Heparinoid suppresses Der p‐induced IL‐1β production by inhibiting ERK and p38 MAPK pathways in keratinocytes

Epidermal keratinocytes initiate skin inflammation by activating immune cells. The skin barrier is disrupted in atopic dermatitis (AD) and epidermal keratinocytes can be exposed to environmental stimuli, such as house dust mite (HDM) allergens. We showed previously that HDM allergens activate the NLRP3 inflammasome of keratinocytes, thereby releasing pro‐inflammatory cytokines. Heparinoid is an effective moisturizer for atopic dry skin. However, a recent report showed that heparinoid treatment can improve inflammation of lichen planus. Therefore, we hypothesized that it acts on epidermal keratinocytes not only as a moisturizer, but also as a suppressant of the triggers of skin inflammation. We found that HDM allergen‐induced interleukin (IL)‐1β release from keratinocytes was inhibited significantly by heparinoid pretreatment without affecting cell viability. However, heparinoid did not affect caspase‐1 release, suggesting that heparinoid did not affect HDM allergen‐induced inflammasome activation. Heparinoid treatment not only decreased intracellular levels of pro‐IL‐1β, but also suppressed IL‐1β messenger RNA (mRNA) expression in keratinocytes. Among the intracellular signalling pathways, the activation of extracellular signal‐regulated kinase and p38 pathways, which are required for IL‐1β expression in keratinocytes, was inhibited by heparinoid treatment. The inhibitory effect of heparinoid on IL‐1β mRNA expression was also confirmed with living skin equivalents. Our results demonstrated that heparinoid suppresses the initiation of keratinocyte‐mediated skin inflammation.     

IL‐17A synergistically enhances TLR3‐mediated IL‐36γ production by keratinocytes: A potential role in injury‐amplified psoriatic inflammation

Skin injury can trigger formation of new lesions in psoriasis (Koebner phenomenon). The mechanisms through which injury exacerbates psoriasis are unclear. During wound repair, epidermal keratinocytes are activated and produce abundant IL‐36γ, further promoting the skin inflammation. IL‐17A is the cornerstone cytokine in the pathogenesis of psoriasis. We sought to investigate the effects of IL‐17A on injury‐induced keratinocyte activation and IL‐36γ production. Here, we demonstrated that dsRNA released from necrotic keratinocytes induced the expression of IL‐36γ. Silencing of TLR3 by siRNA decreased the IL‐36γ induction by necrotic keratinocyte supernatant. Co‐stimulation with dsRNA and IL‐17A synergistically increased the expression of IL‐36γ and other proinflammatory mediators (CCL20, CXCL8, DEFB4 and LCN2) in keratinocytes. The synergistic effects were not dependent on TLR3 upregulation, TNF receptor signalling and mRNA stabilization. Co‐stimulation with dsRNA and IL‐17A resulted in an accumulation of IκBζ. The synergistic upregulation of IL‐36γ and proinflammatory mediators were inhibited by IκBζ siRNA. Co‐stimulation with IL‐17A and poly(I:C) markedly activated the p38 MAPK and NF‐κB pathway, compared with poly(I:C). Blockade of p38 MAPK and NF‐κB suppressed dsRNA/IL‐17A–mediated IκBζ and IL‐36γ induction. These findings demonstrated that IL‐17A synergistically enhanced the dsRNA‐mediated IL‐36γ production through a p38 MAPK‐, NF‐κB–, and IκBζ‐dependent mechanism.     

Malassezia yeasts activate the NLRP3 inflammasome in antigen‐presenting cells via Syk‐kinase signalling

Although being a normal part of the skin flora, yeasts of the genus Malassezia are associated with several common dermatologic conditions including pityriasis versicolour, seborrhoeic dermatitis (SD), folliculitis, atopic eczema/dermatitis (AE/AD) and dandruff. While Malassezia spp. are aetiological agents of pityriasis versicolour, a causal role of Malassezia spp. in AE/AD and SD remains to be established. Previous reports have shown that fungi such as Candida albicans and Aspergillus fumigatus are able to efficiently activate the NLRP3 inflammasome leading to robust secretion of the pro‐inflammatory cytokine IL‐1β. To date, innate immune responses to Malassezia spp. are not well characterized. Here, we show that different Malassezia species could induce NLRP3 inflammasome activation and subsequent IL‐1β secretion in human antigen‐presenting cells. In contrast, keratinocytes were not able to secrete IL‐1β when exposed to Malassezia spp. Moreover, we demonstrate that IL‐1β secretion in antigen‐presenting cells was dependent on Syk‐kinase signalling. Our results identify Malassezia spp. as potential strong inducers of pro‐inflammatory responses when taken up by antigen‐presenting cells and identify C‐type lectin receptors and the NLRP3 inflammasome as crucial actors in this process.     

