CLDN1 knock out keratinocytes as a model to investigate multiple skin disorders.

The transmembrane protein claudin-1 is critical for formation of the epidermal barrier structure called tight junctions (TJ) and has been shown to be important in multiple disease states. These include neonatal ichthyosis and sclerosing cholangitis syndrome, atopic dermatitis and various viral infections. To develop a model to investigate the role of claudin-1 in different disease settings, we used CRISPR/Cas9 to generate human immortalized keratinocyte (KC) lines lacking claudin-1 (CLDN1 KO). We then determined whether loss of claudin-1 expression affects epidermal barrier formation/function and KC differentiation/stratification. The absence of claudin-1 resulted in significantly reduced barrier function in both monolayer and organotypic cultures. CLDN1 KO cells demonstrated decreases in gene transcripts encoding the barrier protein filaggrin and the differentiation marker cytokeratin-10. Marked morphological differences were also observed in CLDN1 KO organotypic cultures including diminished stratification and reduced formation of the stratum granulosum. We also detected increased proliferative KC in the basale layer of CLDN1 KO organotypic cultures. These results further support the role of claudin-1 in epidermal barrier and suggest an additional role of this protein in appropriate stratification of the epidermis.

Myricetin treatment has ameliorative effects in DNFB-induced atopic dermatitis mice under high-fat conditions.

Atopic dermatitis (AD) is a chronic, inflammatory cutaneous disorder. Obesity is associated with increased prevalence and severity of AD for reasons that remain poorly understood. Myricetin, a dietary flavonoid found in fruits and vegetables, is known to have anti-inflammatory effects, but its role in AD is unclear. Thus, we investigated the effects of obesity on exacerbation AD lesions and evaluated the effects of myricetin on obese AD. Mice were fed normal diet (ND) or high-fat diet, and then 2,4-dinitrofluorobenzene was used to induce AD-like lesions. We found that obesity exacerbated AD lesions, and myricetin topical administration ameliorated symptoms and skin lesions of obsess AD mice, such as dermatitis scores, scratching behavior, epidermal thickness, and mast cell infiltration. In addition, myricetin reduced the levels of immunoglobulin E and histamine, inhibited the infiltration of CD4+T cells, and modulated the expression of Th1, Th2, Th17, and Th22 cytokines and pro-inflammatory factors (CCL17, CCL22, IL-1ß, and TGF-ß). Moreover, myricetin restored impaired barrier function by reducing transepidermal water loss, increasing lamellar body secretion, as well as upregulating the mRNA and protein expression of filaggrin. Western blot results showed that significantly increased levels of phosphorylated IκB and NF-κB p65 was observed in the obese AD mice compared with the AD mice fed ND, whereas the myricetin could downregulated the phosphorylations of IκB and NF-κB, and inhibited mRNA expression of iNOS and COX2. Taken together, our results suggest that myricetin treatment exhibits potentially protective effects against the obeseassociated AD by inhibiting inflammatory response and restoring skin barrier function.

Filaggrin and cytokines in respiratory samples of preterm infants at risk for respiratory viral infection.

Respiratory viral infections (RVIs) are frequent in preterm infants possibly inducing long-term impact on respiratory morbidity. Immune response and respiratory barriers are key defense elements against viral insults in premature infants admitted to Neonatal Intensive Care Units (NICUs). Our main goals were to describe the local immune response in respiratory secretions of preterm infants with RVIs during NICU admission and to evaluate the expression and synthesis of lung barrier regulators, both in respiratory samples and in vitro models. Samples from preterm infants that went on to develop RVIs had lower filaggrin gene and protein levels at a cellular level were compared to never-infected neonates (controls). Filaggrin, MIP-1α/CCL3 and MCP-1 levels were higher in pre-infection supernatants compared to controls. Filaggrin, HIF-1α, VEGF, RANTES/CCL5, IL-17A, IL-1ß, MIP-1α and MIP-1ß/CCL5 levels were higher during and after infection. ROC curve and logistic regression analysis shows that these molecules could be used as infection risk biomarkers. Small airway epithelial cells stimulated by poly:IC presented reduced filaggrin gene expression and increased levels in supernatant. We conclude that filaggrin gene and protein dysregulation is a risk factor of RVI in newborns admitted at the NICU.

