Increased expression of Filaggrin and Claudin-1 in the ocular surface of patients with atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) is an itchy, chronic and inflammatory skin condition, with dysfunctional immune response and skin barrier defects. Reduction of filaggrin (FLG) and tight junctions (TJ) proteins, such as claudin-1 (CLDN-1), expression in cutaneous epithelial barrier is remarkable in AD pathogenesis. Ocular involvement occurs in approximately 40% of AD patients leading to changes in the structure of the conjunctiva. OBJECTIVES: We aimed to evaluate the expression of FLG and CLDN-1 in the ocular surface of adults with AD, analysing bulbar conjunctival cells collected by a novel non-invasive cellular imprint. METHODS: Bulbar conjunctival epithelial cells were collected by cellular imprint technique, and FLG and CLDN-1 expression were assessed by immunofluorescence (IF) and real-time polymerase chain reaction (RT-PCR). RESULTS: We detected increased expression of FLG and CLDN-1, as well as their transcript levels in AD patients compared with healthy controls (HC). There was a positive correlation between tear film break-up time (TBUT) and FLG expression. Fluorescein staining was inversely associated with FLG expression. CONCLUSIONS: Our results may reflect a reactive response of the ocular surface to AD-related ocular inflammation and associated dry eye disease. Further investigations focusing on the role of FLG and TJ expression in the ocular surface of AD patients may increment the understanding of the pathophysiology of extracutaneous AD and developing future targeted therapies.

Increased expression of in situ IL-31RA and circulating CXCL8 and CCL2 in pemphigus herpetiformis suggests participation of the IL-31 family in the pathogenesis of the disease.

BACKGROUND: Pemphigus herpetiformis (PH) is a rare clinical subtype of pemphigus with the presence of urticarial plaques, severe pruritus, rare acantholysis and eosinophilic spongiosis. OBJECTIVES: The aim of this study was to investigate the influence of IL-31 and pro-inflammatory cytokines/chemokines in the pathogenesis of PH. METHODS: Twenty-five patients with PH and three groups: pemphigus foliaceus (PF = 14), pemphigus vulgaris (PV = 15) and healthy controls (HC = 20) were selected for this study. The groups were analysed by immunohistochemistry utilizing IL-31, IL-31RA, IL-4, IL-17 and TNF-α antibodies. Serum levels of IL-4, IL-13, TNF, CXCL8, CCL5 and CCL2 were evaluated by cytometric bead array. RESULTS: Analysis of IL-31 family of PH patients revealed the following findings: (i) Enhanced in situ expression of IL-31 in PH samples, compared to PF and to PV (epidermis); (ii) Cutaneous IL-31RA expression in PH samples was higher than in PF, PV and HC groups (epidermis and dermis); (iii) PF patients that evolved to PH showed significant increased IL-31RA epidermal expression during the PH phase. Profile of pro-inflammatory cytokines (IL-4, IL-17 and TNF-α) in PH patients' skin exhibited: (i) Enhanced IL-4 expression, when compared to patients with PF (epidermis and dermis) and with PV (epidermis); (ii) Augmented IL-17 expression than PF and PV patients (epidermis); (iii) Augmented expression of TNF-α when compared to PF at the epidermal level. Evaluation of circulating cytokines and chemokines showed higher levels of CXCL8 and CCL2 in PH sera compared to HC group. CONCLUSIONS: IL-31 and IL-31RA, cytokines related to pruritus, and pro-inflammatory chemokines (CXCL8 and CCL2) seem to exert a role in the pathogenesis of PH. These findings support future studies to clarify the role of IL-31 pathway as a potential therapeutic target for patients with PH.

Impaired CD23 and CD62L expression and tissue inhibitors of metalloproteinases secretion by eosinophils in adults with atopic dermatitis.

BACKGROUND: Eosinophils are multifunctional, polymorphonuclear leucocytes that secrete proteins within cytoplasmic granules, such as cytokines, chemokines, metalloproteinases (MMPs) and metalloproteinases tissue inhibitors (TIMPs). Although eosinophilia is a hallmark of atopic dermatitis (AD), several functional aspects of eosinophils remain unknown. OBJECTIVE: We aimed to evaluate the phenotype and functional response of eosinophils under staphylococcal enterotoxin B (SEB) and Toll-like receptor (TLR)-2/6 (FSL-1) stimulation in the secretion of CCL5, MMPs and TIMPs in adults with AD. METHODS: Forty-one adult patients with AD and 45 healthy controls enrolled for the study. Phenotype of eosinophils from granulocytes of peripheral blood was analysed by flow cytometry. We performed evaluation of CCL5 (cytometric bead array), MMP and TIMP (ELISA) secretion, in culture supernatants of purified eosinophils stimulated with SEB or TLR2/6 agonist (FSL-1). RESULTS: We found a higher frequency of LIN1- CCR3+ eosinophils, and decreased expression of CD23 and CD62L receptors in eosinophils of AD patients. There was no difference in MMP and TIMP serum levels between the evaluated groups. However, we detected decreased basal levels of TIMP-1, TIMP-2 and CCL5 in culture supernatants from purified, unstimulated eosinophils from AD patients. CONCLUSION: In adults with AD, phenotypical features of eosinophils reveal decreased expression of early activation and L-selectin receptors. Regarding the functional profile of purified eosinophils related to tissue remodelling in atopic dermatitis, innate immune stimulation (TLR2/6 agonist and SEB) did not affect the ratio of MMP/TIMPs secretion in AD. Our findings reinforce the potential breakdown in tissue remodelling process mediated by eosinophils in AD.

Profile of skin barrier proteins (filaggrin, claudins 1 and 4) and Th1/Th2/Th17 cytokines in adults with atopic dermatitis.

BACKGROUND: Atopic dermatitis (AD) in adults and profile of skin barrier proteins and inflammatory cytokines. OBJECTIVE: Evaluation of the expression of skin barrier proteins such as filaggrin, claudins 1 and 4 and of circulating inflammatory cytokines (Th1/Th2/Th17) in adults with AD. METHODS: Thirty-three adult patients with AD diagnosed according to the Hanifin & Rajkacriteria, and 25 healthy controls were enrolled in the study. AD severity was measured by Eczema Area and Severity Index (EASI). Laboratory assays included immunohistochemistry analysis of skin barrier proteins, such as filaggrin, claudins 1 and 4 and interleukin-17 (IL-17) from skin samples and determination of circulating cytokine levels (IL-2, 4, 5, 6, 10, 17A, TNF and IFN-γ) by flow cytometry (Cytometric Bead Array). RESULTS: We observed a reduced expression of filaggrin and claudin 1 in lesional skin of AD patients, when compared to controls. There was an inverse correlation of filaggrin expression and disease severity. In addition, IL-17 expression was enhanced in AD patients. Similarly, higher levels of inflammatory cytokines (IL-2, 5, 6, 10, 17A and IFN-γ) were found in AD patients. CONCLUSION: Our data reinforce the role of an altered skin barrier in the pathogenesis of AD. Our results show not only reduced expression of filaggrin and claudin 1 in lesional atopic skin but also inverse correlation of filaggrin expression and disease severity. Moreover, elevation of in situ IL-17 and of circulating interleukin levels in AD emphasize the systemic, inflammatory profile of this defective skin barrier dermatosis.