Evidence for a regulatory loop between IFN‐γ and IL‐33 in skin inflammation

Interleukin‐33 has recently gained much attention due to its role in allergic responses. It has been shown to amplify Th2 responses and to act as a damage‐associated molecular pattern. IL‐33 acts on a broad range of cells and has been proposed to link innate and adaptive features of allergic responses. It was the aim of this study to investigate this property of IL‐33 in the inflammatory response characterising atopic dermatitis (AD). We have analysed the response of skin‐resident cells derived from patients with AD and healthy donors with regard to the expression of IL‐33 and its receptor ST2. The functional impact of IL‐33 on CD4+ T cells was investigated. Keratinocytes and dermal fibroblasts clearly differ in their regulation of IL‐33. In fibroblasts, the concerted action of TNF‐α and IL‐1β was the strongest inducer, whereas IFN‐γ is clearly the key molecule that upregulates IL‐33 in keratinocytes with a more pronounced response of cells derived from patients with AD. Keratinocytes from patients with AD showed a markedly higher constitutive expression level of surface ST2. CD4+ T cells respond to IL‐33. Unexpectedly, IL‐33 failed to induce a significant secretion of IL‐5 or IL‐13. By contrast, high amounts of IFN‐γ were detectable if IL‐33 was added to the T‐cell receptor‐stimulated cells or in combination with IL‐12. These results suggest that IL‐33 and IFN‐γ are closely interlinked in epidermal AD inflammation. IFN‐γ induces IL‐33 in keratinocytes and IL‐33 acts on activated T cells to further increase the release of IFN‐γ, therefore contributing to drive skin inflammation towards chronic responses.     

S‐allyl cysteine inhibits TNF‐α‐induced inflammation in HaCaT keratinocytes by inhibition of NF‐ κB‐dependent gene expression via sustained ERK activation

Tumor necrosis factor‐α (TNF‐α)‐induced keratinocyte inflammation plays a key role in the pathogenesis of multiple inflammatory skin diseases. Here we investigated the anti‐inflammatory effect of S‐allyl cysteine (SAC) on TNF‐α‐induced HaCaT keratinocyte cells and the mechanism behind its anti‐inflammatory potential. SAC was found to inhibit TNF‐α‐stimulated cytokine expression. Further, SAC was found to inhibit TNF‐α‐induced activation of p38, JNK and NF‐κB pathways. Interestingly, SAC was found to differentially regulate ERK MAP kinase in cells. TNF‐α‐induced transient ERK activation and SAC treatment resulted in sustained ERK activation both in the presence and absence of TNF‐α. Additionally, SAC failed to inhibit the TNF‐α‐induced expression of the pro‐inflammatory cytokines TNF‐α and IL‐1β when cells were treated with the MEK inhibitor PD98059, suggesting that the anti‐inflammatory effect of SAC is via sustained activation of the ERK pathway. Since ERK activation has been reported to negatively regulate NF‐κB‐driven gene expression and we find that SAC activates ERK and negatively regulates NF‐κB, we investigated whether there existed any crosstalk between the ERK and the NF‐κB pathways. NF‐κB‐dependent reporter assay, visualization of the nuclear translocation of NF‐κB‐p65 subunit and determination of the cellular levels of I‐κB, the inhibitor of NF‐κB, revealed that SAC inhibited TNF‐α‐induced NF‐κB activation, and PD98059 treatment reversed this effect. These results collectively suggest that SAC inhibits TNF‐α‐induced inflammation in HaCaT cells via a combined effect entailing the inhibition of the p38 and the JNK pathways and NF‐κB pathway via the sustained activation of ERK.     