FRA1:c-JUN:HDAC1 complex down-regulates filaggrin expression upon TNFα and IFNγ stimulation in keratinocytes.

Filaggrin (FLG), an essential structural protein for skin barrier function, is down-regulated under chronic inflammatory conditions, leading to disruption of the skin barrier. However, the detailed molecular mechanisms of how FLG changes in the context of chronic inflammation are poorly understood. Here, we identified the molecular mechanisms by which inflammatory cytokines inhibit FLG expression in the skin. We found that the AP1 response element within the -343/+25 of the FLG promoter was necessary for TNFα + IFNγ-induced down-regulation of FLG promoter activity. Using DNA affinity precipitation assay, we observed that AP1 subunit composition binding to the FLG promoter was altered from c-FOS:c-JUN (at the early time) to FRA1:c-JUN (at the late time) in response to TNFα + IFNγ stimulation. Knockdown of FRA1 or c-JUN abrogated TNFα + IFNγ-induced FLG suppression. Histone deacetylase (HDAC) 1 interacted with FRA1:c-JUN under TNFα + IFNγ stimulation. Knockdown of HDAC1 abrogated the inhibitory effect of TNFα + IFNγ on FLG expression. The altered expression of FLG, FRA1, c-JUN, and HDAC1 was confirmed in mouse models of 2,4-dinitrochlorobenzene-induced atopic dermatitis and imiquimod-induced psoriasis. Thus, the current study demonstrates that TNFα + IFNγ stimulation suppresses FLG expression by promoting the FRA1:c-JUN:HDAC1 complex. This study provides insight into future therapeutic strategies targeting the FRA1:c-JUN:HDAC1 complex to restore impaired FLG expression in chronic skin inflammation.

Influence of FLG loss-of-function mutations in host-microbe interactions during atopic skin inflammation.

BACKGROUND: Loss-of-function mutations in the filaggrin (FLG) gene directly alter skin barrier function and critically influence atopic inflammation. While skin barrier dysfunction, Th2-associated inflammation and bacterial dysbiosis are well-known characteristics of atopic dermatitis (AD), the mechanisms interconnecting genotype, transcriptome and microbiome remain largely elusive. OBJECTIVE: In-depth analysis of FLG genotype-associated skin gene expression alterations and host-microbe interactions in AD. METHODS: Multi-omics characterization of a cohort of AD patients carrying heterozygous loss-of-function mutations in the FLG gene (ADMut) (n = 15), along with matched wild-type (ADWt) patients and healthy controls. Detailed clinical characterization, microarray gene expression and 16 S rRNA-based microbial marker gene data were generated and analyzed. RESULTS: In the context of filaggrin dysfunction, the transcriptome was characterized by dysregulation of barrier function and water homeostasis, while the lesional skin of ADWt demonstrated the specific upregulation of pro-inflammatory cytokines and T-cell proliferation. S. aureus dominated the microbiome in both patient groups, however, shifting microbial communities could be observed when comparing healthy with non-lesional ADWt or ADMut skin, offering the opportunity to identify microbe-associated transcriptomic signatures. Moreover, an AD core signature with 28 genes, including CCL13, CCL18, BTC, SCIN, RAB31 and PCLO was identified. CONCLUSIONS: Our integrative approach provides molecular insights for the concept that FLG loss-of-function mutations are a genetic shortcut to atopic inflammation and unravels the complex interplay between genotype, transcriptome and microbiome in the human holobiont.

The Discovery and Function of Filaggrin.

