Hana Lee

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

IL-33–activated mast cells and ILC2s, and may be involved in severe asthma via the recruitment of neutrophils and smooth muscle activation. Furthermore, the presence of other immune cells in the airway in an IL-33 milieu may result in an abundance of non–type 2 mediators, which could affect differential gene expression not otherwise seen in the coculture system. Thus, further studies need to be conducted in vivo to examine the effects of anti–IL-13 on airway inflammation, mucus expression, and hyperreactivity in response to IL-33. In conclusion, we have shown that IL-33–activated mast cells and ILC2s drive predominantly IL-13–regulated gene expression when cocultured with NHBE cells, which initiates a transcriptional program that can subsequently exacerbate airway pathology and contribute to fibrosis, eosinophilia, and mucous metaplasia. Although mast cells and ILC2s produce a diverse set of cytokines and other mediators of intercellular communication in response to IL-33, the predominant effect on airway epithelium in this in vitro coculture system is mediated by IL-13.

UVB Induces HIF-1α-Dependent TSLP Expression via the JNK and ERK Pathways

Thymic stromal lymphopoietin (TSLP) may have a key role in the initiation and maintenance of allergic inflammatory diseases, including atopic dermatitis. The present study revealed that UVB radiation exposure could induce TSLP expression in human keratinocytes and a human skin equivalent model. In addition, we investigated the regulatory mechanism of UVB-induced TSLP expression in keratinocytes. TSLP expression was upregulated by transfection with pcDNA3-hypoxia-inducible factor (HIF)-1α (P402A and P564A), which stably expresses HIF-1α protein. UVB-induced TSLP induction in keratinocytes was suppressed in the treatment of mitogen-activated protein kinase inhibitors or small interfering RNAs against HIF-1α. The results of chromatin immunoprecipitation assays indicate the direct involvement of HIF-1α in UVB-mediated TSLP induction. Taken together, these findings indicate that UVB exposure may increase TSLP expression through a HIF-1α-dependent mechanism via the c-JUN N-terminal kinase and extracellular signal-regulated kinase pathways in human keratinocytes. Our data showed that UVB-induced TSLP might increase secretion of the T-helper type 2-attracting chemokine (c-c motif) ligand 17 by human dendritic cells. The present study suggests an important role of HIF-1α in UVB-mediated immune response in keratinocytes.

IL‐33 induces Egr‐1‐dependent TSLP expression via the MAPK pathways in human keratinocytes

Atopic dermatitis (AD) is a chronic inflammatory skin disease in which T‐helper type 2 (Th2)‐type immune responses are dominant. Th2 cytokine, interleukin (IL)‐33 and thymic stromal lymphopoietin (TSLP) have been suggested to have an important role in AD. IL‐33 is highly expressed in AD, but its role in AD has not yet been fully understood. To further identify the role of IL‐33 in AD, we investigated the expression of TSLP induced by IL‐33 in keratinocytes. This study revealed that IL‐33 induced TSLP expression in human keratinocytes. Early growth response protein 1 (Egr)‐1, which is an inflammatory transcriptional factor, is induced by IL‐33. IL‐33‐mediated TSLP induction in keratinocytes was suppressed by treatment with mitogen‐activated protein kinase (MAPK) inhibitors or small interfering RNA against Egr‐1. Chromatin immunoprecipitation (ChIP) assay indicated the direct involvement of Egr‐1 in IL‐33‐mediated TSLP induction. Taken together, these findings indicate that IL‐33 may increase TSLP expression through an Egr‐1‐dependent mechanism via ERK1/2, JNK and p38 activation in keratinocytes. These data suggest that the IL‐33‐ERK/JNK/p38/Egr‐1/TSLP axis is involved in allergic skin Th2 inflammation, and it may be a novel therapeutic target.

TSLP Down-Regulates S100A7 and ß-Defensin 2 Via the JAK2/STAT3-Dependent Mechanism