Dual oxidase 2 is essential for house dust mite‐induced pro‐inflammatory cytokine production in human keratinocytes

House dust mites (HDMs) are known to trigger chronic inflammation through Toll‐like receptors (TLRs) and their signalling cascades. In this study, we found that TLR2 ligation by HDMs induced the activation of dual oxidase 2 (Duox2) and nuclear factor‐κB (NF‐κB), leading to the production of pro‐inflammatory cytokines in human keratinocytes. Stimulation of human keratinocytes with HDMs resulted in increases in interleukin‐8 (IL‐8) and chemokine (C–C motif) ligand 20 (CCL20) levels. However, pro‐inflammatory cytokine production was abolished in keratinocytes transfected with TLR2 siRNA, indicating that HDM‐induced cytokine production was mediated via TLR2 signalling. We also examined the function of Duox1/2 isozymes, which are primarily expressed in keratinocytes, in HDM‐mediated pro‐inflammatory cytokine production. Human keratinocytes transfected with control siRNA or Duox1 siRNA showed no inhibition of IL‐8 or CCL20 production in response to HDMs, whereas the silencing of Duox2 expression resulted in a failure to induce cytokine production. Moreover, the phosphorylation and nuclear localization of RelA/p65, a component of NF‐κB, were induced by HDMs in human keratinocytes. Transfection of human keratinocytes with TLR2 siRNA or Duox2 siRNA resulted in the complete abolishment of RelA/p65 nuclear localization in response to HDMs. Taken together, these results indicate that the HDM‐dependent TLR2‐Duox2 signalling axis indeed promotes NF‐κB activation, which induces IL‐8 and CCL20 production and mediates epidermal keratinocyte inflammation.     

Intercellular pathway through hyaluronic acid in UVB‐induced inflammation

Ultraviolet B (UVB) radiation induces inflammation in the skin specifically at the site of exposure. We unexpectedly found that UVB‐induced inflammation was not induced in gp91phox‐depleted mice. To test whether gp91phox is directly involved in UVB‐induced inflammation, neutrophil‐ and hyaluronic acid–depleted mice were also irradiated and examined for their response. Hyaluronic acid–depleted mice showed strongly inhibited UVB‐induced inflammation, but the neutrophil‐depleted mice did not exhibit any suppressed UVB‐induced inflammation. To elucidate the pathway by which UVB irradiation induced inflammation, we examined the expression of nucleotide‐binding domain, leucine‐rich‐containing family, pyrin domain‐containing‐3 (NLRP3) and caspase‐1 in the mouse skin. An increase in the expression of NLRP3 and caspase‐1 was seen following the UVB irradiation of C57BL mice; however, the UVB‐irradiated gp91phox‐knockout (gp91phox^?/?) mice did not have this increase in expression. Furthermore, the plasma IL‐1β level increased after the UVB irradiation in C57BL mice, but there was no change in the gp91phox^?/? mice. These results clearly indicate that nicotinamide adenine dinucleotide phosphate oxidase is activated by gp91phox, which is expressed on the surface in response to the increased expression of hyaluronic acid induced by UVB irradiation, and as result, the generation of reactive oxygen species (ROS) increases. This ROS activate NLRP3, and NLRP3 leads to the production of caspase‐1, which subsequently increases IL‐1β, thereby finally inducing inflammation. It is thought that this system may play an important role in the damage and ageing of skin, and further studies are necessary to confirm these finding.     

High calcium enhances the expression of double‐stranded RNA sensors and antiviral activity in epidermal keratinocytes

Double‐stranded RNA (dsRNA) sensors including TLR3, MDA5 and RIG‐I are expressed in epidermal keratinocytes and play an important immunological role by enhancing various innate and adaptive immune responses. Although the role of elevated extracellular calcium concentration in keratinocyte differentiation is well understood, the effect of high calcium on dsRNA sensors is not well studied. We investigated alterations in dsRNA sensor expression and antiviral activity induced by a high extracellular concentration of calcium in epidermal keratinocytes. Normal human epidermal keratinocytes (NHEKs) were stimulated with high calcium and/or synthetic dsRNA, poly (I:C). TLR3, IFIH1 (MDA5) and DDX58 (RIG‐I) expression were measured via qPCR, and IFN‐β and human beta‐defensin 2 (HBD2) levels were measured using ELISA. TLR3 localization was evaluated with immunocytofluorescence. Antiviral activity was quantified with virus plaque assays using herpes simplex virus type 1 (HSV‐1). High calcium significantly upregulated mRNA expression of TLR3, IFIH1 and DDX58 in NHEKs. In addition, high calcium significantly enhanced poly (I:C)‐induced anti‐HSV‐1 activity in NHEKs. The antiviral molecule HBD2 but not IFN‐β induction by poly (I:C) was enhanced by high calcium. Our findings indicate that high levels of extracellular calcium enhance the expression of dsRNA sensors and augment antiviral activity in epidermal keratinocytes.     

