A molecular mechanism for IL-4 suppression of loricrin transcription in epidermal keratinocytes: implication for atopic dermatitis pathogenesis

Skin barrier defects play an important role in atopic dermatitis (AD) pathogenesis. Loricrin, an important barrier protein suppressed in human AD, is down-regulated by IL-4 in keratinocytes. However, the molecular mechanism is unknown. Since loricrin transcription requires p300/CBP, and Stat6 also recruits this common coactivator for its stimulated factors, we hypothesize that IL-4-activated Stat6 competes for the available endogenous p300/CBP, leading to loricrin transcription inhibition. First, we showed that loricrin is suppressed in the skin of IL-4 transgenic mice, an AD mouse model. In human keratinocytes, IL-4 down-regulation of loricrin is abrogated by a pan-Jak inhibitor, suggesting that the Jak-Stat pathway is involved. To further investigate the downstream molecular mechanism, we transfected HaCat cells with a loricrin promoter and then treated them with either IL-4 or vehicle. Not surprisingly, IL-4 greatly suppressed the promoter activity. Interestingly, this suppression was prevented when we knocked down Stat6, indicating that Stat6 participates in IL-4 regulation of loricrin. A Stat6-specific inhibitor confirmed the knockdown study. Finally, IL-4 suppression of loricrin was reversed with transfection of a CBP expression vector in a dose-dependent manner. Taken together, for the first time, we delineate a molecular mechanism for IL-4 down-regulation of loricin expression in human keratinocytes, which may play an important role in AD pathogenesis.

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IL-24 Negatively Regulates Keratinocyte Differentiation Induced by Tapinarof, an Aryl Hydrocarbon Receptor Modulator: Implication in the Treatment of Atopic Dermatitis

International Journal of Molecular Sciences ◽

10.3390/ijms21249412 ◽

2020 ◽

Vol 21 (24) ◽

pp. 9412

Author(s):

Yen Hai Vu ◽

Akiko Hashimoto-Hachiya ◽

Masaki Takemura ◽

Ayako Yumine ◽

Yasutaka Mitamura ◽

...

Keyword(s):

Atopic Dermatitis ◽

Aryl Hydrocarbon Receptor ◽

Skin Barrier ◽

Janus Kinase ◽

Keratinocyte Differentiation ◽

Epidermal Keratinocytes ◽

Dependent Manner ◽

Jak Inhibitors ◽

Barrier Dysfunction ◽

Aryl Hydrocarbon

Skin barrier dysfunction, including reduced filaggrin (FLG) and loricrin (LOR) expression, plays a critical role in atopic dermatitis (AD) development. Since aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor, mediates keratinocyte differentiation, it is a potential target for AD treatment. Recently, clinical studies have shown that tapinarof, an AHR modulator, attenuated the development of AD. To examine the molecular mechanism involved in this, we analyzed tapinarof-treated normal human epidermal keratinocytes (NHEKs). Tapinarof upregulated FLG and LOR mRNA and protein expression in an AHR-dependent manner. Tapinarof also induced the secretion of IL-24, a cytokine that activates Janus kinase (JAK)-signal transducer and activator of transcription (STAT), leading to the downregulation of FLG and LOR expression. Knockdown of either IL-24 or STAT3 expression by small interfering RNA (siRNA) transfection augmented the upregulation of FLG and LOR expression induced by tapinarof, suggesting that inhibition of the IL-24/STAT3 axis during AHR activation supports the improvement of skin barrier dysfunction. Furthermore, tapinarof alone could restore the downregulation of FLG and LOR expression induced by IL-4, a key cytokine of AD, and its combination with JAK inhibitors enhanced this effect. These findings provide a new strategy for treating AD using AHR modulators and JAK inhibitors.

