Skin Barrier Proteins Research Articles

Upregulation of FLG, LOR, and IVL Expression by Rhodiola crenulata Root Extract via Aryl Hydrocarbon Receptor: Differential Involvement of OVOL1.

Rhodiola species are antioxidative, salubrious plants that are known to inhibit oxidative stress induced by ultraviolet and γ-radiation in epidermal keratinocytes. As certain phytochemicals activate aryl hydrocarbon receptors (AHR) or OVO-like 1 (OVOL1) to upregulate the expression of epidermal barrier proteins such as filaggrin (FLG), loricrin (LOR), and involucrin (IVL), we investigated such regulation by Rhodiola crenulata root extract (RCE). We demonstrated that RCE induced FLG and LOR upregulation in an AHR-OVOL1-dependent fashion. However, RCE-mediated IVL upregulation was AHR-dependent but OVOL1-independent. Coordinated upregulation of skin barrier proteins by RCE via AHR may be beneficial in the management of barrier-disrupted inflammatory skin diseases such as atopic dermatitis.

Protective role of 6-formylindolo[3,2-b]carbazole (FICZ), an endogenous ligand for arylhydrocarbon receptor, in chronic mite-induced dermatitis

BackgroundChronic eczema such as atopic dermatitis imposes significant socio-econo-psychologic burdens on the affected individuals. In addition to conventional topical treatments, phototherapy is recommended for patients with extensive lesions. Although immunosuppression is believed to explain its primary effectiveness, the underlying mechanisms of phototherapy remain unsolved. Ultraviolet irradiation generates various tryptophan photoproducts including 6-formylindolo[3,2-b]-carbazole (FICZ). FICZ is known to be a potent endogenous agonist for aryl hydrocarbon receptor (AHR); however, the biological role of FICZ in chronic eczema is unknown. ObjectiveTo investigate the effect of FICZ on chronic eczema such as atopic dermatitis. MethodsWe stimulated HaCaT cells and normal human epidermal keratinocytes (NHEKs) with or without FICZ and then performed quantitative reverse transcriptase polymerase chain reaction, immunofluorescence, and siRNA treatment. We used the atopic dermatitis-like NC/Nga murine model and treated the mice for 2 weeks with either Vaseline® as a control, FICZ ointment, or betamethasone 17-valerate ointment. The dermatitis score, transepidermal water loss, histology, and expression of skin barrier genes and proteins were evaluated. ResultsFICZ significantly upregulated the gene expression of filaggrin in both HaCaT cells and NHEKs in an AHR-dependent manner, but did not affect the gene expression of other barrier-related proteins. In addition, FICZ improved the atopic dermatitis-like skin inflammation, clinical scores, and transepidermal water loss in NC/Nga mice compared with those of control mice. On histology, FICZ significantly reduced the epidermal and dermal thickness as well as the number of mast cells. Topical FICZ also significantly reduced the gene expression of Il22. ConclusionThese findings highlight the beneficial role of FICZ-AHR and provide a new strategic basis for developing new drugs for chronic eczema.

The Development of Diagnosis for Atopic Dermatitis by Evaluating the Expression of Skin Barrier Proteins Using a Non-Invasive Method

The Development of Diagnosis for Atopic Dermatitis by Evaluating the Expression of Skin Barrier Proteins Using a Non-Invasive Method

P38/ERK MAPK signaling pathways are involved in the regulation of filaggrin and involucrin by IL‑17.

Atopic dermatitis (AD) is characterized by a defective skin barrier, which increases the penetration of allergens and pathogens through the skin. The role of interleukin (IL)-17, a pro-inflammatory cytokine, in the pathogenesis of AD remains to be elucidated. The present study aimed to examine the effects of IL-17 on skin barrier proteins in the HaCaT cell line. The expression levels of filaggrin (FLG) and involucrin (IVL) were evaluated by reverse transcription-quantitative polymerase chain reaction and western blot analyses of the HaCaT cells following IL-17 simulation. The role of IL-17 was further examined by using small molecule inhibitors of extracellular signal-regulated kinase (ERK) and P38. Treatment of the HaCaT cells with IL-17 resulted in reduced expression levels of FLG and IVL at the mRNA and protein levels. In addition, the gene expression levels of FLG and IVL were significantly reduced in the HaCaT cells by IL-4. Treatment with the mitogen-activated protein kinase (MAPK) inhibitors, SB203580 and PD98059, significantly inhibited the effects of IL-17 on the gene and protein expression levels of FLG and IVL. Finally, the protein levels of phosphorylated ERK and P38 were significantly increased following IL-17 stimulation. Taken together, the results revealed that IL-17 reduced the expression of FLG and IVL in HaCaT cells, and this effect involved the P38/ERK MAPK signaling pathways.

