Atopic Dermatitis Skin Lesions Research Articles

The Role of OX40-OX40L Axis in the Pathogenesis of Atopic Dermatitis.

OX40 is a co-stimulatory immune checkpoint molecule that promotes the activation and the effector function of T lymphocytes through interaction with its ligand (OX40L) on antigen-presenting cells. OX40-OX40L axis plays a crucial role in Th1 and Th2 cell expansion, particularly during the late phases or long-lasting response. Atopic dermatitis is characterized by an immune dysregulation of Th2 activity and by an overproduction of proinflammatory cytokines such as interleukin (IL)-4 and IL-13. Other molecules involved in its pathogenesis include thymic stromal lymphopoietin, IL-33, and IL-25, which contribute to the promotion of OX40L expression on dendritic cells. Lesional skin in atopic dermatitis exhibits a higher level of OX40L+-presenting cells compared with other dermatologic diseases or normal skin. Recent clinical trials using antagonizing anti-OX40 or anti-OX40L antibodies have shown symptom improvement and cutaneous manifestation alleviation in patients with atopic dermatitis. These findings suggest the relevance of the OX40-OX40L axis in atopic dermatitis pathogenesis.

Inhibition of RNase 7 by RNase inhibitor promotes inflammation and Staphylococcus aureus growth: Implications for atopic dermatitis.

The antimicrobial ribonuclease RNase 7 is abundantly expressed in the epidermis of lesional skin of atopic dermatitis (AD). Host RNase inhibitor (RI) binds to RNase 7 and blocks its ribonuclease activity. This study aimed to evaluate the impact of RNase 7-RI interactions on AD. Cultured human primary keratinocytes, with siRNA-mediated downregulation of RNase 7 and RI, were stimulated with the synthetic RNA polyinosinic-polycytidylic acid (poly I:C). Induction of proinflammatory mediators was analyzed by real-time PCR and ELISA. RI expression in AD non-lesional and lesional skin biopsies and healthy controls was analyzed by real-time PCR and immunostaining. RI protein release invivo on the AD skin surface was determined by western blot. Antimicrobial and ribonuclease assays were used to investigate the functional role of RI. RNase 7 inhibited the RNA-induced expression of proinflammatory mediators in keratinocytes. Accordingly, downregulation of RNase 7 in keratinocytes enhanced RNA-mediated induction of proinflammatory mediators, whereas downregulation of RI had the opposite effect. RI was released by damaged keratinocytes and epidermis. Invivo expression and release of RI on the skin surface were enhanced in lesional AD skin. Rinsing solution from the surface of lesional AD skin blocked the ribonuclease activity of RNase 7. The anti-Staphylococcus aureus activity of RNase 7 was abrogated by RI. Our data suggest a novel role of RI as a trigger factor of inflammation in AD by blocking the ribonuclease and antimicrobial activity of RNase 7, thereby enhancing RNA-mediated inflammation and S. aureus growth.

Interplay of cytokines in the pathophysiology of atopic dermatitis: insights from Murin models and human.

The pathogenesis of atopic dermatitis (AD) is understood to be crucially influenced by three main factors: dysregulation of the immune response, barrier dysfunction, and pruritus. In the lesional skin of AD, various innate immune cells, including Th2 cells, type 2 innate lymphoid cells (ILC2s), and basophils, produce Th2 cytokines [interleukin (IL)-4, IL-5, IL-13, IL-31]. Alarmins such as TSLP, IL-25, and IL-33 are also produced by epidermal keratinocytes, amplifying type 2 inflammation. In the chronic phase, not only Th2 cells but also Th22 and Th17 cells increase in number, leading to suppression of filaggrin expression by IL-4, IL-13, and IL-22, which further deteriorates the epidermal barrier function. Dupilumab, which targets IL-4 and IL-13, has shown efficacy in treating moderate to severe AD. Nemolizumab, targeting IL-31RA, effectively reduces pruritus in AD patients. In addition, clinical trials with fezakinumab, targeting IL-22, have demonstrated promising results, particularly in severe AD cases. Conversely, in murine models of AD, several cytokines, initially regarded as promising therapeutic targets, have not demonstrated sufficient efficacy in clinical trials. IL-33 has been identified as a potent activator of immune cells, exacerbating AD in murine models and correlating with disease severity in human patients. However, treatments targeting IL-33 have not shown sufficient efficacy in clinical trials. Similarly, thymic stromal lymphopoietin (TSLP), integral to type 2 immune responses, induces dermatitis in animal models and is elevated in human AD, yet clinical treatments like tezepelumab exhibit limited efficacy. Therapies targeting IL-1α, IL-5, and IL-17 also failed to achieve sufficient efficacy in clinical trials. It has become clear that for treating AD, IL-4, IL-13, and IL-31 are relevant therapeutic targets during the acute phase, while IL-22 emerges as a target in more severe cases. This delineation underscores the necessity of considering distinct pathophysiological aspects and therapeutic targets in AD between mouse models and humans. Consequently, this review delineates the distinct roles of cytokines in the pathogenesis of AD, juxtaposing their significance in human AD from clinical trials against insights gleaned from AD mouse models. This approach will improve our understanding of interspecies variation and facilitate a deeper insight into the pathogenesis of AD in humans.