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Siraitia grosvenorii Residual Extract Attenuates Atopic Dermatitis by Regulating Immune Dysfunction and Skin Barrier Abnormality

Nutrients ◽

10.3390/nu12123638 ◽

2020 ◽

Vol 12 (12) ◽

pp. 3638

Author(s):

Yoon-Young Sung ◽

Heung-Joo Yuk ◽

Won-Kyung Yang ◽

Seung-Hyung Kim ◽

Dong-Seon Kim

Keyword(s):

Atopic Dermatitis ◽

Immunoglobulin E ◽

Allergic Inflammation ◽

Skin Barrier ◽

Human Keratinocytes ◽

Skin Lesions ◽

Protein Levels ◽

Siraitia Grosvenorii ◽

Ifn Γ

Atopic dermatitis is a persistent inflammatory skin disorder. Siraitia grosvenorii fruits (monk fruit or nahangwa in Korean, NHG) are used as a natural sweetener and as a traditional medicine for the treatment of asthma and bronchitis. We evaluated the activity of S. grosvenorii residual extract (NHGR) on allergic inflammation of atopic dermatitis in a Dermatophagoides farinae mite antigen extract (DfE)-treated NC/Nga murine model and in vitro. Oral administration of NHGR significantly reduced epidermal hyperplasia and inflammatory cell infiltration in the skin lesions of DfE-induced atopic dermatitis, as well as the dermatitis severity score. NHGR reduced serum immunoglobulin E levels. Splenic concentrations of IFN-γ, interleukin (IL)-4, IL-5, and IL-13 were reduced by NHGR administration. Immunohistofluorescence staining showed that NHGR administration increased the protein levels of claudin-1, SIRT1, and filaggrin in atopic dermatitis skin lesions. In addition, NHGR inhibited the phosphorylation of mitogen-activated protein kinases and decreased filaggrin and chemokine protein expression in TNF-α/IFN-γ-induced human keratinocytes. Moreover, NHGR also inhibited histamine in mast cells. The quantitative analysis of NHGR revealed the presence of grosvenorine, kaempferitrin, and mogrosides. These results demonstrate that NHGR may be an efficient therapeutic agent for the treatment of atopic dermatitis.

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Critical role for Epac1 in inflammatory pain controlled by GRK2-mediated phosphorylation of Epac1

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1516036113 ◽

2016 ◽

Vol 113 (11) ◽

pp. 3036-3041 ◽

Cited By ~ 61

Author(s):

Pooja Singhmar ◽

XiaoJiao Huo ◽

Niels Eijkelkamp ◽

Susana Rojo Berciano ◽

Faiza Baameur ◽

...

Keyword(s):

Chronic Pain ◽

Molecular Mechanism ◽

Inflammatory Pain ◽

Critical Role ◽

Specific Inhibitor ◽

Mechanical Hyperalgesia ◽

Dependent Manner ◽

Phosphorylation Event

cAMP signaling plays a key role in regulating pain sensitivity. Here, we uncover a previously unidentified molecular mechanism in which direct phosphorylation of the exchange protein directly activated by cAMP 1 (EPAC1) by G protein kinase 2 (GRK2) suppresses Epac1-to-Rap1 signaling, thereby inhibiting persistent inflammatory pain. Epac1−/− mice are protected against inflammatory hyperalgesia in the complete Freund’s adjuvant (CFA) model. Moreover, the Epac-specific inhibitor ESI-09 inhibits established CFA-induced mechanical hyperalgesia without affecting normal mechanical sensitivity. At the mechanistic level, CFA increased activity of the Epac target Rap1 in dorsal root ganglia of WT, but not of Epac1−/−, mice. Using sensory neuron-specific overexpression of GRK2 or its kinase-dead mutant in vivo, we demonstrate that GRK2 inhibits CFA-induced hyperalgesia in a kinase activity-dependent manner. In vitro, GRK2 inhibits Epac1-to-Rap1 signaling by phosphorylation of Epac1 at Ser-108 in the Disheveled/Egl-10/pleckstrin domain. This phosphorylation event inhibits agonist-induced translocation of Epac1 to the plasma membrane, thereby reducing Rap1 activation. Finally, we show that GRK2 inhibits Epac1-mediated sensitization of the mechanosensor Piezo2 and that Piezo2 contributes to inflammatory mechanical hyperalgesia. Collectively, these findings identify a key role of Epac1 in chronic inflammatory pain and a molecular mechanism for controlling Epac1 activity and chronic pain through phosphorylation of Epac1 at Ser-108. Importantly, using the Epac inhibitor ESI-09, we validate Epac1 as a potential therapeutic target for chronic pain.

