) sur des explants de peau saine. Dans les DCs de peau ou dérivées de monocytes (MonoDCs), nous avons démontré que la vitamine D3 provoque de profonds changements phénotypiques, caractérisés par l’expression du marqueur CD14. Ces cellules conduisent spécifiquement à une orientation Th2 (IL-4+) TSLP-indépendante. En parallèle, nous avons constitué un modèle 3D de peau contenant des DCs, des neurones et des kératinocytes, dans lequel nous évaluerons si la vitamine D3 et d’autres agents peuvent induire une réponse Th2, et si celle-ci peut être influencée par les neurones. Cette étude aidera à la conception de thérapies spécifiques de la DA.

In the mouse, cutaneous application of vitamin D3 results in a Th2-driven inflammation resembling atopic dermatitis. This response relies on Thymic Stromal Lymphopoietin (TSLP) production by keratinocytes and T cell orientation by TSLP-stimulated skin dendritic cells (DCs). Nevertheless, similar effects of vitamin D3 have not been confirmed thus far by in vitro models involving human skin. We have studied in human skin explants whether vitamin D3 analogs allow emigrating skin DCs to promote human Th2 responses. In line with a previous report, topical application of calcipotriol cream or calcitriol addition in the culture medium increased the rate of CD14+ DCs recovered after 3 days of culture. Using immunofluorescence microscopy of cultured skin and monocyte-derived dendritic cells (MonoDCs) stimulated with calcitriol, we showed that CD14 upregulation is a direct consequence of vitamin D3 exposure rather than an increased migration of CD14+ dermal DCs.Skin-emigrating DCs were collected and cocultured with nau00efve CD4+ T cells from a different donor. This resulted in a clear orientation towards Th2 differentiation, for which CD14+ DCs appeared sufficient. Finally, we found that both analogs of vitamin D3 induced TSLP production by human skin. However, poor correlation between TSLP amounts and the Th2-driving capacity of skin DCs suggested a TSLP-independent effect. Of note, vitamin D3-stimulated MonoDCs also yielded strong Th2 differentiation in the absence of TSLP.Therefore, our findings reveal possible TSLP-independent effects of vitamin D3 on T cell responses, which are currently under investigation. Altogether, we confirmed vitamin D3 as a potent Th2 inducer in a human skin system, paving the way towards accurate modeling of the initiation steps of atopic dermatitis.