Magnolol Reduces Atopic Dermatitis-like Symptoms in BALB/c Mice.

In traditional Korean medicines, Magnolia officinalis is commonly included for the remedy of atopic dermatitis, and magnolol is a major constituent of Magnolia officinalis. Its pharmacological effects include anti-inflammatory, hepatoprotective, and antioxidant effects. Using BALB/c mice repeatedly exposed to 1-chloro-2,4-dinitrobenzene (DNCB), magnolol was evaluated in atopic dermatitis-like lesions. Administration of magnolol (10 mg/kg, intraperitoneal injection) markedly relieved the skin lesion severity including cracking, edema, erythema, and excoriation, and significantly inhibited the increase in IgE levels in the peripheral blood. A DNCB-induced increase in mast cell accumulation in atopic dermatitis skin lesions was reversed by magnolol administration, as well as a rise in expression levels of pro-inflammatory Th2/Th17/Th1 cytokines' (IL-4, IL-13, IL-17A, IFN-γ, IL-12A, TARC, IL-8, and IL-6) mRNAs in the lymph nodes and skin (n = 5 per group). In lymph nodes, magnolol reversed DNCB's increase in CD4+RORγt+ Th17 cell fraction and decrease in CD4+FoxP3+ regulatory T cell fraction. The results also showed that magnolol suppressed T cell differentiation into Th17 and Th2 cells, but not Th1 cells. Magnolol suppresses atopic dermatitis-like responses in the lymph nodes and skin, suggesting that it may be feasible to use it as a treatment for atopic dermatitis through its suppression of Th2/Th17 differentiation.

Changes in the composition of molecular species of covalently bound and free ceramides [EOS], and their correlation with disease severity in atopic dermatitis.

Ceramides are major constituents of stratum corneum (SC) intercellular lipids involved in skin barrier function. The ratio of molecular species of ceramides and their correlation with disease severity was examined in patients with atopic dermatitis (AD). Thirty-eight patients with AD and 32 healthy controls (HCs) were assessed for transepidermal water loss, SC collection and clinical assessment. The ceramide content of different molecular species in the samples was quantified using high-performance liquid chromatography coupled with tandem mass spectrometry. Unsaturated acyl chains of both covalently bound and free ceramides [EOS] were higher in AD lesional skin than those in AD non-lesional or normal HC skin. The proportion of unsaturated acyl chains (C30:1, C32:1 and C34:1) was higher than other ceramide molecular species among covalently bound and free ceramides [EOS] in patients with AD. The proportion of unsaturated acyl chains in covalently bound ceramides was positively correlated with transepidermal water loss (r = 0.600) when considering the total number of non-lesional and lesional skin. Additionally, thymus and activation-regulated chemokine (TARC) showed a positive correlation with unsaturated acyl chains proportion in AD non-lesional (r = 0.676) and lesional (r = 0.503) skin. Our study is the first to show the increase in unsaturated acyl chains of both covalently bound and free ceramides [EOS] in lesional and non-lesional skin in AD for each molecular species. This increase is associated with dryness and impaired barrier function, which correlates with TARC levels, a marker for the degree of type 2 inflammation. We speculate that type 2 inflammation exacerbation leads to abnormal epidermal lipid metabolism in the skin of patients with AD.

Epidermal loss of Bcl6 exacerbates MC903-induced atopic dermatitis-like skin inflammation

Atopic dermatitis (AD) is a chronic inflammatory skin disease where Th2-type immune responses are dominant. In the lesional skin of AD, keratinocytes show differentiation defects and secrete proinflammatory cytokines and chemokines, amplifying Th2-type responses in AD. We previously reported that inducible loss of B-cell lymphoma 6 (Bcl6), a transcription repressor and a master transcriptional regulator of follicular helper T cells and germinal center B cells, in the whole body results in upregulation of Th2-related cytokines in mouse skin. However, the role of Bcl6 in keratinocytes remains to be clarified. Here, we observed that BCL6 positively regulates the expression of keratinocyte differentiation markers and plasma membrane localization of adherence junctional proteins in keratinocyte cell culture. Although keratinocyte-specific loss of Bcl6 alone did not induce AD-like skin inflammation, it aggravates MC903-induced AD-like skin inflammation in mice. In addition, Bcl6 expression is decreased in the epidermis of lesional skin from MC903-induced AD-like skin inflammation in mice. These results strongly suggest that Bcl6 downregulation in keratinocytes contributes to the development and aggravation of AD-like skin inflammation in mice.