Interleukin‐1 from epithelial cells fosters T cell‐dependent skin inflammation

Background Chronic inflammatory skin diseases such as atopic dermatitis and psoriasis are characterized by the infiltration of lymphocytes into the epidermal compartment. Several studies point to an active role of skin epithelial cells in the pathophysiology of such diseases. Objectives In this study we addressed the regulatory function of primary human keratinocytes in the interaction with autologous T cells and monocytes. Methods We used a human coculture model with keratinocytes grown from epidermal stem cells of the outer root sheath of human hair follicles and autologous T cells. Results In our coculture system we observed a high production of interferon (IFN)‐γ, but not Th2 cytokines, in the presence of superantigen or antigen‐pulsed autologous monocytes. Critical parameters for this effect were: (i) T‐cell receptor activation, (ii) an intercellular adhesion molecule‐1/lymphocyte function‐associated antigen (LFA)‐1‐dependent interaction between keratinocytes and T cells, and (iii) secretion of interleukin (IL)‐1β. Remarkably, in the presence of activated T cells, epithelial cells seemed to be a more significant source of IL‐1β than monocytes. Application of the LFA‐1 blocker efalizumab or IL‐1 receptor antagonist anakinra enabled us to suppress completely the production of IFN‐γ by T cells in the coculture. Conclusions IL‐1 secretion and the physical contact between keratinocytes and activated, infiltrating T cells may be central for the development of chronic inflammatory skin conditions.     

Antitumor activity of sulforaphane in mice model of skin cancer via blocking sulfatase‐2

Although there are many treatment options for skin cancer, the chemotherapeutic agents for skin cancer are linked with many adverse effects as well as the development of multidrug resistance. Sulforaphane is an isothiocyanate, which is found in cruciferous vegetables. Consumption of sulforaphane‐rich diet has been linked to inhibition of UV‐exposed skin carcinogenesis. Therefore, the goal of this study was to determine the ability of sulforaphane to reduce skin cancer in mice through inhibition of sulfatase‐2 enzyme. Epicutaneous application of 7,12‐dimethylbenz (a) anthracene was performed on the shaved dorsal skin of mice followed by croton oil. Sulforaphane (9 μmol/mouse/day) was administered to mice orally. Skin was removed from the dorsal area for assessment of sulfatase‐2, glypican‐3, heparan sulphate proteoglycans (HSPGs), nuclear factor (NF)κB, nuclear factor E2‐related factor 2 (Nrf2), tumor necrosis factor (TNF)‐α, IL‐1β and caspase‐3. In addition, skin sections were stained with haematoxylin/eosin, Mallory and cytokeratin immunostaining. We found that, sulforaphane blocked sulfatase‐2 activity, leading to significant elevation in HSPGs as well as significant reduction in glypican‐3. In addition, sulforaphane significantly activated Nrf2 and reduced both the gene and protein expression of NFκB, TNF‐α, IL‐1β and caspase‐3. In parallel, stained sections obtained from skin cancer mice treated with sulforaphane showed significant reduction in hyperkeratosis, acanthosis and epithelial dysplasia. The collective results indicate that sulforaphane suppresses skin cancer via blocking sulfatase‐2 with subsequent elevation in HSPGs and reduction in glypican‐3. Moreover, sulforaphane attenuated skin cancer‐induced activation of inflammatory and apoptotic pathways.     

IL‐17F regulates psoriasis‐associated genes through IκBζ

Psoriasis is a common chronic inflammatory and immune‐mediated skin disease. Antagonists of TNF‐α and, recently, IL‐17 have proven to be highly effective in the treatment for psoriasis; however, the molecular mechanisms involved in the pathogenesis of psoriasis are poorly understood. Recently, we presented evidence that IκBζ is a key regulator in the development of psoriasis through its role in mediating IL‐17A‐driven effects. Like IL‐17A, IL‐17F is produced by a variety of immune cells, and the expression of IL‐17F is increased in psoriatic skin. The purpose of this study was to characterize the role of IL‐17F in the regulation of IκBζ expression and to investigate whether IL‐17F regulates psoriasis‐associated genes in human keratinocytes through IκBζ. Here, we demonstrate that IL‐17F stimulation induces IκBζ expression at both the mRNA and the protein levels in normal human keratinocytes. Moreover, silencing IκBζ by siRNA revealed that IκBζ is a key regulator of specific IL‐17F‐inducible psoriasis‐associated genes and proteins, including DEFB4/hBD2, S100A7, CCL20, IL‐8 and CHI3L1. In addition, IL‐17F‐induced IκBζ expression is mediated by a mechanism involving the p38 MAPK and NF‐κB signalling pathways, as shown by the clear reduction in IL‐17F‐mediated expression of IκBζ during chemical inhibition of these two signalling pathways. In summary, we present IκBζ as a novel key regulator of IL‐17F‐driven effects in psoriasis. Thus, antagonists to IκBζ could potentially provide a more targeted approach for treating psoriasis as well as for treating the other inflammatory and immune‐mediated diseases for which IL‐17‐targeting drugs have recently been approved.     