Feeding filaggrin: effects of l-histidine supplementation in atopic dermatitis.

Atopic dermatitis (AD), also known as eczema, is one of the most common chronic skin conditions worldwide, affecting up to 16% of children and 10% of adults. It is incurable and has significant psychosocial and economic impacts on the affected individuals. AD etiology has been linked to deficiencies in the skin barrier protein, filaggrin. In mammalian skin, l-histidine is rapidly incorporated into filaggrin. Subsequent filaggrin proteolysis releases l-histidine as an important natural moisturizing factor (NMF). In vitro studies were conducted to investigate the influence of l-histidine on filaggrin processing and barrier function in human skin-equivalent models. Our further aim was to examine the effects of daily oral l-histidine supplementation on disease severity in adult AD patients. We conducted a randomized, double-blind, placebo-controlled, crossover, nutritional supplementation pilot study to explore the effects of oral l-histidine in adult AD patients (n=24). In vitro studies demonstrated that l-histidine significantly increased both filaggrin formation and skin barrier function (P<0.01, respectively). Data from the clinical study indicated that once daily oral l-histidine significantly reduced (P<0.003) AD disease severity by 34% (physician assessment using the SCORingAD tool) and 39% (patient self-assessment using the Patient Oriented Eczema Measure tool) after 4 weeks of treatment. No improvement was noted with the placebo (P>0.32). The clinical effect of oral l-histidine in AD was similar to that of mid-potency topical corticosteroids and combined with its safety profile suggests that it may be a safe, nonsteroidal approach suitable for long-term use in skin conditions that are associated with filaggrin deficits such as AD.

Antioxidant Artemisia princeps Extract Enhances the Expression of Filaggrin and Loricrin via the AHR/OVOL1 Pathway.

The Japanese mugwort, Artemisia princeps (yomogi in Japanese), has anti-inflammatory and antioxidant effects. Skin care products containing Artemisia princeps extract (APE) are known to improve dry skin symptoms in atopic dermatitis. Atopic dry skin is associated with a marked reduction of skin barrier proteins, such as filaggrin (FLG) and loricrin (LOR). Recently, aryl hydrocarbon receptor (AHR), and its downstream transcription factor OVO-like 1 (OVOL1), have been shown to regulate the gene expression of FLG and LOR. The focus of this paper is to evaluate the effects of APE on the AHR/OVOL1/FLG or LOR pathway since they have remained unknown to this point. We first demonstrated that non-cytotoxic concentrations of APE significantly upregulated antioxidant enzymes, NAD(P)H dehydrogenase quinone 1 and heme oxygenase 1, in human keratinocytes. Even at these low concentrations, APE induced nuclear translocation of AHR and significantly upregulated CYP1A1 (a specific target gene for AHR activation), FLG, and LOR expression. AHR knockdown downregulated OVOL1 expression. The APE-induced upregulation of FLG and LOR was canceled in keratinocytes with AHR or OVOL1 knockdown. In conclusion, antioxidant APE is a potent phytoextract that upregulates FLG and LOR expression in an AHR/OVOL1-dependent manner and this may underpin the barrier-repairing effects of APE in treating atopic dry skin.

135 Comparative genomics of skin barrier proteins reveals environment-dependent dispensability of the suprabasal epidermal keratins K1, K2, and K10

The epidermis has been shaped by evolutionary adaptations to the environment. In humans, the most abundant proteins of the suprabasal epidermis are keratins K1, K2 and K10 which together form the cytoskeleton at the skin barrier. Mutations in K1 and K10 cause epidermolytic hyperkeratosis. Here, we performed comparative genomics to determine whether mutations in keratin genes were tolerated under different environmental constraints. Genome sequences of a phenotypically diverse set of mammalian species were screened by bioinformatic methods. The basal epidermal keratins K5 and K14 as well as the hyperproliferation-associated keratins K6 and K17 are highly conserved among mammals. By contrast, the late differentiation-associated keratin K2 has been lost in at least 4 phylogenetic lineages, indicating dispensability of K2 for skin barrier formation. K1 and K10 have been conserved in terrestrial mammals, but both have been inactivated in fully aquatic mammals of the phylogenetic clade comprising dolphins and whales. The results of this study identify specific components of the epidermal cytoskeleton that are associated with the skin barrier function in a terrestrial environment.