Characterization of the Pro-Inflammatory and Pruritogenic Transcriptome in Skin Lesions of the Experimental Canine Atopic Acute IgE-Mediated Late Phase Reactions Model and Correlation to Acute Skin Lesions of Human Atopic Dermatitis.

Intradermal injection of anti-immunoglobulin E (IgE) antibodies in dogs grossly and histologically resemble naturally occurring atopic dermatitis (AD). However, the activated inflammatory and pruritic pathways have not been characterized. The objectives of this study were to characterize the inflammatory transcriptome of experimental acute canine IgE-induced lesions and to determine how these correlate to the transcriptome of naturally occurring human and canine acute atopic dermatitis. Biopsies were collected at 6 and 24 h after intradermal injections of anticanine-IgE antibodies to eight healthy male castrated Beagles; healthy and saline-injected skin served as controls. We extracted total RNA from skin biopsies and analyzed transcriptome using RNA-sequencing. Gene expressions of IgE-induced biopsies were compared to that of controls from the same subject (1.5-fold change, p-adjusted value ≤ 0.05). Acute IgE-mediated lesions had a significant upregulation of pro-inflammatory (e.g., LTB, IL-1B, PTX3, CCL2, IL6, IL8, IL18), T helper-(Th)1/IFNγ signal (e.g., STAT-1, OASL, MX-1, CXCL10, IL-12A) and Th2 (e.g., IL4R, IL5, IL13, IL33 and POSTN) genes, as well as Th2 chemokines (CCL17, CCL24). Pathway analysis revealed strong significant upregulation of JAK-STAT, histamine, IL-4 and IL13 signaling. Spearman correlation coefficient for the shared DEGs between canine anti-canine-IgE and human AD samples revealed a significant moderate positive correlation for anti-canine-IgE 6-h samples (r = 0.53) and 24-h samples (r = 0.47). In conclusion, acute canine IgE-mediated skin lesions exhibit a multipolar immunological axis upregulation (Th1, Th2 and Th17) in healthy dogs, resembling acute spontaneous human AD lesions.

563 - Dupilumab improves skin lipid composition in atopic dermatitis irrespective of patient filaggrin (FLG) mutation status

Abstract Introduction/Background Type 2 inflammatory cytokines interleukin 4 (IL-4) and IL-13 play an important role in skin barrier disruption in atopic dermatitis (AD). Filaggrin and ceramides play a crucial role in skin barrier integrity. Loss of function mutations in the FLG gene are associated with the impaired skin barrier function and more severe AD.1,2 FLG mutations only affect a minority of AD patients; however, increased IL-4 and IL-13 cytokines are a common cause of reduced filaggrin expression in patients with AD, independent of FLG genotype. Objectives To explore whether dupilumab treatment improves skin barrier function in patients with or without FLG mutations. Methods In the BArrier function and LIpidomics STudy in Atopic Dermatitis (BALISTAD; NCT04447417), a 16-week study in patients with AD aged 12 to 65 years, adult patients with AD received dupilumab 300 mg every 2 weeks; adolescent patients with AD received dupilumab 200 mg every 2 weeks if their baseline weight was <60 kg and 300 mg if ≥60 kg. FLG mutations were evaluated in DNA from blood samples of consenting patients with AD and healthy volunteers. Transepidermal water loss (TEWL) was assessed longitudinally after 5 skin tape strippings (STS) from AD lesions (n = 26) and from the skin of healthy participants (n =26) (age: 12 to 63 years) over a 16-week course of dupilumab treatment. Quantitative N(C18)S-ceramide analysis of STS samples collected on Days 1, 15, 29, 57, and 85, and at Week 16 from AD lesions and from the skin of healthy participants was performed using liquid chromatography tandem mass spectrometry. Results At baseline, mean TEWL after 5 STS (TEWL5) was significantly higher in AD lesional skin than healthy skin (p<0.0001). The mean TEWL5 in AD lesions in subjects with FLG mutations (n = 6/19) was significantly higher at baseline than in AD subjects without mutations (P < 0.0001). Dupilumab treatment significantly reduced TEWL5 in AD lesional skin as early as Week 2 with a progressive decrease through Week 16 (P < 0.0001). Reduction in mean TEWL5 was similar from Week 2 to Week 16 in AD patients with and without FLG mutations. At Week 16, TEWL5 was comparable to healthy skin in the lesional skin of AD patients with and without FLG mutations (P > 0.05). AD skin lesions had increased levels of N(C18)S-ceramides at baseline (P < 0.0001); but no differences were noted in subjects with or without FLG mutations (P > 0.05). Dupilumab treatment significantly reduced levels of N(C18)S-ceramides in AD lesional skin as early as Week 2 with a progressive decrease through Week 16 (P < 0.0001). Dupilumab treatment decreased levels of N(C18)S-ceramides in STS samples similarly in subjects with and without FLG mutations from Week 2 to Week 16. Conclusions Dupilumab treatment normalizes TEWL5 and decreases levels of N(C18)S-ceramides in AD lesional skin of subjects with and without FLG mutations.