Keratohyalin granules were discovered in the mid-19th century in cells that terminally differentiate to form the outer, cornified layer of the epidermis. The first indications of the composition of these structures emerged in the 1960s from a histochemical stain for histidine, followed by radioautographic evidence of a high incidence of histidine incorporation into newly synthesized proteins in cells containing the granules. Research during the next three decades revealed the structure and function of a major protein in these granules, which was initially called the 'histidine-rich protein'. Steinert and Dale named the protein 'filaggrin' in 1981 because of its ability to aggregate keratin intermediate filaments. The human gene for the precursor, 'profilaggrin,' was reported in 1991 to encode 10, 11 or 12 nearly identical repeats. Remarkably, the mouse and rat genes encode up to 20 repeats. The lifetime of filaggrin is the time required for keratinocytes in the granular layer to move into the inner cornified layer. During this transition, filaggrin facilitates the collapse of corneocytes into 'building blocks' that become an impermeable surface barrier. The subsequent degradation of filaggrin is as remarkable as its synthesis, and the end-products aid in maintaining moisture in the cornified layer. It was apparent that ichthyosis vulgaris and atopic dermatitis were associated with the absence of this protein. McLean's team in 2006 identified the cause of these diseases by discovering loss-of-function mutations in the profilaggrin gene, which led to dysfunction of the surface barrier. This story illustrates the complexity in maintaining a healthy, functional epidermis.

Polymorphisms of the filaggrin gene are associated with atopic dermatitis in the Caucasian population of Central Russia.

Association of the filaggrin (FLG) gene with atopic dermatitis (AD) in Caucasians from Central Russia was studied in the sample of 700 patients and 612 controls. In total ten SNPs of the gene (rs61816761, rs12130219, rs77199844, rs558269137, rs4363385, rs12144049, rs471144, rs6661961, rs10888499, rs3126085), their haplotypes and interlocus interactions were analyzed using logistic regression. The functional effects of the AD risk candidate loci and their proxies (136 SNPs) were evaluated by in silico analysis. All analyzed SNPs were associated with AD: two SNPs (rs3126085 and rs12144049) manifested the independent association, nine SNPs were associated within 30 haplotypes, and seven SNPs showed interlocus interaction effects within ten most significant epistatic models. Alleles A rs3126085 and C rs12144049 were associated with a higher risk of AD according to the allelic (ORs being 1.75, pperm = 0.002 and 1.45, pperm = 0.011 respectively), additive (ORs being 1.69, pperm = 0.004 and 1.47, pperm = 0.011 respectively) and dominant (ORs being 1.79, pperm = 0.004 and 1.63, pperm = 0.005 respectively) genetic models. Three haplotypes, GT[rs3126085-rs12144049] (OR = 0.60), GGT[rs61816761-rs3126085-rs12144049] (OR = 0.59), and AWGGT[rs12130219-rs558269137-rs61816761-rs3126085-rs12144049] (OR = 0.63) demonstrated the protective effect (pperm = 0.001). The in silico analysis suggested that the AD risk variants and their proxies apparently produce various effects on 38 genes in various tissue/organs (including 20 genes in the skin). The biological process enrichment analyses suggest that the target AD candidate genes influence the formation of the cornified envelope, keratinization and cornification, and more than twenty other pathways related to skin development, programmed cell death, and regulation of water loss via skin.

Novel role for caspase recruitment domain family member 14 and its genetic variant rs11652075 in skin filaggrin homeostasis.

BACKGROUND: Low epidermal filaggrin (FLG) is a risk factor for atopic dermatitis (AD) and allergic comorbidity. FLG mutations do not fully explain the variation in epidermal FLG levels, highlighting that other genetic loci may also regulate FLG expression. OBJECTIVE: We sought to identify genetic loci that regulate FLG expression and elucidate their functional and mechanistic consequences. METHODS: A genome-wide association study of quantified skin FLG expression in lesional and baseline non(never)-lesional skin of children with AD in the Mechanisms of Progression of Atopic Dermatitis to Asthma in Children cohort was conducted. Clustered regularly interspaced short palindromic repeat approaches were used to create isogenic human keratinocytes differing only at the identified variant rs11652075, and caspase recruitment domain family member 14 (CARD14)-deficient keratinocytes for subsequent mechanistic studies. RESULTS: The genome-wide association study identified the CARD14 rs11652075 variant to be associated with FLG expression in non(never)-lesional skin of children with AD. Rs11652075 is a CARD14 expression quantitative trait locus in human skin and primary human keratinocytes. The T variant destroys a functional cytosine-phosphate-guanine site, resulting in reduced cytosine-phosphate-guanine methylation at this site (but not neighboring sites) in TT and CT compared with CC primary human keratinocytes and Mechanisms of Progression of Atopic Dermatitis to Asthma in Children children's skin samples, and rs11652075 increases CARD14 expression in an allele-specific fashion. Furthermore, studies in clustered regularly interspaced short palindromic repeat-generated CC and TT isogenic keratinocytes, as well as CARD14-haplosufficient and deficient keratinocytes, reveal that IL-17A regulates FLG expression via CARD14, and that the underlying mechanisms are dependent on the rs11652075 genotype. CONCLUSIONS: Our study identifies CARD14 as a novel regulator of FLG expression in the skin of children with AD. Furthermore, CARD14 regulates skin FLG homeostasis in an rs11652075-dependent fashion.