The "Alarmins" HMBG1 and IL-33 Downregulate Structural Skin Barrier Proteins and Impair Epidermal Growth.

The epidermal-derived "alarmins" high-mobility group box 1 (HMGB1) protein and interleukin-33 (IL-33) are upregulated in patients with atopic dermatitis. How-ever, their capacity as pro-inflammatory cytokines and their derived effects on skin barrier regulation are poorly elucidated. We investigated the impact of HMGB1 and IL-33 on gene transcription, protein expression and epidermal differentiation across 3 distinct keratinocyte in vitro models. Primary keratinocytes from healthy donors were used in submerged monolayer cultures, 3D human epidermis equivalents and 3D human skin equivalents. All keratinocyte models underwent 96-h stimulation with HMGB1 (100 μM) or IL-33 (100 ng/ml) using IL-4 (50 ng/ml) as positive control of regulation and vehicle as negative control. We found that HMGB1 and IL-33 downregulated transcription of several genes from members of the epidermal differentiation complex, including filaggrin. Furthermore, HMGB1 downregulated the expression of the encoded proteins in the upper epidermis. Finally, IL-33 and HMGB1 ultimately led to impaired epidermal growth and maturation. In conclusion, HMGB1 and IL-33 could play a significant role in the atopic dermatitis pathophysiology due to negative regulation of structural proteins, stratum corneum formation and epidermal growth.

Profile of skin barrier proteins and cytokines in adults with atopic dermatitis.

Atopic dermatitis (AD), an inflammatory skin disorder with chronic course and characterized by intense pruritus, is a dermatosis of high prevalence of childhood. However, persistence of the disease in adolescents and adults may occur, and more studies regarding the interactions of the complex triggering factors, especially between the adaptive and innate immune alterations and skin barrier defects are needed. In this review the authors summarize the major novel findings of a dysfunctional skin barrier in AD, with emphasis on tight junction components, such as claudins and on proteins of the keratinocyte differentiation, such as filaggrin. This review also provides an update on the characterization of immune response in adults with atopic dermatitis. The adaptive immune dysfunction in AD, classically known as a Th2/Th1 model, has changed its profile, with recent reported cytokines such as interleukins 17, 22, and 31; as for the innate immune system scenario in AD, the characterization of skin microbiome opens new frontiers for the understanding of such a complex inflammatory disease.

Atopic dermatitis: A tale of two distinct pathomechanisms that make you itch.

Atopic dermatitis (AD) or eczema is the most common chronic inflammatory skin disease. It is a multifactorial disease with local and systemic immune changes. Current therapies focus on restoring the local skin barrier or inhibiting immune responses. In this issue of the European Journal of Immunology, Sehra etal. [Eur. J. Immunol. 2016. 46:2609-2613] describe a mouse model with T-cell-specific expression of constitutively active Stat6 in Flaky tail mice, which have mutations in the Flg and Tmem79 genes. The authors describe that it is the combination of changes in the skin barrier proteins filaggrin and Tmem79, together with Th2 cytokine signaling in the constitutively active Stat6 transgene, that drives the immune-pathomechanism in AD. These results are consistent with human studies where it is demonstrated that diminished filaggrin expression in skin is a predisposing factor for AD, but is neither required nor sufficient for disease indicating that additional factors are required for disease development. The current mouse model by Sehra etal. could be instrumental in evaluation new therapeutic strategies for AD.

Impact of systemic alitretinoin treatment on skin barrier gene and protein expression in patients with chronic hand eczema.