520 - Evaluating the expression of OX40 and OX40 ligand in lesional skin versus non-lesional skin in volunteers with atopic dermatitis and in healthy control skin

Abstract Introduction/Background Growing evidence suggests that blocking the OX40 ligand (OX40L)/OX40 costimulatory pathway could have therapeutic effects in inflammatory diseases such as atopic dermatitis (AD). Amlitelimab (SAR445229; KY1005) is a potential first-in-class, fully human, non-depleting anti-OX40L monoclonal antibody that blocks OX40L-OX40 interactions. We report OX40L and OX40 expression from skin biopsies collected in two studies: an amlitelimab Phase 2a (Ph2a) study in patients with moderate-to-severe AD, and a small-scale study of patients with mild AD and healthy volunteers. Objectives Assess OX40L and OX40 expression in AD skin (lesional and non-lesional) and healthy control skin. Methods As part of a Ph2a randomized, double-blind, placebo-controlled study (NCT03754309) of amlitelimab in moderate-to-severe AD, 174 baseline-paired lesional/non-lesional skin biopsies were collected from all patients (n=87). In a separate small-scale study, 24 skin biopsies were collected, including healthy control skin from healthy volunteers (n=6) and paired lesional/non-lesional skin from patients with mild AD (n=9). Each biopsy underwent immunohistochemical (IHC; formalin-fixed paraffin-embedded) analyses using various antibodies (OX40L: rabbit IgG antibody from Cell Signaling Technology for Ph2a biopsies, proprietary mouse IgG2a KY1005 for small-scale study biopsies; OX40: clone Ber-ACT35 from BioLegend for all biopsies). A trained pathologist assessed IHC staining in the dermis and/or epidermis using semi-quantitative scoring (0 [no stained cells] to 10 [all cells stained]) or quantitative methods (cells/mm2, Tissue Studio 4.0, and HALO link image analysis software). Wilcoxon matched-pairs signed-rank test was used to assess IHC staining of Ph2a biopsies. Ph2a biopsies also underwent transcriptomic analyses (stored in RNALater) for OX40L (TNFSF4) and OX40 (TNFRSF4) on sequenced RNA using a linear mixed model with Benjamini-Hochberg adjustment. Results In baseline Ph2a biopsies, IHC analysis revealed elevated OX40L and OX40 staining in lesional versus non-lesional AD skin (P <0.0001 for both markers). For OX40L, the median score was 3 (lesional) versus 1 (non-lesional) and the Wilcoxon matched-pairs signed-rank test found P<0.0001. For OX40+ cells, the median expression was 179.8 cells/mm2 in lesional AD skin versus 54.45 cells/mm2 in non-­lesional AD skin (P<0.0001 using the Wilcoxon matched-pairs signed-rank test). These findings were confirmed by RNA sequencing analysis, with TNFSF4 (OX40L) and TNFRSF4 (OX40) differentially expressed in lesional and non-lesional AD skin (P<0.01 and P<0.0001, respectively). A larger scale study is ongoing that may further support this finding. In the small-scale study including patients with mild AD and healthy volunteers, there was a trend for an increase of mean OX40L+ and OX40+ cells/mm2 in lesional skin versus non-lesional skin and healthy control skin in the dermis and epidermis (OX40L+ dermal lesional, non-lesional, healthy, respectively: 17.6, 11.2, 11.5; OX40L+ epidermal: 5.8, 2.1, 1.9; OX40+ dermal: 5.7, 2.1, 0.4; OX40+ epidermal: 10.9, 9.0, 1.8); the increase was significant for lesional versus healthy control skin for dermal OX40+ cells/mm2 (P<0.001). Conclusions We confirm with IHC and RNA sequencing that OX40L and OX40 expression is elevated in skin lesions of patients with moderate-to-severe AD. We also demonstrate a trend to elevated OX40L and elevated OX40 in mild AD skin versus healthy volunteers. A potential explanation for the lower levels of OX40L and OX40 in patients with mild AD in this study may be their lower disease severity versus Ph2a study patients. These findings support the OX40L/OX40 pathway as a relevant target in T cell immune dysregulation; its blockade may represent a novel treatment approach for moderate-to-severe AD.