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20-Hydroxyvitamin D2is a noncalcemic analog of vitamin D with potent antiproliferative and prodifferentiation activities in normal and malignant cells

AJP Cell Physiology ◽

10.1152/ajpcell.00203.2010 ◽

2011 ◽

Vol 300 (3) ◽

pp. C526-C541 ◽

Cited By ~ 67

Author(s):

Andrzej T. Slominski ◽

Tae-Kang Kim ◽

Zorica Janjetovic ◽

Robert C. Tuckey ◽

Radoslaw Bieniek ◽

...

Keyword(s):

Vitamin D ◽

Catalytic Efficiency ◽

Human Keratinocytes ◽

Melanoma Cells ◽

Time Dependent ◽

Epidermal Keratinocytes ◽

Dependent Manner ◽

Cell Type ◽

Comparable Rate

20-hydroxyvitamin D2[20(OH)D2] inhibits DNA synthesis in epidermal keratinocytes, melanocytes, and melanoma cells in a dose- and time-dependent manner. This inhibition is dependent on cell type, with keratinocytes and melanoma cells being more sensitive than normal melanocytes. The antiproliferative activity of 20(OH)D2is similar to that of 1,25(OH)2D3and of newly synthesized 1,20(OH)2D2but significantly higher than that of 25(OH)D3. 20(OH)D2also displays tumorostatic effects. In keratinocytes 20(OH)D2inhibits expression of cyclins and stimulates involucrin expression. It also stimulates CYP24 expression, however, to a significantly lower degree than that by 1,25(OH)2D3or 25(OH)D3. 20(OH)D2is a poor substrate for CYP27B1 with overall catalytic efficiency being 24- and 41-fold lower than for 25(OH)D3with the mouse and human enzymes, respectively. No conversion of 20(OH)D2to 1,20(OH)2D2was detected in intact HaCaT keratinocytes. 20(OH)D2also demonstrates anti-leukemic activity but with lower potency than 1,25(OH)2D3. The phenotypic effects of 20(OH)D2are mediated through interaction with the vitamin D receptor (VDR) as documented by attenuation of cell proliferation after silencing of VDR, by enhancement of the inhibitory effect through stable overexpression of VDR and by the demonstration that 20(OH)D2induces time-dependent translocation of VDR from the cytoplasm to the nucleus at a comparable rate to that for 1,25(OH)2D3. In vivo tests show that while 1,25(OH)2D3at doses as low as 0.8 μg/kg induces calcium deposits in the kidney and heart, 20(OH)D2is devoid of such activity even at doses as high as 4 μg/kg. Silencing of CY27B1 in human keratinocytes showed that 20(OH)D2does not require its transformation to 1,20(OH)2D2for its biological activity. Thus 20(OH)D2shows cell-type dependent antiproliferative and prodifferentiation activities through activation of VDR, while having no detectable toxic calcemic activity, and is a poor substrate for CYP27B1.

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Tissue-specific contributions of Tmem79 to atopic dermatitis and mast cell-mediated histaminergic itch

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1814132115 ◽

2018 ◽

Vol 115 (51) ◽

pp. E12091-E12100 ◽

Cited By ~ 11

Author(s):

Joshua J. Emrick ◽

Anubhav Mathur ◽

Jessica Wei ◽

Elena O. Gracheva ◽

Karsten Gronert ◽

...