Therapeutic potential of topically administered γ‐AlOOH on 2,4‐dinitrochlorobenzene‐induced atopic dermatitis‐like lesions in Balb/c mice

Boehmite (γ‐AlOOH) has a wide range of applications in a variety of industrial and biological fields. However, little is known about its potential roles in skin diseases. The current study investigated its effect on atopic dermatitis (AD). Following characterization, cytotoxicity, pro‐inflammatory response and oxidative stress associated with boehmite were assessed, using TNF‐α‐induced keratinocytes and mast cells. In addition, therapeutic effects of boehmite, topically administered to Balb/c mice induced by 2,4‐dinitrochlorobenzene (DNCB), were evaluated. Expression of cytokines (TLSP, IL‐25 and IL‐33) and the generation of ROS from keratinocytes induced by TNF‐α were significantly inhibited by boehmite without affecting cell viability. MAPKs (ERK, JNK and p38) required for cytokine expression were suppressed by boehmite treatment. Up‐regulation of cytokines (TSLP, IL‐4, IL‐5, IL‐13, RANTES) in human mast cells treated with phorbol 12‐myristate 13‐acetate and calcium ionophore was also suppressed by boehmite. Boehmite improved the AD severity score, epidermal hyperplasia and transepidermal water loss in DNCB‐induced AD‐like lesions. Moreover, Th2‐mediated cytokine expression, mast cell hyperplasia and destruction of the skin barrier were improved by boehmite treatment. Overall, we demonstrated that boehmite may potentially protect against AD.     

Inhibition of Poly(I:C)-Induced Inflammation by Salvianolic Acid A in Skin Keratinocytes

BackgroundSkin keratinocytes participate actively in inducing immune responses when external pathogens are introduced, thereby contributing to elimination of pathogens. However, in condition where the excessive inflammation is occurred, chronic skin disease such as psoriasis can be provoked.ObjectiveWe tried to screen the putative therapeutics for inflammatory skin disease, and found that salvianolic acid A (SAA) has an inhibitory effect on keratinocyte inflammatory reaction. The aim of this study is to demonstrate the effects of SAA in poly(I:C)-induced inflammatory reaction in skin keratinocytes.MethodsWe pre-treated keratinocytes with SAA then stimulated with poly(I:C). Inflammatory reaction of keratinocytes was verified using real-time polymerase chain reaction, enzyme-linked immunosorbent assay and Western blot.ResultsWhen skin keratinocytes were pre-treated with SAA, it significantly inhibited poly (I:C)-induced expression of inflammatory cytokines including interleukin (IL)-1β, IL-6, IL-8, tumor necrosis factor-α, and CCL20. SAA inhibited poly(I:C)-induced activation of nuclear factor-κB signaling. And SAA also inhibited inflammasome activation, evidenced by decrease of IL-1β secretion. Finally, SAA markedly inhibited poly(I:C)-induced NLRP3 expression.ConclusionThese results demonstrate that SAA has an inhibitory effect on poly(I:C)-induced inflammatory reaction of keratinocytes, suggesting that SAA can be developed for the treatment of inflammatory skin diseases such as psoriasis.     