Chronic hand eczema (CHE) is a common inflammatory skin disease that affects approximately 10% of the population. Systemic alitretinoin has been shown to be effective in patients with CHE who are refractory to topical corticosteroids. To analyse the impact of alitretinoin on the skin barrier genes and protein expression in the skin lesions of patients with CHE. Fifteen patients with CHE were treated with 30 mg daily of alitretinoin for up to 27 weeks. Disease severity was assessed using a clinical score. Skin biopsies from all the patients were evaluated before and after therapy for the expression of Ki-67, various skin barrier genes and thymic stromal lymphopoietin (TSLP) by real-time quantitative polymerase chain reaction and immunohistochemistry. After alitretinoin application, an improvement in the clinical severity of CHE was observed in the majority of patients. Analysis of skin biopsies before treatment showed a significant increase in Ki-67-positive cells in the suprabasal layer and a dysregulated expression of various skin barrier genes, such as claudin 1, loricrin, filaggrin and cytokeratin 10, which were normalized after treatment. TSLP was significantly upregulated in patients with CHE and also normalized after alitretinoin treatment and negatively correlated with filaggrin. Our data indicate that the expression of barrier genes and proteins was normalized following treatment with alitretinoin in patients with CHE. The change in expression levels of these genes correlated with the clinical efficacy, suggesting that alitretinoin exhibits a disease-modifying activity. TSLP is upregulated in CHE and seems to counteract filaggrin expression in the skin.

Comprehensive Approach of Treatment of Atopic Dermatitis including Intensive Skin Hydration Therapy (Wet Wrap Therapy–WWT) – A Review.

Atopic dermatitis (AD), the most common skin disease in children, is the earliest presentation of allergic diseases among childhood. Although AD is commonly seen within the infantile age group, it can manifest its initial presentation in late childhood and in adulthood. The classic sites of the rash, the chronic relapsing nature of the disease along with itching establishes the diagnosis of AD. Despite the fact that ‘atopic’ denotes the relationship with IgE-mediated mechanism, atopy could be demonstrated in approximately 50% of AD cases. Common allergens found in association with infantile AD are food allergens (commonly cow’s milk and eggs) whereas inhalant allergens are found among AD cases of older ages. Recently, defects of skin barrier proteins which protect the transepidermal loss of water were found among skin of AD patients. The classic molecule that has been extensively studied was filaggrin. With the combination of defective skin barrier protein and altered immune mechanism, AD skin became dry, itchy and inflamed. Treatment of AD requires several modalities of approach including hydration of skin, use of moisturizers, use of anti-inflammatory agents, treatment of skin infections, and avoidance of allergens. This multilevel approach, although difficult to accomplish, can be adequately arranged even in busy clinics and thus leading to a higher level of success in the treatment of severe AD cases. Hydration of dry skin is an important step in the treatment of AD and in severe cases, a more complex hydration approach is needed. This can be achieved by comprehensive hydration therapy with prolonged wrapping with saline or water (wet wrap therapy–WWT). This article reviews important salient points for the pathogenesis of AD and also in-depth discussion on techniques of WWT. Effective therapy of AD can be achieved in a short period of time using WWT and can help sustain an optimal use of moisturizers and intermittent inflammatory agents.

Filaggrin inhibits generation of CD1a neolipid antigens by house dust mite-derived phospholipase.

Atopic dermatitis is a common pruritic skin disease in which barrier dysfunction and cutaneous inflammation contribute to pathogenesis. Mechanisms underlying the associated inflammation are not fully understood, and although Langerhans cells expressing the nonclassical major histocompatibility complex (MHC) family member CD1a are known to be enriched within lesions, their role in clinical disease pathogenesis has not been studied. We observed that house dust mite (HDM) allergen generates neolipid antigens presented by CD1a to T cells in the blood and skin lesions of affected individuals. HDM-responsive CD1a-reactive T cells increased in frequency after birth in individuals with atopic dermatitis and showed rapid effector function, consistent with antigen-driven maturation. In HDM-challenged human skin, we observed phospholipase A2 (PLA2) activity in vivo. CD1a-reactive T cell activation was dependent on HDM-derived PLA2, and such cells infiltrated the skin after allergen challenge. Moreover, we observed that the skin barrier protein filaggrin, insufficiency of which is associated with atopic skin disease, inhibited PLA2 activity and decreased CD1a-reactive PLA2-generated neolipid-specific T cell activity from skin and blood. The most widely used classification schemes of hypersensitivity suggest that nonpeptide stimulants of T cells act as haptens that modify peptides or proteins; however, our results show that HDM proteins may also generate neolipid antigens that directly activate T cells. These data define PLA2 inhibition as a function of filaggrin, supporting PLA2 inhibition as a therapeutic approach.