Keyword(s):

Oxidative Stress ◽

Atopic Dermatitis ◽

Mast Cell ◽

Transmembrane Protein ◽

Skin Barrier ◽

Histamine Receptor ◽

Cyclooxygenase Inhibitors ◽

Dependent Manner ◽

Common Skin ◽

Ep3 Receptor

Atopic dermatitis (AD) is the most common skin disease in children. It is characterized by relapsing inflammation, skin-barrier defects, and intractable itch. However, the pathophysiology of itch in AD remains enigmatic. Here, we examine the contribution of Tmem79, an orphan transmembrane protein linked to AD in both mice and humans. We show that Tmem79 is expressed by both keratinocytes and sensory neurons, but that loss of keratinocytic Tmem79 is sufficient to elicit robust scratching. Tmem79−/− mice demonstrate an accumulation of dermal mast cells, which are diminished following chronic treatment with cyclooxygenase inhibitors and an EP3 receptor antagonist. In Tmem79−/− mice, mast cell degranulation produces histaminergic itch in a histamine receptor 1/histamine receptor 4 (H4R/H1R)-dependent manner that may involve activation of TRPV1− afferents. TMEM79 has limited sequence homology to a family of microsomal glutathione transferases and confers protection from cellular accumulation of damaging reactive species, and may thus play a role in regulating oxidative stress. In any case, mechanistic insights from this model suggest that therapeutics targeting PGE2 and/or H1R/H4R histaminergic signaling pathways may represent useful avenues to treat Tmem79-associated AD itch. Our findings suggest that individuals with mutations in Tmem79 develop AD due to the loss of protection from oxidative stress.

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Crocin Ameliorates Atopic Dermatitis Symptoms by down Regulation of Th2 Response via Blocking of NF-κB/STAT6 Signaling Pathways in Mice

Nutrients ◽

10.3390/nu10111625 ◽

2018 ◽

Vol 10 (11) ◽

pp. 1625 ◽

Cited By ~ 10

Author(s):

Yoon-Young Sung ◽

Ho Kim

Keyword(s):

Atopic Dermatitis ◽

Signaling Pathways ◽

Crude Extract ◽

Th2 Cells ◽

Major Constituent ◽

Therapeutic Effects ◽

Dependent Manner ◽

Dermatophagoides Farinae ◽

Th2 Response ◽

Down Regulation

Crocin, a major constituent of Gardenia jasminoides, is a natural colorant carotenoid compound that has been reported to have anti-inflammatory effects. This study investigated the therapeutic effects of crocin on mice with atopic dermatitis induced by Dermatophagoides farinae crude extract, which is a common environmental allergen in house dust that causes atopic dermatitis in humans. Crocin application ameliorated Dermatophagoides farinae crude extract-induced atopic dermatitis symptoms by inhibiting the dermatitis severity score, ear thickness, and serum immunoglobulin E levels in NC/Nga mice. The increases in epidermal thickness and dermal inflammatory cells (eosinophil and mast cells) infiltrations observed on the dorsal back skin of atopic dermatitis control mice were inhibited in a dose-dependent manner by topical application of crocin in atopic dermatitis treatment mice. Crocin inhibited the Dermatophagoides farinae crude extract-induced increase of thymus and activation-regulated chemokines, interleukin (IL)-4, and IL-13 on the dorsal skin of mice. Crocin also inhibited Dermatophagoides farinae crude extract-induced activation of nuclear factor-κB (NF-κB) and signal transducer and activator of transcription (STAT) 6. These results show that crocin ameliorates atopic dermatitis symptoms by down regulation of the Th2 cells-mediated immune response via blocking of NF-κB/STAT6 signaling pathways.

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1-Ethyl-β-N-acetylglucosaminide increases hyaluronan production in human keratinocytes by being converted to N-acetylglucosamine via β-N-acetylglucosaminidase-dependent manner

Bioscience Biotechnology and Biochemistry ◽

10.1093/bbb/zbab060 ◽

2021 ◽

Author(s):

Yumiko Akazawa ◽

Hiroyuki Yoshida ◽

Yoko Endo ◽

Jun Sugita ◽

Masafumi Yakumaru ◽

...