Interleukin‐17A affects extracellular vesicles release and cargo in human keratinocytes

Psoriasis is a chronic inflammatory systemic disease caused by deregulation of the interleukin‐23/‐17 axis that allows the activation of Th17 lymphocytes and the reprogramming of keratinocytes proliferative response, thereby inducing the secretion of cyto‐/chemokines and antimicrobial peptides. Beside cell‐to‐cell contacts and release of cytokines, hormones and second messengers, cells communicate each other through the release of extracellular vesicles containing DNA, RNA, microRNAs and proteins. It has been reported the alteration of extracellular vesicles trafficking in several diseases, but there is scarce evidence of the involvement of extracellular vesicles trafficking in the pathogenesis of psoriasis. The main goal of the study was to characterize the release, the cargo content and the capacity to transfer bioactive molecules of extracellular vesicles produced by keratinocytes following recombinant IL‐17A treatment if compared to untreated keratinocytes. A combined approach of standard ultracentrifugation, RNA isolation and real‐time RT‐PCR techniques was used to characterize extracellular vesicles cargo. Flow cytometry was used to quantitatively and qualitatively analyse extracellular vesicles and to evaluate cell‐to‐cell extracellular vesicles transfer. We report that the treatment of human keratinocytes with IL‐17A significantly modifies the extracellular vesicles cargo and release. Vesicles from IL‐17A‐treated cells display a specific pattern of mRNA which is undid by IL‐17A neutralization. Extracellular vesicles are taken up by acceptor cells irrespective of their content but only those derived from IL‐17A‐treated cells enable recipient cells to express psoriasis‐associated mRNA. The results imply a role of extracellular vesicles in amplifying the pro‐inflammatory cascade induced in keratinocyte by pro‐psoriatic cytokines.     

3‐O‐Laurylglyceryl ascorbate improves the development of sensitive skin through the reduction of oxidative stress

Skin sensitivity is a serious problem for many people, and it can be induced by various factors such as UV irradiation, physical and mental stresses, air pollution, dry air and so on. Skin dryness triggered by UV and dry air is one of the most important causes inducing the development of sensitive skin, and it has been reported that oxidative stress contributes to skin dryness. In this study, we investigated whether treatment with 3‐O‐laurylglyceryl ascorbate (VC‐3LG), which is an amphipathic ascorbic acid derivative, can suppress the development of sensitive skin. The results demonstrate that VC‐3LG restores the expression levels of interleukin‐1α, nerve growth factor and matrix metalloprotease‐9 in the dry skin models of reconstructed human epidermal equivalents (RHEEs) and in H₂O₂‐treated keratinocytes. In addition, VC‐3LG suppresses the dendrite elongation of nerve cells induced in RHEEs by dry skin conditions and by H₂O₂ treatment of keratinocytes. Therefore, we consider that treatment of the skin with VC‐3LG is an effective approach to improve the development of sensitive skin.     

Tropisetron via α7 nicotinic acetylcholine receptor suppresses tumor necrosis factor‐α‐mediated cell responses of human keratinocytes

Tropisetron is a serotonin receptor (5‐HT‐R)‐modulating agent and approved as an antiemetic for patients undergoing chemotherapy. In the gut, it acts via specific serotonin receptors, 5‐HT₃‐R, to elicit its beneficial effects against nausea. We investigated whether tropisetron can affect inflammatory cell responses of human primary epidermal keratinocytes (NHK) which are key cells in the regulation of skin homoeostasis. Tropisetron significantly and dose‐dependently suppressed tumor necrosis factor (TNF)‐α‐mediated mRNA expression and protein secretion of interleukin (IL)‐6 and IL‐8 in these cells. This effect of tropisetron was independent of p65/NF‐κB as shown by various NF‐κB signal transduction read‐outs. Importantly, the anti‐inflammatory tropisetron effect on NHK was neither mediated by 5‐HT₃‐R nor 5‐HT₄‐R since these receptors were absent in NHK. In contrast, NHK expressed α7 nicotinic acetylcholine receptors (α7nAchR) which previously were found to bind tropisetron. The α7nAchR antagonist α‐bungarotoxin neutralized, whereas AR‐R17779, a specific α7nAchR agonist, mimicked the suppressive effect of tropisetron on TNF‐α‐mediated IL‐6 and IL‐8 expression in NHK. Our findings suggest that tropisetron and probably other α7nAchR‐activating agents could be useful for the future therapy of inflammatory skin diseases.     

