T cells expressing the cutaneous lymphocyte antigen (CLA) mediate pathogenic inflammation in atopic dermatitis (AD). The molecular alterations contributing to their dysregulation remain unclear. With the aim to elucidate putative altered pathways in AD we profiled DNA methylation levels and miRNA expression in sorted T cell populations (CD4 + , CD4 + CD45RA + naïve, CD4 + CLA + , and CD8 + ) from adult AD patients and healthy controls (HC). Skin homing CD4 + CLA + T cells from AD patients showed significant differences in DNA methylation in 40 genes compared to HC ( p < 0.05). Reduced DNA methylation levels in the upstream region of the interleukin-13 gene ( IL13 ) in CD4 + CLA + T cells from AD patients correlated with increased IL13 mRNA expression in these cells. Sixteen miRNAs showed differential expression in CD4 + CLA + T cells from AD patients targeting genes in 202 biological processes ( p < 0.05). An integrated network analysis of miRNAs and CpG sites identified two communities of strongly interconnected regulatory elements with strong antagonistic behaviours that recapitulated the differences between AD patients and HC. Functional analysis of the genes linked to these communities revealed their association with key cytokine signaling pathways, MAP kinase signaling and protein ubiquitination. Our findings support that epigenetic mechanisms play a role in the pathogenesis of AD by affecting inflammatory signaling molecules in skin homing CD4 + CLA + T cells and uncover putative molecules participating in AD pathways.

Background: IgE-reactivity to antigens from gram-positive and negative bacteria is common in patients suffering from respiratory and skin manifestations of allergy, but the routes and mechanisms of sensitisation are not fully understood. The analysis of the genome, transcriptome and microbiome of house dust mites has shown that Staphylococcus aureus and Escherichia coli species are abundant bacteria within the house dust mites microbiome. Therefore, our aim was to investigate if house dust mites are carriers of bacterial antigens leading to IgE sensitisation in patients suffering from atopic dermatitis. Methods: Plasma samples from atopic dermatitis patients (n = 179) were analysed for IgE-reactivity to a comprehensive panel of micro-arrayed house dust mites allergen molecules and to Staphylococcus aureus and Escherichia coli by IgE immunoblotting. Antibodies specific for Staphylococcus aureus and Escherichia coli antigens were tested for reactivity to nitrocellulose-blotted extract from purified house dust mites bodies and the IgE-reactive antigens were detected by IgE-immunoblot inhibition experiments. IgE antibodies directed to bacterial antigens in house dust mites were quantified by IgE ImmunoCAPTM inhibition experiments. Results: IgE-reactivity to bacterial antigens was significantly more frequent in atopic dermatitis patients sensitised to house dust mites than in atopic dermatitis patients without house dust mites sensitisation. Staphylococcus aureus and Escherichia coli antigens were detected in immune-blotted house dust mites extract and the presence of IgE-reactive antigens in house dust mites was demonstrated by qualitative and quantitative IgE inhibition experiments. Conclusions: House dust mites may serve as carriers of bacteria responsible for the induction of IgE sensitisation to microbial antigens.

Background Atopic dermatitis (AD) is a complex chronic inflammatory disease where allergens can act as specific triggering factors. Aim To characterize the specificities of IgE-reactivity in patients with AD to a broad panel of exogenous allergens including microbial and human antigens. Methodology Adult patients with AD were grouped according to the SCORAD index, into severe (n = 53) and moderate AD (n = 126). As controls 43 patients were included with seborrhoeic eczema and 97 individuals without history of allergy or skin diseases. Specific IgE reactivity was assessed in plasma using Phadiatop®, ImmunoCap™, micro-arrayed allergens, dot-blotted recombinant Malassezia sympodialis allergens, and immune-blotted microbial and human proteins. Results IgE reactivity was detected in 92% of patients with severe and 83% of patients with moderate AD. Sensitization to cat allergens occurred most frequently, followed by sensitization to birch pollen, grass pollen, and to the skin commensal yeast M. sympodialis. Patients with severe AD showed a significantly higher frequency of IgE reactivity to allergens like cat (rFel d 1) and house dust mite (rDer p 4 and 10), to Staphylococcus aureus, M. sympodialis, and to human antigens. In contrast, there were no significant differences in the frequencies of IgE reactivity to the grass pollen allergens rPhl p 1, 2, 5b, and 6 between the two AD groups. Furthermore the IgE reactivity profile of patients with severe AD was more spread towards several different allergen molecules as compared to patients with moderate AD. Conclusion We have revealed a hitherto unknown difference regarding the molecular sensitization profile in patients with severe and moderate AD. Molecular profiling towards allergen components may provide a basis for future investigations aiming to explore the environmental, genetic and epigenetic factors which could be responsible for the different appearance and severity of disease phenotypes in AD.