Keyword(s):

Mrna Expression ◽

Human Keratinocytes ◽

Epidermal Keratinocytes ◽

Dependent Manner ◽

Intracellular Level ◽

Human Epidermal Keratinocytes ◽

Synthetic Substrate ◽

Epidermal Homeostasis ◽

Ha Synthase

Abstract Regulation of hyaluronan (HA) is important for the maintenance of epidermal homeostasis. Here we examined the mechanism by which 1-ethyl-β-N-acetylglucosaminide (β-NAG2), a newly developed N-acetylglucosamine (NAG) derivative, increases HA production in cultured human epidermal keratinocytes. When keratinocytes were treated with β-NAG2, mRNA expression of HA synthase 3, which is responsible for HA production in human keratinocytes, was not influenced, but the intracellular level of UDP-NAG, a substrate used for HA synthesis, was increased. By using a synthetic substrate for β-N-acetylglucosaminidase (β-NAGase), keratinocytes were found to possess β-NAGase activity, and treatment of o-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino N-phenyl carbamate (PUGNAc), an inhibitor of β-NAGase, abolished the release of NAG from β-NAG2 in keratinocytes. Furthermore, PUGNAc attenuated the β-NAG2-induced intracellular UDP-NAG and HA production in keratinocytes. These results suggest that β-NAG2 is converted to NAG by endogenous β-NAGase in keratinocytes, and the resulting NAG is further metabolized to UDP-NAG and utilized for HA production.

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Yokukansan, a Traditional Japanese Medicine, Adjusts Glutamate Signaling in Cultured Keratinocytes

BioMed Research International ◽

10.1155/2014/364092 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 7

Author(s):

Maki Wakabayashi ◽

Toshio Hasegawa ◽

Takuji Yamaguchi ◽

Naoko Funakushi ◽

Hajime Suto ◽

...

Keyword(s):

Glutamate Release ◽

Skin Barrier ◽

Human Keratinocytes ◽

Dependent Manner ◽

Extracellular Glutamate ◽

Concentration Dependent Manner ◽

Glutamate Signaling ◽

Cultured Keratinocytes ◽

Traditional Japanese Medicine ◽

Mrna Expression Levels

Glutamate plays an important role in skin barrier signaling. In our previous study, Yokukansan (YKS) affected glutamate receptors in NC/Nga mice and was ameliorated in atopic dermatitis lesions. The aim of this study was to assess the effect of YKS on skin and cultured human keratinocytes. Glutamate concentrations in skin of YKS-treated and nontreated NC/Nga mice were measured. Then, glutamate release from cultured keratinocytes was measured, and extracellular glutamate concentrations in YKS-stimulated cultured human keratinocytes were determined. The mRNA expression levels of NMDA receptor 2D (NMDAR2D) and glutamate aspartate transporter (GLAST) were also determined in YKS-stimulated cultured keratinocytes. The glutamate concentrations and dermatitis scores increased in conventional mice, whereas they decreased in YKS-treated mice. Glutamate concentrations in cell supernatants of cultured keratinocytes increased proportionally to the cell density. However, they decreased dose-dependently with YKS. YKS stimulation increased NMDAR2D in a concentration-dependent manner. Conversely, GLAST decreased in response to YKS. Our findings indicate that YKS affects peripheral glutamate signaling in keratinocytes. Glutamine is essential as a transmitter, and dermatitis lesions might produce and release excess glutamate. This study suggests that, in keratinocytes, YKS controls extracellular glutamate concentrations, suppresses N-methyl-D-aspartate (NMDA) receptors, and activates glutamate transport.