Staphylococcal α‐toxin induces a functional upregulation of TLR‐2 on human peripheral blood monocytes

Resistance to bacterial skin infections, for example with Staphylococcus aureus (S. aureus), is based on the function of intact innate immune mechanisms. Toll‐like receptor (TLR)‐2 recognizes components of S. aureus and is known to be expressed on monocytes. Staphylococcal exotoxins such as staphylococcal enterotoxin B (SEB) or α‐toxin are produced by many S. aureus strains. To investigate TLR‐2 regulation and function on human monocytes upon stimulation with staphylococcal exotoxins to elucidate a putative feedback loop between different staphylococcal components. Monocytes were stimulated with α‐toxin or SEB, respectively. TLR‐2 expression and regulation as well as functional effects of TLR‐2 stimulation with Pam3Cys (TLR‐2/TLR‐1), lipoteichoic acid (LTA) (TLR‐2/TLR‐6) and peptidoglycan (PGN) (TLR‐2 and Nod) were then investigated both at the mRNA and protein level and compared to monocytes from patients with psoriasis. α‐toxin significantly upregulated TLR‐2 expression. TLR‐2 mediated IL‐1β, IL‐6 and IL‐8 secretion was significantly augmented after upregulation with staphylococcal exotoxins. CD36 expression was significantly more downregulated after TLR‐2 upregulation with SEB and consecutive LTA stimulation and TLR‐2 upregulation with α‐toxin following LTA and PGN stimulation, respectively. PGN enhanced CD54 expression after upregulation of the receptor with α‐toxin. Expression of HLA‐DR was unaffected. However, no differences were observed in monocytes from psoriasis patients compared to healthy controls. Together, our findings provide a new link between staphylococcal α‐toxin and TLR‐2 signalling in monocytes which may have implications for skin diseases where skin colonization with S. aureus and dysregulation of TLR‐2 have been described.     

Genetic support for the role of the NLRP3 inflammasome in psoriasis susceptibility

NACHT leucine‐rich repeat‐ and PYD‐containing (NLRP)3 protein controls the inflammasome by regulating caspase‐1 activity and interleukin (IL)‐1β processing. The contribution of IL‐1β in the pathogenesis of psoriasis is well recognized. Polymorphisms in NLRP3 and caspase recruitment domain–containing protein (CARD)8, a negative regulator of caspase‐1 activity, have been associated with susceptibility to common inflammatory diseases, such as Crohn's disease and rheumatoid arthritis. To investigate the role for genetic variants in the NLRP3 inflammasome in psoriasis susceptibility. In a patient sample comprising 1988 individuals from 491 families and 1002 healthy controls, genotypes for four selected single‐nucleotide polymorphisms (SNPs) in NLRP3 (three SNPs) and CARD8 (one SNP) were determined by TaqMan^® Allelic Discrimination. Using the transmission disequilibrium test (TDT), a significant increase in the transmission of the NLRP3 rs10733113G genotype to a subgroup of patients with more widespread psoriasis was demonstrated (P = 0.015). Using logistic regression analysis in 741 patients with psoriasis and 1002 controls, the CARD8 rs2043211 genotype was significantly different in cases and controls in overall terms [OR 1.3 (1.1–1.5), P = 0.004] and for both genders. Our data support the hypothesis that the inflammasome plays a role in psoriasis susceptibility.     

Cytosolic dsDNA triggers apoptosis and pro‐inflammatory cytokine production in normal human melanocytes

Considerable evidence implicates that viral infection might be a participant factor in the pathogenesis of vitiligo. However, it is still unclear how viral infection leads to the melanocyte destruction. To elucidate the effects of viral dsDNA on the viability and cytokine synthesis of normal human melanocytes and to explore the underlying mechanisms, primary cultured normal human melanocytes were transfected with poly(dA:dT). The results demonstrated that poly(dA:dT) triggered apoptosis instead of pyroptosis in melanocytes. Knocking down AIM2 or RIG‐I by RNA interference partially reduced the poly(dA:dT)‐induced LDH release, suggesting the involvement of both nucleic acid sensors in the process of melanocyte death. Poly(dA:dT) induced the expression of pro‐inflammatory cytokine genes including IFN‐β, TNF‐α, IL‐6 and IL‐8 as well, whereas the pro‐inflammatory cytokine production was suppressed by RIG‐I siRNA, but not by AIM2 siRNA. Poly(dA:dT) treatment increased the phosphorylation of p38 and JNK and NFκB. Accordingly, NFκB inhibitor Bay 11‐7082 and JNK inhibitor SP600125 blocked the induction of the cytokine genes except IFN‐β. The production of IL6 and IL8 was also suppressed by p38 inhibitor SB203580. On the contrary, the Poly(dA:dT)‐induced melanocyte death was only decreased by SP600125. This study provides the possible mechanism of melanocyte destruction and immuno‐stimulation in vitiligo by innate immune response following viral infection.