Basophils are important for development of allergic skin inflammation

Atopic dermatitis skin lesions exhibit increased infiltration by basophils. Basophils produce IL-4, which plays an important role in the pathogenesis of atopic dermatitis. We sought to determine the role of basophils in a mouse model of antigen-driven allergic skin inflammation. Wild-type mice, mice with selective and inducible depletion of basophils, and mice expressing Il4-driven enhanced green fluorescent protein were subjected to epicutaneous sensitization with ovalbumin or saline. Sensitized skin was examined by histology for epidermal thickening. Cells were analyzed for surface markers and intracellular expression of enhanced green fluorescent protein by flow cytometry. Gene expression was evaluated by real-time reverse transcription-quantitative PCR. Basophils were important for epidermal hyperplasia, dermal infiltration by CD4+ T cells, mast cells, and eosinophils in ovalbumin-sensitized mouse skin and for the local and systemic TH2 response to epicutaneous sensitization. Moreover, basophils were the major source of IL-4 in epicutaneous-sensitized mouse skin and promote the ability of dendritic cells to drive TH2 polarization of naive T cells. Basophils play an important role in the development of allergic skin inflammation induced by cutaneous exposure to antigen in mice.

GSDMD suppresses keratinocyte differentiation by inhibiting FLG expression and attenuating KCTD6-mediated HDAC1 degradation in atopic dermatitis.

Recent studies have shown that activated pyroptosis in atopic dermatitis (AD) switches inflammatory processes and causes abnormal cornification and epidermal barrier dysfunction. Little research has focused on the interaction mechanism between pyroptosis-related genes and human keratinocyte differentiation. The AD dataset from the Gene Expression Omnibus (GEO) was used to identify differently expressed pyroptosis-related genes (DEPRGs). Hub genes were identified and an enrichment analysis was performed to select epithelial development-related genes. Lesions of AD patients were detected via immunohistochemistry (IHC) to verify the hub gene. Human keratinocytes cell lines, gasdermin D (GSDMD) overexpression, Caspase1 siRNA, Histone Deacetylase1 (HDAC1) siRNA, and HDAC1 overexpression vectors were used for gain-and-loss-of-function experiments. Regulation of cornification protein was determined by qPCR, western blot (WB), immunofluorescence (IF), dual-luciferase reporter assay, co-immunoprecipitation (Co-IP), and chromatin immunoprecipitation (ChIP). A total of 27 DEPRGs were identified between either atopic dermatitis non-lesional skin (ANL) and healthy control (HC) or atopic dermatitis lesional skin (AL) and HC. The enrichment analysis showed that these DEPRGs were primarily enriched in the inflammatory response and keratinocytes differentiation. Of the 10 hub genes identified via the protein-protein interaction network, only GSDMD was statistically and negatively associated with the expression of epithelial tight junction core genes. Furthermore, GSDMD was upregulated in AD lesions and inhibited human keratinocyte differentiation by reducing filaggrin (FLG) expression. Mechanistically, GSDMD activated by Caspase1 reduced FLG expression via HDAC1. HDAC1 decreased FLG expression by reducing histone acetylation at the FLG promoter. In addition, GSDMD blocked the interaction of Potassium Channel Tetramerization Domain Containing 6 (KCTD6) and HDAC1 to prohibit HDAC1 degradation. This study revealed that GSDMD was upregulated in AD lesions and that GSDMD regulated keratinocytes via epigenetic modification, which might provide potential therapeutic targets for AD.

Altered Gene Expression of IL-35 and IL-36α in the Skin of Patients with Atopic Dermatitis.