Nuclear IL-33 Plays an Important Role in IL-31‒Mediated Downregulation of FLG, Keratin 1, and Keratin 10 by Regulating Signal Transducer and Activator of Transcription 3 Activation in Human Keratinocytes.

IL-33, a chromatin-associated multifunctional cytokine, is implicated in the pathogenesis of atopic dermatitis (AD), an inflammatory skin disorder characterized by skin barrier dysfunction. IL-33 accumulates in the nuclei of epidermal keratinocytes (KCs) in AD lesions. However, it is unclear whether nuclear IL-33 directly contributes to the pathogenesis of AD. IL-31, a pruritogenic cytokine primarily produced by T helper type 2 cells, is elevated in AD lesions and promotes AD development by suppressing KC differentiation and inducing itching. In this study, we investigated the involvement of nuclear IL-33 in IL-31‒mediated suppression of KC differentiation. In monolayer cultures and living skin equivalent, IL-31 increased the expression of full-length IL-33 and the phosphorylation of signal transducer and activator of transcription 3 (STAT3) in the nuclei of human KCs, which in turn downregulated the expression of differentiation markers. We found that IL-31 and IL-4/IL-13 use very similar mechanisms to inhibit KC differentiation: nuclear IL-33 combines with phosphorylated STAT3 and functions as a STAT3 transcription cofactor, promoting phosphorylated STAT3 binding to the FLG promoter to inhibit its transcription; moreover, the nuclear IL-33/phosphorylated STAT3 complex drives the downregulation of keratin 1 and keratin 10 by reducing the availability of the transcription factor RunX1. Therefore, nuclear IL-33 plays an important role in IL-31‒mediated differentiation suppression by regulating STAT3 activation in human KCs.

IL-22 Downregulates Peptidylarginine Deiminase-1 in Human Keratinocytes: Adding Another Piece to the IL-22 Puzzle in Epidermal Barrier Formation.

Increased presence of IL-22+ cells in the skin is a characteristic finding in skin barrier defects, such as psoriasis and atopic dermatitis. However, mechanistic insight into effects of IL-22 on epidermal functioning is yet to be elucidated. One crucial step during epidermal differentiation is deimination or citrullination. Here, we show reduced levels of peptidylarginine deiminase 1, an enzyme that converts peptidylarginine into citrulline in lesional psoriatic skin. IL-22 signaling through the IL-22 receptor complex was found to suppress expression of peptidylarginine deiminase 1 in epidermal keratinocytes. Subsequently, total peptidylarginine deiminase activity and extent of protein deimination in keratinocytes treated with IL-22 were reduced together with a significant decrease in deimination of keratin 1 and FLG, both important for epidermal differentiation. Vitamin D and acitretin partly restored the peptidylarginine deiminase 1 defect caused by IL-22. Collectively, we show that IL-22 downregulates deimination, thus identifying a potential target for treatment of skin barrier defects.

Topical Application of Galgeunhwanggeumhwangryeon-Tang Recovers Skin-Lipid Barrier and Ameliorates Inflammation via Filaggrin-Thymic Stromal Lymphopoietin-Interleukin 4 Pathway.