Inhibitory Effect of Valencene on the Development of Atopic Dermatitis-Like Skin Lesions in NC/Nga Mice.

Valencene (VAL) isolated from Cyperus rotundus possesses various biological effects such as antiallergic and antimelanogenesis activity. We investigated the effect of VAL on atopic dermatitis (AD) skin lesions and their molecular mechanisms. We topically applied VAL to 1-chloro-2,4-dinitrobenzene (DNCB) sensitized NC/Nga mice. Modified scoring atopic dermatitis index, scratching behavior, and histological/immunohistochemical staining were used to monitor disease severity. RT-PCR, western blotting, and enzyme-linked immunosorbent assay were used to determine the level of IgE, proinflammatory cytokines/chemokines production, and skin barrier proteins expression. Topical application of VAL significantly reduced AD-like symptoms and recovered decreased expression of filaggrin in DNCB-sensitized NC/Nga mice. The levels of serum IgE, IL-1β, IL-6, and IL-13 in skin/splenic tissue were reduced. In vitro studies using TNF-α and IFN-γ treated HaCaT cells revealed that VAL inhibited the exaggerated expression of Th2 chemokines including TARC/CCL17, MDC/CCL22, and proinflammatory chemokines such as CXCL8, GM-CSF, and I-CAM through blockade of the NF-κB pathway. In addition, expression of the skin barrier protein, involucrin, was also increased by VAL treatment. VAL inhibited the production and expression of proinflammatory cytokines IL-1β and IL-6 in LPS-stimulated RAW 264.7 cells. These results suggest that VAL may serve as a potential therapeutic option for AD.

Thymic stromal lymphopoietin downregulates filaggrin expression by signal transducer and activator of transcription 3 (STAT3) and extracellular signal-regulated kinase (ERK) phosphorylation in keratinocytes

Thymic stromal lymphopoietin downregulates filaggrin expression by signal transducer and activator of transcription 3 (STAT3) and extracellular signal-regulated kinase (ERK) phosphorylation in keratinocytes

Immunolocalization of loricrin in the maturing α-layer of normal and regenerating epidermis of the lizard Anolis carolinensis.

Numerous corneous proteins are produced during the differentiation of the complex lizard epidermis, comprising hard β-layers and softer α-layers. In the present ultrastructural and immunocytochemical study, we have localized a homolog of the mammalian skin barrier protein loricrin in the skin of the green anole lizard (Anolis carolinensis). We used an antibody specific to the carboxyterminus of loricrin 1, a gene of the epidermal differentiation complex (EDC) of A. carolinensis. Lizard loricrin is present in the maturing α-layer (lacunar cells) of normal scale epidermis and in the accumulating corneocytes of the wound epidermis (lacunar cells) of the regenerating epidermis. The protein appears as a component of the α-layer but not of the β-layer. Lizard loricrin is diffused in the cytoplasm of pre-corneous α-keratinocytes but eventually concentrates in the packing corneous material of the maturing corneocytes of the α-layer (lacunar) in normal epidermis or in the wound epidermis of regenerating epidermis. The protein likely contributes to the composition and pliability of the corneous material but is not specifically accumulated on the corneous cell envelope (marginal layer) that is scarcely differentiated in these cells. The study contributes to the knowledge on the distribution of specific corneous proteins that give rise to the different material properties of α-layers versus β-layers in lizard epidermis.

The connection between seasonal allergies, food allergies, and rhinosinusitis

Rhinosinusitis affects an estimated one in seven adults in the United States. Otolaryngologists are intimately involved in the care of patients with rhinosinusitis and other upper airway inflammatory conditions through procedures such as endoscopic sinus surgery and, therefore, would benefit from a deeper understanding of the associated comorbidities and their management. Recent evidence has suggested several connections between the underlying disease of rhinosinusitis, seasonal allergies, and food allergies. The authors of the present review seek to provide a focused analysis of the recent literature with respect to epidemiology, pathophysiology, and treatment options concerning these conditions. Evidence has connected the function of filaggrin, a skin barrier protein, with the pathogenesis of allergic rhinosinusitis and food allergy. Additionally, decreased levels of regulatory B cells and T cells are associated with and play a role in atopic disease. Overlapping treatment modalities between these conditions suggest similar conclusions. Future research into the role of the skin barrier, regulatory immune cell functioning, transforming growth factor-β, and other cytokine signaling, and treatment options such as omalizumab and azelastine is likely to have profound impact on clinicians' management of patients with these disorders and their comorbidities.