A comprehensive understanding of atopic dermatitis (AD) pathogenesis is desired, especially in the current era of novel biologics and small molecule drugs. In recent years, new cytokines have emerged that may play a significant role in the pathogenesis of AD. Using the tape stripping (TS) method, this study analyzed the gene expression of IL-35 and IL-36α in lesional and nonlesional AD skin compared with healthy skin and their association with the clinical features of AD among the Polish population. Ten AD patients and seven healthy individuals were enrolled. The lesional skin of the AD patients showed significantly higher expression levels of IL-35 compared to healthy skin (p = 0.0001). The expression level of IL-36α was significantly higher in lesional AD skin than in nonlesional AD skin (p = 0.0039) and healthy skin (p = 0.0045). There was a significant negative correlation between AD severity and the expression level of IL-35 in both lesional (R = -0.66, p = 0.048) and nonlesional skin (R = -0.9, p = 0.0016). In summary, both IL-35 and IL-36α appear to play a role in the pathogenesis of AD. Furthermore, it might be speculated that IL-35 and IL-36α may be potential candidates for disease biomarkers. However, further studies are needed to verify these assumptions and comprehensively elucidate their importance in the pathogenesis of AD.

Expression of hornerin in skin lesions of atopic dermatitis and skin diseases.

We have previously identified the filaggrin (FLG)-like protein, hornerin (HRNR). Recently, there have been several reports regarding the relationship between HRNR and atopic dermatitis (AD). In the present study, we examined HRNR expression in the skin lesions of seven unrelated patients with AD to clarify the role of HRNR in the pathogenesis of AD. HRNR was detected in chronic AD lesions (n = 4), whereas no HRNR signals were observed in acute AD lesions (n = 3). HRNR was detected in the cytokeratin 6-expressing epidermis, and Ki67-positive keratinocytes were more abundant in the HRNR-positive epidermis. These findings suggest that HRNR may be associated with epidermal hyperproliferation in AD lesions. Next, we examined HRNR expression in skin diseases associated with hyperkeratosis. HRNR signals were irregularly observed in different cells from those expressing FLG in epidermolytic ichthyosis and actinic keratosis. Therefore, HRNR may play a unique role in the molecular process of cornification.

Blockade of OSMRβ signaling ameliorates skin lesions in a mouse model of human atopic dermatitis.

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by severe pruritus and eczematous skin lesions. Although IL-31, a type 2 helper T (Th2)-derived cytokine, is important to the development of pruritus and skin lesions in AD, the blockade of IL-31 signaling does not improve the skin lesions in AD. Oncostatin M (OSM), a member of IL-6 family of cytokines, plays important roles in the regulation of various inflammatory responses through OSM receptor β subunit (OSMRβ), a common receptor subunit for OSM and IL-31. However, the effects of OSM on the pathogenesis of AD remain to be elucidated. When AD model mice were treated with OSM, skin lesions were exacerbated and IL-4 production was increased in the lymph nodes. Next, we investigated the effects of the monoclonal antibody (mAb) against OSMRβ on the pathogenesis of AD. Treatment with the anti-OSMRβ mAb (7D2) reduced skin severity score in AD model mice. In addition to skin lesions, scratching behavior was decreased by 7D2 mAb with the reduction in the number of OSMRβ-positive neurons in the dorsal root ganglia of AD model mice. 7D2 mAb also reduced the serum concentration of IL-4, IL-13, and IgE as well as the gene expressions of IL-4 and IL-13 in the lymph nodes of AD model mice. Blockade of both IL-31 and OSM signaling is suggested to suppress both pruritus and Th2 responses, resulting in the improvement of skin lesions in AD. The anti-OSMRβ mAb may be a new therapeutic candidate for the treatment of AD.

Clinically uninvolved but not healthy-The skin of patients with atopic dermatitis is primed for itch and inflammation.

Atopic dermatitis (AD) is a highly prevalent inflammatory skin disorder characterized by episodic exacerbations and remissions. Why the clinically healthy skin of AD patients becomes rapidly inflamed and very pruritic is poorly understood. To investigate cowhage- and histamine-induced itch and skin expression levels of their target receptors in lesional and non-lesional skin of AD, compared to the skin of patients with psoriasis, chronic spontaneous urticaria (CSU) and healthy subjects. Patients with AD, psoriasis and chronic spontaneous urticaria (CSU) as well as healthy control subjects (HC) (n = 20 each) were assessed for differences in itch parameters, neurogenic flare reaction and local blood flow responses to skin provocations with cowhage and histamine. Skin biopsies from 10 AD, 10 psoriasis,11 CSU and 12 HC were obtained to assess expression of protease-activated receptors 2 and 4 (PAR-2, PAR-4), histamine H1 and H4 receptors (H1R, H4R), and mast cells. Provocation of non-lesional skin of AD patients with cowhage resulted in prolonged itch (p = 0.020), which was not observed in psoriasis and CSU. Significantly prolonged and more intense cowhage- and histamine-induced itch (for duration, peak and overall intensity) was also observed in lesional AD skin. Diminished neurogenic flare reaction and blood flow after histamine provocation were shown in AD and psoriasis patients. Non-lesional AD skin along with lesional AD and psoriasis skin showed an increased expression of PAR-2 and PAR-4, H1R and H4R. Mast cell number was higher in lesional AD and psoriasis skin (p = 0.006 and p = 0.006, respectively). The non-lesional skin of AD patients markedly differs from healthy skin in cowhage-induced itch responses and the expression of receptors for proteases and histamine. Proactive therapeutic interventions that downregulate these receptors may prevent episodic exacerbation in AD.