Background and objectives: The purpose of this study was to confirm the effect of Galgeunhwanggeumhwangryeon-tang (GGRT) on the skin barrier integrity and inflammation in an atopic dermatitis-like animal model. Materials and Methods: The model was established using lipid barrier elimination (LBE) in BALB/c mice. Ceramide 3B, a control drug, and GGRT were applied to the skin of LBE mice. Gross observation and histological examination were combined with measurement of skin score, trans-epidermal water loss, and pH. The expression of filaggrin, kallikrein-related peptidase 7 (KLK7), protease-activated receptor-2 (PAR-2), thymic stromal lymphopoietin (TSLP), and interleukin 4 (IL-4) was examined. Results: The effect of GGRT on atopic dermatitis was estimated in silico using two individual gene sets of human atopic dermatitis. In animal experiments, GGRT treatment reduced atopic dermatitis-like symptoms, as confirmed via gross and histological observations, skin score, pH change, and trans-epidermal water loss. The expression level of filaggrin increased in the skin of GGRT-treated mice compared to that in the LBE group. The expression levels of KLK7, PAR2, TSLP, and IL-4 were decreased in GGRT-treated mice skin compared to those in LBE mice. Conclusions: We demonstrated that GGRT restored the skin barrier and reduced inflammatory reactions in a murine model of atopic dermatitis.

Pathogenic Mechanism of Der p 38 as a Novel Allergen Homologous to RipA and RipB Proteins in Atopic Dermatitis.

Atopic dermatitis (AD) is a chronic relapsing pruritic disease encompassing skin inflammation and barrier dysfunction. House dust mites are key allergens that augment the development of atopic dermatitis. We aimed to investigate the pathogenic mechanism of AD due to Der p 38, recently identified by us. The frequency of IgE reactivity to Der p 38 in AD subjects was 52.6% (10/19) in the skin prick test and 57.9% (11/19) in the dot blot assay. In human keratinocyte HaCaT cells, Der p 38 triggered the impairment of filaggrin expression and induced pro-inflammatory cytokines such as IL-6, IL-8 and MCP-1 through TLR4, PI3K, AKT, c-Jun N-terminal kinase (JNK) and NF-κB pathway. Supernatants from Der p 38-treated cells blocked filaggrin expression and neutrophil apoptosis. The anti-apoptotic effect of the Der p 38-released molecules on neutrophils was accomplished by inhibition of the caspase 9/3 pathway, and by increased MCL-1 expression and BCL-2/BAX expression ratio. In C57BL/6 wild type (WT) mice, Der p 38 induced a dose-dependent increase of AD-like skin lesions, with enhanced expressions of total and Der p 38-specific IgE. Der p 38 also diminished the expressions of skin barrier proteins and induced JNK activation. However, the AD-like features following cutaneous Der p 38 exposure were observed to be reduced in the TLR4 knockout (KO) group, as compared to the WT group. Skin infiltration of neutrophils, eosinophils and mast cells was increased in the WT mice, but was not portrayed in the TLR4 KO mice. These findings indicate that Der p 38 is a novel mite allergen that triggers AD by lowering skin barrier proteins and increasing inflammatory cells. Results of this study have thereby paved the way to unveil the pathogenic mechanisms of AD.

K15 promoter-driven enforced expression of NKIRAS exhibits tumor suppressive activity against the development of DMBA/TPA-induced skin tumors.

NKIRAS1 and NKIRAS2 (also called as κB-Ras) were identified as members of the atypical RAS family that suppress the transcription factor NF-κB. However, their function in carcinogenesis is still controversial. To clarify how NKIRAS acts on cellular transformation, we generated transgenic mice in which NKIRAS2 was forcibly expressed using a cytokeratin 15 (K15) promoter, which is mainly activated in follicle bulge cells. The ectopic expression of NKIRAS2 was mainly detected in follicle bulges of transgenic mice with NKIRAS2 but not in wild type mice. K15 promoter-driven expression of NKIRAS2 failed to affect the development of epidermis, which was evaluated using the expression of K10, K14, K15 and filaggrin. However, K15 promoter-driven expression of NKIRAS2 effectively suppressed the development of skin tumors induced by treatment with 7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoylphorbol 13-acetate (TPA). This observation suggested that NKIRAS seemed to function as a tumor suppressor in follicle bulges. However, in the case of oncogenic HRAS-driven cellular transformation of murine fibroblasts, knockdown of NKIRAS2 expression drastically suppressed HRAS-mutant-provoked cellular transformation, suggesting that NKIRAS2 was required for the cellular transformation of murine fibroblasts. Furthermore, moderate enforced expression of NKIRAS2 augmented oncogenic HRAS-provoked cellular transformation, whereas an excess NKIRAS2 expression converted its functional role into a tumor suppressive phenotype, suggesting that NKIRAS seemed to exhibit a biphasic bell-shaped enhancing effect on HRAS-mutant-provoked oncogenic activity. Taken together, the functional role of NKIRAS in carcinogenesis is most likely determined by not only cellular context but also its expression level.

