Langerhans and inflammatory dendritic epidermal cells in atopic dermatitis are tolerized toward TLR2 activation.

BACKGROUND: The skin of atopic dermatitis (AD) patients presents a significant dysbalance of the microbiome with a high colonization by Staphylococcus aureus (S. aureus), which positively correlates with the severity of the disease. OBJECTIVE: Understanding the role of epidermal dendritic cells (DC) as link between the innate and the adaptive immune systems in AD. METHODS: Comparative phenotypic and functional analysis of TLR2 on Langerhans cells (LC) and inflammatory dendritic epidermal cells (IDEC) in organotypic models as well as freshly isolated cells from healthy and AD skin. RESULTS: In situ analysis of freshly isolated LC and IDEC from AD skin revealed decreased TLR2 expression compared to LC from healthy skin. In contrast to IDEC, LC from AD skin failed to display any evidence for in situ activation. Exposure to TLR2 ligand Pam3Cys resulted in maturation and increased migratory activity of LC from normal skin. LC and IDEC from AD were unresponsive to TLR2 ligand in that they failed to mature and displayed a high spontaneous migratory activity. Keratinocytes from both healthy and AD skin expressed similar levels of TLR2. The production of IL-6 and IL-10 was impaired by Pam3Cys in supernatants from AD skin. IL-18 was significantly higher in supernatants from AD skin and not influenced by TLR2 ligation, when compared to healthy skin. CONCLUSION: Our results suggest that TLR2-mediated sensing of S. aureus-derived signals is strongly impaired in LC from AD skin. This phenomenon may partly contribute to the immune deviation in AD and the lack of S. aureus clearance.

AhR mediates an anti-inflammatory feedback mechanism in human Langerhans cells involving FcεRI and IDO.

BACKGROUND: Aryl hydrocarbon receptor (AhR), an important regulator of immune responses, is activated by UVB irradiation in the skin. Langerhans cells (LC) in the epidermis of patients with atopic dermatitis (AD) carry the high-affinity receptor for IgE, FcεRI, and are crucially involved in the pathogenesis of AD by inducing inflammatory responses and regulating tolerogenic processes. OBJECTIVES: We investigated AhR and AhR repressor (AhRR) expression and functional consequences of AhR activation in human ex vivo skin cells and in in vitro-generated LC. METHODS: Epidermal cells from healthy skin were analyzed for their expression of AhR and AhRR. LC generated from CD34+ hematopoietic stem cells (CD34LC) were treated with the UV photoproduct and AhR ligand 6-formylindolo[3,2-b]carbazole (FICZ). Cell surface receptors, transcription factors, and the tolerogenic tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO) were analyzed using flow cytometry and quantitative PCR. RESULTS: Epidermal LC and CD34LC express AhR and AhRR. AhR was also found in keratinocytes, which lack AhRR. AhR activation of LC by FICZ caused downregulation of FcεRI in CD34LC without affecting their maturation. AhR-mediated regulation of FcεRI did not involve any known transcription factors related to this receptor. Furthermore, we could show upregulation of IDO mediated by AhR engagement. CONCLUSIONS: Our study shows that AhR activation by FICZ reduces FcεRI and upregulates IDO expression in LC. This AhR-mediated anti-inflammatory feedback mechanism may dampen the allergen-induced inflammation in AD.

TLR2 down-regulates FcεRI and its transcription factor PU.1 in human Langerhans cells.

BACKGROUND: Epidermal Langerhans cells (LC) expressing the high-affinity receptor for IgE (FcεRI) play a key role in atopic dermatitis (AD). AD skin is highly colonized with Staphylococcus aureus (S.a.), which are sensed by Toll-like receptor 2 (TLR2). We hypothesized that TLR2 may impact on the expression of FcεRI on LC. OBJECTIVES: To study a putative impact of TLR2 signaling on FcεRI, we analyzed FcεRI and known transcription factors of the receptor after ligand binding to TLR2. METHODS: We generated LC from CD34(+) progenitors in vitro (CD34LC) expressing FcεRI and TLR2 as well as its partners TLR1 and TLR6. The expression of FcεRI and known transcription factors of the receptor was analyzed on the protein and RNA level by flow cytometry, Western blotting, and real-time PCR. RESULTS: For CD34LC from 123 donors, we observed a high heterogeneity in FcεRI surface expression correlating with mRNA level of its α-chain. Stimulation of TLR1/2 or TLR2/6 dramatically down-regulated FcεRI on protein and mRNA level of both α- and γ-chain. Further analysis of putative transcription factors for FCER1A revealed the lack of GATA1 in CD34LC, weak expression of ELF1 and YY1, and high expression of PU.1. While ELF1 and YY1 appeared to be little affected by TLR2 engagement, PU.1 was significantly down-regulated. CONCLUSIONS: Taken together, our findings show that in human, LC ligation of TLR2 by S.a.-derived products down-regulates FcεRI and its transcription factor PU.1, thus suggesting that FcεRI is controlled by PU.1 in these cells.