Investigating dysregulation of TGF-β1/SMAD3 signaling in atopic dermatitis: a molecular and immunohistochemical analysis.

Atopic dermatitis (AD) is a persistent and recurring inflammatory condition affecting the skin. An expanding corpus of evidence indicates the potential participation of transforming growth factor-β1 (TGF-β1) in the modulation of inflammation and tissue remodeling in AD. The primary objective of this study was to examine the aberrant modulation of TGF-β1/small mothers against decapentaplegic homolog 3 (SMAD3) signaling through a comprehensive analysis of their molecular and protein expression profiles. The study encompassed an aggregate of 37 participants, which included 25 AD patients and 12 controls. The assessment of mRNA and protein levels of TGF-β1 and SMAD3 was conducted utilizing quantitative real-time PCR and immunohistochemistry (IHC), whereas serum IgE and vitamin D levels were estimated by ELISA and chemiluminescence, respectively. Quantitative analysis demonstrated a 2.5-fold upregulation of TGF-β1 mRNA expression in the lesional AD skin (P < 0.0001). IHC also exhibited a comparable augmented pattern, characterized by moderate to strong staining intensities. In addition, TGF-β1 mRNA showed an association with vitamin D deficiency in serum (P < 0.02), and its protein expression was linked with the disease severity (P < 0.01) Furthermore, a significant decrease in the expression of the SMAD3 gene was observed in the affected skin (P = 0.0004). This finding was further confirmed by evaluating the protein expression and phosphorylation of SMAD3, both of which exhibited a decrease. These findings suggest that there is a dysregulation in the TGF-β1/SMAD3 signaling pathway in AD. Furthermore, the observed augmentation in mRNA and protein expression of TGF-β1, along with its correlation with the disease severity, holds considerable clinical significance and emphasizes its potential role in AD pathogenesis.

Unbalanced Aryl Hydrocarbon Receptor Expression in Peripheral and Lesional T Cell Subsets of Atopic Dermatitis.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor, which is involved in the pathogenesis of a variety of skin diseases such as atopic dermatitis (AD). In this study, we aimed to study the AhR-expressing cells in T helper 17 (Th17), T helper 22 (Th22), regulatory T cells (Treg) and B cells in peripheral blood and in AD skin lesions. Twenty AD patients defined according to the Chinese criteria of atopic dermatitis and eighteen healthy subjects were included in our study. The AhR-expressing Th17, Th22, Treg and total B cells in peripheral blood were measured by flow cytometry. The AhR+ Th17 cells and AhR+ Th22 cells in AD skin lesions were measured by immunofluorescence. The mRNA of AhR, interleukin (IL)-22, IL-17A, IL-10, Foxp3, RORγT and TGF-β in peripheral blood mononuclear cells (PBMCs) was measured by real-time quantitative polymerase chain reaction. The expression of AhR in peripheral CD4+ T cells, Th22 cells, Treg cells and total B cells was significantly increased in AD. AhR+IL-17A+ and AhR+IL-22+ lymphocytes were also increased in AD skin lesions. The mRNA levels of AhR, IL-22 and IL-17A in PBMCs in AD patients were significantly higher. AhR mRNA levels in PBMCs positively correlated with peripheral basophil count, peripheral eosinophil count and mRNA levels of IL-22. AhR was highly expressed in subpopulations of CD4+ T cells in peripheral blood and skin lesions of AD, suggesting that AhR might contribute to the pathogenesis of AD.