S100A7 Co-localization and Up-regulation of Filaggrin in Human Sinonasal Epithelial Cells.

OBJECTIVE: Filaggrin (FLG) is a protein expressed in the epidermis and involved in the maintenance of the epidermal barrier. However, the expression and localization of FLG in the upper airway remain controversial. The present study aimed to determine the significance of FLG and the effect of S100A7 on FLG expression in the upper respiratory mucosa. METHODS: Human nasal epithelial cells (HNECs) were cultured and examined for FLG expression and S100A7 effects by real-time polymerase chain reaction and Western blotting. The localization and distribution of FLG were assessed using sinonasal mucosa. RESULTS: A significant expression of FLG was detected at the mRNA and protein levels in HNECs. A moderate FLG immunoreactivity was observed in the epithelial cells, but no staining was seen in epithelial goblet cells. S100A7 increased the FLG mRNA level in HNECs in a dose-dependent manner and also up-regulated the FLG protein in a dose-dependent manner. CONCLUSION: This study significantly contributes to a better understanding of the role of FLG in the pathogenesis of airway inflammation from the viewpoint of the epithelial barrier function. FLG-related events in response to S100A7 protein may represent novel therapeutic targets for the treatment of upper airway inflammation.

Sensitization to peanut, egg or pets is associated with skin barrier dysfunction in children with atopic dermatitis.

BACKGROUND: Children with atopic dermatitis (AD) are often sensitized to food and aeroallergens, but sensitization patterns have not been analysed with biologic measures of disease pathogenicity. OBJECTIVE: We sought to define allergen sensitization grouping(s) using unbiased machine learning and determine their associations with skin filaggrin (FLG) and transepidermal water loss (TEWL) (assesses skin barrier integrity), S100A8 and S100A9 expression (assesses skin inflammation) and AD severity. METHODS: We studied 400 children with AD in the Mechanisms of Progression from Atopic Dermatitis to Asthma in Children (MPAACH) cohort to identify groupings of food and aeroallergen sensitizations. MPAACH is a paediatric AD cohort, aged 1-2, recruited through hospital/community settings between 2016 and 2018. We analysed these groupings' associations with AD biomarkers: skin FLG, S100A8 and S100A9 expression, total IgE, TEWL and AD severity. RESULTS: An unbiased machine learning approach revealed five allergen clusters. The most common cluster (N = 131), SPTPEP, had sensitization to peanut, egg and/or pets. Three low prevalence clusters, which included children with allergen sensitization other than peanut, egg or pets, were combined into SPTOther . SPTNEG included children with no sensitization(s). SPTPEP children had higher median non-lesional TEWL (16.9 g/m2 /h) and IgE (90 kU/L) compared with SPTOTHER (8.8 g/m2 /h and 24 kU/L; p = .01 and p < .001) and SPTNEG (9 g/m2 /h and 26 kU/L; p = .003 and p < .001). SPTPEP children had lower median lesional (0.70) and non-lesional (1.09) FLG expression compared with SPTOTHER (lesional: 0.9; p = .047, non-lesional: 1.78; p = .01) and SPTNEG (lesional: 1.47; p < .001, non-lesional: 2.21; p < .001). There were no differences among groupings in S100A8 or S100A9 expression. CONCLUSIONS AND CLINICAL RELEVANCE: In this largely clinic-based cohort of young children with AD, allergic sensitization to peanut, egg, cat or dog was associated with more severe disease and skin barrier function but not markers of cutaneous inflammation. These data need replicating in a population-based cohort but may have important implications for understanding the interaction between AD and allergic sensitization.