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The role of interleukin-24 in atopic dermatitis

10.37349/ei.2021.00002 ◽

2021 ◽

Author(s):

Yen Hai Vu ◽

Masutaka Furue ◽

Gaku Tsuji

Keyword(s):

Atopic Dermatitis ◽

Inflammatory Responses ◽

Skin Barrier ◽

Human Keratinocytes ◽

Barrier Disruption ◽

Aryl Hydrocarbon ◽

Receptor Modulator ◽

Chronic Pruritus

Atopic dermatitis (AD) is characterized by skin barrier disruption, type 2 immune dysregulation, chronic pruritus, and abnormal colonization by Staphylococcus aureus (S. aureus). Tapinarof, an aryl hydrocarbon receptor modulator, has been demonstrated to attenuate the development of AD in clinical studies. Recently, we found that tapinarof upregulated the expression of filaggrin and loricrin, which are essential proteins in skin barrier functions. Paradoxically, tapinarof induced interleukin (IL)-24 secretion by normal human keratinocytes. IL-24 is produced by T helper 2 lymphocytes and keratinocytes following stimulation by type 2 cytokines, and IL-24 is upregulated in the skin of patients with AD. Furthermore, IL-24 contributes to skin barrier disruption and hyperplasia in AD, and it may exacerbate skin inflammatory responses, itch, and S. aureus infection. In this review, we summarized the current findings regarding the detrimental role of IL-24 in AD, thereby suggesting that co-treatment of tapinarof with therapeutics that block IL-24 signaling may represent a promising strategy for managing AD.

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Low-Temperature Argon Plasma Regulates Skin Moisturizing and Melanogenesis-Regulating Markers through Yes-Associated Protein

International Journal of Molecular Sciences ◽

10.3390/ijms22041895 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1895

Author(s):

Hae-Young Kim ◽

Gaurav Agrahari ◽

Min Jung Lee ◽

Lee-Jung Tak ◽

Won-Kook Ham ◽

...

Keyword(s):

Low Temperature ◽

Argon Plasma ◽

Skin Barrier ◽

Dermal Fibroblasts ◽

Expression Level ◽

Epidermal Keratinocytes ◽

Type I ◽

Dependent Manner ◽

Expression Levels ◽

Normal Human

Extensive water loss and melanin hyperproduction can cause various skin disorders. Low-temperature argon plasma (LTAP) has shown the possibility of being used for the treatment of various skin diseases, such as atopic dermatitis and skin cancer. However, the role of LTAP in regulating skin moisturizing and melanogenesis has not been investigated. In this study, we aimed to determine the effect of LTAP on yes-associated protein (YAP), a major transcriptional coactivator in the Hippo signaling pathway that is involved in skin moisturizing and melanogenesis-regulating markers. In normal human epidermal keratinocytes (NHEKs), the human epidermal keratinocyte line HaCaT, and human dermal fibroblasts (HDFs), we found that LTAP exhibited increased expression levels of YAP protein. In addition, the expression levels of filaggrin (FLG), which is involved in natural moisturizing factors (NMFs), and hyaluronic acid synthase (HAS), transglutaminase (TGM), and involucrin (IVL), which regulate skin barrier and moisturizing, were also increased after exposure to LTAP. Furthermore, collagen type I alpha 1 and type III alpha 1 (COL1A1, COL3A1) were increased after LTAP exposure, but the expression level of matrix metalloproteinase-3 (MMP-3) was reduced. Moreover, LTAP was found to suppress alpha-melanocyte stimulating hormone (α-MSH)-induced melanogenesis in murine melanoma B16F10 cells and normal human melanocytes (NHEMs). LTAP regulates melanogenesis of the melanocytes through decreased YAP pathway activation in a melanocortin 1 receptor (MC1R)-dependent manner. Taken together, our data show that LTAP regulates skin moisturizing and melanogenesis through modulation of the YAP pathway, and the effect of LTAP on the expression level of YAP varies from cell to cell. Thus, LTAP might be developed as a treatment method to improve the skin barrier, moisture content, and wrinkle formation, and to reduce melanin generation.

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