Histone deacetylase 3 inhibition alleviates 2,4-dinitrochlorobenzene-induced atopic dermatitis via epigenetically upregulating Nrf2/HO-1 signaling pathway

Atopic dermatitis (AD) is a frequent skin disorder that is associated with immune dysfunction and skin inflammation. Histone deacetylase 3 (HDAC3) possesses strong immune and inflammatory modulatory properties in multiple diseases. However, the role and mechanism of HDAC3 in AD remain unknown. Here, we reported that HDAC3 expression was aberrantly upregulated in 2,4-dinitrochlorobenzene (DNCB)-induced lesional AD skin in mice. Inhibition of HDAC3 by RGFP966 protected against DNCB-induced AD, indicated by improved histological damages, relieved inflammatory and immune dysfunction. Nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) signaling pathway activity in lesional AD skin was significantly decreased and RGFP966 attenuated the decrease. Inhibition of Nrf2/HO-1 signaling pathway via Nrf2 inhibitor ML385 blunted anti-AD effect of RGFP966 in DNCB-treated mice. Mechanistically, RGFP966 promoted Nrf2 expression and upregulated H3K27ac deposition on the promoter region of Nrf2. Collectively, HDAC3 inhibition protects against AD via epigenetically activating Nrf2 transcription to upregulate Nrf2/HO-1 signaling pathway activity. HDAC3 may act as a promising therapeutic target for the treatment of AD.

Oncostatin M suppresses IL31RA expression in dorsal root ganglia and interleukin-31-induced itching.

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by intermittent itchy rash. Type 2 inflammatory cytokines such as interleukin (IL)-4, IL-13, and IL-31 are strongly implicated in AD pathogenesis. Stimulation of IL-31 cognate receptors on C-fiber nerve endings is believed to activate neurons in the dorsal root ganglion (DRG), causing itch. The IL-31 receptor is a heterodimer of OSMRβ and IL31RA subunits, and OSMRβ can also bind oncostatin M (OSM), a pro-inflammatory cytokine released by monocytes/macrophages, dendritic cells, and T lymphocytes. Further, OSM expression is enhanced in the skin lesions of AD and psoriasis vulgaris patients. The current study aimed to examine the contributions of OSM to AD pathogenesis and symptom expression. The expression levels of the OSM gene (OSM) and various cytokine receptor genes were measured in human patient skin samples, isolated human monocytes, mouse skin samples, and mouse DRG by RT-qPCR. Itching responses to various pruritogens were measured in mice by counting scratching episodes. We confirmed overexpression of OSM in skin lesions of patients with AD and psoriasis vulgaris. Monocytes isolated from the blood of healthy subjects overexpressed OSM upon stimulation with IL-4 or GM-CSF. Systemic administration of OSM suppressed IL31RA expression in the mouse DRG and IL-31-stimulated scratching behavior. In contrast, systemic administration of OSM increased the expression of IL-4- and IL-13-related receptors in the DRG. These results suggest that OSM is an important cytokine in the regulation of skin monocytes, promoting the actions of IL-4 and IL-13 in the DRG and suppressing the action of IL-31. It is speculated that OSM released from monocytes in skin modulates the sensitivity of DRG neurons to type 2 inflammatory cytokines and thereby the severity of AD-associated skin itch.

Ethnic endotypes in paediatric atopic dermatitis depend on immunotype, lipid composition and microbiota of the skin.

Atopic dermatitis (AD) endotypes differ with ethnicity. We examined the skin microbiota, cytokine and lipid profiles in Greenlandic Inuit and Danish children with AD. Twenty-five Inuit children with AD and 25 Inuit control children were clinically examined and compared to previously collected data from 25 Danish children with AD. Skin tape strips and skin swabs were collected from lesional and non-lesional skin. Levels of cutaneous immune biomarkers, free sphingoid bases and their (glycosyl)ceramides were analysed. Skin swabs were analysed with 16S rRNA and tuf gene for characterization of bacterial species communities. Bacterial β-diversity was significantly different between Inuit and Danish AD skin, in both lesional (p < 0.001) and non-lesional (p < 0.001) AD skin, and there was a higher relative abundance of Staphylococcus aureus in Danish compared to Inuit lesional (53% vs. 8%, p < 0.01) and non-lesional skin (55% vs. 5%, p < 0.001). Danish AD children had a higher α-diversity than Inuit children in non-lesional (p < 0.05) but not in lesional skin. Significantly higher levels of type 2 immunity cytokine interleukin (IL)-4 (p < 0.05) and IL-5 (p < 0.01) were identified in Inuit compared to Danish AD children. In contrast, IL-33 (p < 0.01) was higher in Danish lesional and non-lesional AD skin. Higher levels of long-chain glucosylceramide (GlcCER)[S](d26:1) were found in lesional (p < 0.001) and non-lesional (p < 0.001) Inuit skin compared with Danish AD skin. NMF levels were similar in Inuit and Danish AD skin. Skin microbiota, cytokine and lipid composition differed significantly between Inuit and Danish children with AD and showed a stronger type 2 immune signature in Inuit children.