Particulate matter causes skin barrier dysfunction.

The molecular mechanisms that underlie the detrimental effects of particulate matter (PM) on skin barrier function are poorly understood. In this study, the effects of PM2.5 on filaggrin (FLG) and skin barrier function were investigated in vitro and in vivo. The levels of FLG degradation products, including pyrrolidone carboxylic acid, urocanic acid (UCA), and cis/trans-UCA, were significantly decreased in skin tape stripping samples of study subjects when they moved from Denver, an area with low PM2.5, to Seoul, an area with high PM2.5 count. Experimentally, PM2.5 collected in Seoul inhibited FLG, loricrin, keratin-1, desmocollin-1, and corneodesmosin but did not modulate involucrin or claudin-1 in keratinocyte cultures. Moreover, FLG protein expression was inhibited in human skin equivalents and murine skin treated with PM2.5. We demonstrate that this process was mediated by PM2.5-induced TNF-α and was aryl hydrocarbon receptor dependent. PM2.5 exposure compromised skin barrier function, resulting in increased transepidermal water loss, and enhanced the penetration of FITC-dextran in organotypic and mouse skin. PM2.5-induced TNF-α caused FLG deficiency in the skin and subsequently induced skin barrier dysfunction. Compromised skin barrier due to PM2.5 exposure may contribute to the development and the exacerbation of allergic diseases such as atopic dermatitis.

Deacetylasperulosidic Acid Ameliorates Pruritus, Immune Imbalance, and Skin Barrier Dysfunction in 2,4-Dinitrochlorobenzene-Induced Atopic Dermatitis NC/Nga Mice.

The prevalence of atopic dermatitis (AD), a disease characterized by severe pruritus, immune imbalance, and skin barrier dysfunction, is rapidly increasing worldwide. Deacetylasperulosidic acid (DAA) has anti-atopic activity in the three main cell types associated with AD: keratinocytes, mast cells, and eosinophils. Our study investigated the anti-atopic activity of DAA in 2,4-dinitrochlorobenzene-induced NC/Nga mice. DAA alleviated the symptoms of AD, including infiltration of inflammatory cells (mast cells and eosinophils), epidermal thickness, ear thickness, and scratching behavior. Furthermore, DAA reduced serum IgE, histamine, and IgG1/IgG2a ratio and modulated the levels of AD-related cytokines and chemokines, namely interleukin (IL)-1ß, IL-4, IL-6, IL-9, IL-10, IL-12, tumor necrosis factor-α, interferon-γ, thymic stromal lymphopoietin, thymus and activation-regulated chemokine, macrophage-derived chemokine, and regulated on activation the normal T cell expressed and secreted in the serum. DAA restored immune balance by regulating gene expression and secretion of Th1-, Th2-, Th9-, Th17-, and Th22-mediated inflammatory factors in the dorsal skin and splenocytes and restored skin barrier function by increasing the expression of the pro-filaggrin gene and barrier-related proteins filaggrin, involucrin, and loricrin. These results suggest DAA as a potential therapeutic agent that can alleviate the symptoms of AD by reducing pruritus, modulating immune imbalance, and restoring skin barrier function.