Insight from the Air–Skin Interface

The epidermis is a relatively hypoxic tissue, despite being continually exposed to air. The role of hypoxia in epidermal differentiation and skin barrier function is incompletely understood. In this issue, Wong et al. show that hypoxia-inducible factors are central to the processes of epidermal differentiation and barrier formation, in particular by promoting the expression of the key skin barrier protein filaggrin.

RETRACTED: Skin barrier dysfunction measured by transepidermal water loss at 2 days and 2 months predates and predicts atopic dermatitis at 1 year

BackgroundLoss-of-function mutations in the skin barrier protein filaggrin (FLG) are a major risk for atopic dermatitis (AD). The pathogenic sequence of disturbances in skin barrier function before or during the early development of AD is not fully understood. A more detailed understanding of these events is needed to develop a clearer picture of disease pathogenesis. A robust, noninvasive test to identify babies at high risk of AD would be important in planning early intervention and/or prevention studies.ObjectivesTo ascertain whether a noninvasive measurement of skin barrier function at day 2 after birth and at 2 months predicts the development of AD at 1 year. Furthermore, to determine whether increases in transepidermal water loss (TEWL) predate the development of clinical AD.MethodsA total of 1903 infants were enrolled in the Cork Babies After Scope: Evaluating the Longitudinal Impact Using Neurological and Nutritional Endpoints Birth Cohort study from July 2009 to October 2011. Measurements of TEWL were made at birth (day 2) and at 2 and 6 months. The presence of AD was ascertained at 6 and 12 months, and disease severity was assessed by using the SCORing Atopic Dermatitis clinical tool at 6 months and by using both the SCORing Atopic Dermatitis clinical tool and Nottingham Severity Score at 12 months. A total of 1300 infants were genotyped for FLG mutations.ResultsAt 6 months, 18.7% of the children had AD, and at 12 months, 15.53%. In a logistic regression model, day 2 upper quartile TEWL measurement was significantly predictive of AD at 12 months (area under the receiver operating characteristic curve, 0.81; P < .05). Lowest quartile day 2 TEWL was protective against AD at 12 months. An upper quartile 2 month TEWL was also strongly predictive of AD at 12 months (area under the receiver operating characteristic curve, 0.84; P < .05). At both ages, this effect was independent of parental atopy, FLG status, or report of an itchy flexural rash at 2 months. Associations were increased when parental atopy status or child FLG mutation status was added into the linear regression model.ConclusionsImpairment of skin barrier function at birth and at 2 months precedes clinical AD. In addition to providing important mechanistic insights into disease pathogenesis, these findings have implications for the optimal timing of interventions for the prevention of AD.

Immune Alterations in IgE and Non IgE-Associated Atopic Dermatitis

Atopic dermatitis is a complex disease in which a strong interaction between alterations of skin barrier and the adaptive immune system coexists. In the recent years, new findings have underlined the importance of skin proteins, especially filaggrin, which participate to the outmost layers of the skin. To strengthen this physical barrier, many factors are available, such as antimicrobial peptides, chemokines and cytokines produced by keratinocytes. Skin disruption can easily allow the allergen penetration and the local keratinocytes can promote the adaptive immune response toward a Th2 phenotype. On the other side, allergic Th2 cytokines may downregulate the production of skin barrier proteins, facilitating the penetration of allergens. Moreover, data on murine models show the absolute relevance of the systemic immune system to develop clinical skin reaction. Since the clinical aspect of patients with AD does not show different patterns whatever is the prevalent underlying mechanism, in clinical practice it is difficult to translate the different endotypes beside the IgE and non IgE associated forms. The aim of this review is to point out to the most recent knowledge in this field, which makes AD more difficult to frame in a unique clinical entity.