Immunohistochemical expression of filaggrin is decreased in proton pump inhibitor non-responders compared with proton pump inhibitor responders of eosinophilic esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is an allergic gastrointestinal disease that features eosinophilic infiltration of esophageal mucosa, but the role of barrier dysfunction of the epithelium in its pathogenesis remains to be elucidated. Clinically, EoE is divided into proton pump inhibitor-non-responders (PPI-NR) and PPI-responders (PPI-R). Our main aims were to investigate the differences of expression of epidermal differential complex (EDC) proteins and desmoglein that are considered to play important roles in formation of the epidermal skin barrier between these two conditions and to seek the usefulness of the differences in pathological diagnosis. Conventional histopathological findings and allergic background were also compared. METHODS: Twenty-nine PPI-NR and 44 PPI-R were recruited, and 35 reflux esophagitis patients were also enrolled. After clinical information and histopathological findings were reviewed, immunohistochemical expression of EDC proteins (filaggrin, loricrin, and involucrin) and desmoglein in all three groups were examined and semi-quantitatively scored. RESULTS: Regarding allergic conditions, the prevalence of asthma was significantly higher in PPI-NR than in PPI-R. Other allergic conditions showed no differences. Histopathological findings did not exhibit the statistical difference between PPI-NR and PPI-R. However, immunostaining score of filaggrin in PPI-NR was significantly lower than in PPI-R, although the expressions of involucrin, loricrin and desmoglein demonstrated no differences. CONCLUSIONS: The results suggest a role of reduced filaggrin expression in the difference of effectiveness of PPI treatment between PPI-NR and PPI-R. Moreover, immunohistochemical determination of filaggrin expression in EoE patients could be informative in the clinical decision of how to treat the patients.

The vitamin A ester retinyl propionate has a unique metabolic profile and higher retinoid-related bioactivity over retinol and retinyl palmitate in human skin models.

Human skin is exposed daily to environmental stressors, which cause acute damage and inflammation. Over time, this leads to morphological and visual appearance changes associated with premature ageing. Topical vitamin A derivatives such as retinol (ROL), retinyl palmitate (RPalm) and retinyl propionate (RP) have been used to reverse these changes and improve the appearance of skin. This study investigated a stoichiometric comparison of these retinoids using in vitro and ex vivo skin models. Skin biopsies were treated topically to compare skin penetration and metabolism. Treated keratinocytes were evaluated for transcriptomics profiling and hyaluronic acid (HA) synthesis and treated 3D epidermal skin equivalents were stained for epidermal thickness, Ki67 and filaggrin. A retinoic acid receptor-alpha (RARα) reporter cell line was used to compare retinoid activation levels. Results from ex vivo skin found that RP and ROL have higher penetration levels compared with RPalm. RP is metabolized primarily into ROL in the viable epidermis and dermis whereas ROL is esterified into RPalm and metabolized into the inactive retinoid 14-hydroxy-4,14-retro-retinol (14-HRR). RP treatment yielded higher RARα activation and HA synthesis levels than ROL whereas RPalm had a null effect. In keratinocytes, RP and ROL stimulated similar gene expression patterns and pathway theme profiles. In conclusion, RP and ROL show a similar response directionality whereas RPalm response was inconsistent. Additionally, RP has a consistently higher magnitude of response compared with ROL or RPalm.

Topical application with conjugated linoleic acid ameliorates 2, 4-dinitrofluorobenzene-induced atopic dermatitis-like lesions in BALB/c mice.

Atopic dermatitis (AD) is a multifactorial chronic inflammatory skin disease characterized by skin barrier dysfunction, eczematous lesions, pruritus, and abnormal immune responses. In this study, we assessed the therapeutic effect of topical applied conjugated linoleic acid (CLA) on a murine AD model that was developed by repetitive applications of 2, 4-dinitrofluorobenzene (DNFB). 2% or 5% CLA could markedly ameliorate AD-like skin lesions, scratching behaviour and skin inflammation as evidenced by the reduced inflammatory blood cells, IgE and Th2-related cytokine levels, and the infiltration of mast cells and inflammatory cells to the dermal tissues. Moreover, topical application with CLA modulated skin barrier repair including maintaining a balanced skin pH and increasing skin hydration, partially mediated by upregulating skin barrier-related protein, filaggrin (FLG). In addition, topical CLA significantly dose-dependently inhibited pro-inflammatory cytokines including interleukin (IL)-6, IL-1ß, tumour necrosis factor (TNF)-α and pro-inflammatory enzyme expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in inflamed mice skin. Its anti-inflammatory effect was associated with the inhibition of DNFB-stimulated IκBα and NF-κB p65 phosphorylation in mouse skin. Taken together, our results suggest that locally applied CLA exerts potentially protective effects against AD lesional skin at least in part, due to regulation of skin barrier function and inflammatory response.