Elevated T-helper type 2 cytokines in atopic skin, such as IL-4 and IL-13, were thought to be responsible for an impaired expression of antimicrobial proteins, which may contribute to the increased susceptibility to skin infections in patients with atopic dermatitis. In this study, the relationship between thymic stromal lymphopoietin and antimicrobial proteins and the involved molecular pathway was defined in normal human epidermal keratinocytes and human skin equivalent model. Stimulation of normal human epidermal keratinocytes with thymic stromal lymphopoietin decreased both mRNA and levels of S100A7 and human β-defensin 2 in a dose-dependent manner, and the regulation was JAK2/STAT3-dependent. Thymic stromal lymphopoietin decreased the antimicrobial protein expression, even in the presence of IL-17, which is their strong inducer. STAT3 directly regulated the S100A7 and human β-defensin 2 promoters in normal human epidermal keratinocytes. Immunohistochemically, lesional atopic skin stained more intensely with phospho-STAT3 compared with healthy control. Our results show that up-regulated thymic stromal lymphopoietin may contribute to the deficiency of antimicrobial proteins in atopic dermatitis, including S100A7 and human β-defensin 2, by a JAK2/STAT3-dependent mechanism and that STAT3/Sin3a might directly control the transcriptional activity of the antimicrobial protein promoters in normal human epidermal keratinocytes. Taken together, a key role of the JAK2/STAT3/Sin3a signaling pathway in thymic stromal lymphopoietin-mediated immune response in normal human epidermal keratinocytes might give us clues to understanding the pathological signal transductions in atopic dermatitis.

Egr-1 is a key regulator of IL-17A-induced psoriasin upregulation in psoriasis

The early growth response (Egr)‐1 is a transcriptional factor which plays an important role in the regulation of cell growth, differentiation, cell survival and immune responses. Emerging evidences including our data demonstrate that the Egr‐1 expression is up‐regulated in the psoriatic skin lesions. The purpose of this study was to investigate the significance and regulatory mechanism of Egr‐1 in the pathogenesis of psoriasis. Through microarray analysis, we found out that psoriasin (S100A7) expression was increased in the Egr‐1 overexpressed cells. Our results showed that IL‐17A increased Egr‐1 expression in the skin of psoriatic patients and cultured human keratinocytes. We then investigated activation of mitogen‐activated protein kinase as an upstream signal regulator of Egr‐1 expression. IL‐17A‐induced Egr‐1 expression was suppressed by ERK inhibitor. In addition, IL‐17A induced psoriasin expression in cultured keratinocytes and the skin of IL‐17A intradermally injected mouse. IL‐17A‐mediated psoriasin upregulation was reduced after treatment of small interfering RNAs against Egr‐1. Furthermore, the results of chromatin immunoprecipitation assays demonstrated that Egr‐1 directly binds the psoriasin promoter. Our findings present a novel signalling mechanism by which IL‐17A can induce the Egr‐1‐dependent psoriasin expression via the ERK pathway in human keratinocytes. This study suggests that Egr‐1 may be a novel and important modulator in IL‐17A‐mediated immune response in psoriasis.

Toluene induces early growth response-1 dependent thymic stromal lymphopoietin expression in human keratinocytes

Volatile organic compounds (VOCs) can be an important environmental risk factor in the development and/or aggravation of atopic dermatitis (AD). In a several studies to evaluate the clinical effects of air pollution including VOCs on skin symptoms in AD patients. Although these accumulated data shows an association between exposure to VOCs and aggravation of AD, the molecular effects of VOCs on the aggravation of AD is still not clear. The present study demonstrates increases of thymic stromal lymphopoietin (TSLP) in keratinocytes following exposure to toluene. We further investigated the regulatory mechanisms of toluene-induced TSLP expression in human keratinocytes. Toluene induces early growth response-1 (Egr-1) protein expression in human keratinocytes. Moreover, treatment of Egr-1 siRNA abolished toluene-induced TSLP protein expression. Taken together, these findings suggest that in human keratinocytes, the up-regulation of TSLP by toluene is induced through an Egr-1 dependent mechanism.

Chloroform induces cystein-rich 61, a mediator of collagen homeostasis via early growth response-1 dependent pathway in human skin dermal fibroblasts

Volatile organic compounds (VOCs) are major air pollutants known to have adverse effects on skin. Many reports have shown an association between air pollutants and skin aging. However, there is no report on VOCs association with skin aging. The aim of this study is to identify the biologic effects of VOCs on fibroblasts. We investigated the effects of chloroform on cysteine-rich 61 (CYR61) expression in human skin fibroblasts using western blot and reverse transcriptionpolymerase chain reaction. We also observed the change of type I procollagen and matrix metalloproteinase after exposure to chloroform using western blot. We explored the involvement of early growth response-1 (Egr-1) using siRNA s in the fibroblasts. Chloroform exposure increased the expression of CYR61 protein and mRNA level. Elevated CYR61 downregulates type I procollagen and upregulates MMP-1. Egr-1 overexpression upregulates CYR61 promoter activity. CYR61 promoter activity was lower in Egr-1 siRNA-transfected fibroblasts. The present study demonstrates that CYR61 is transcriptionally regulated by chloroform exposure through transcription factor Egr-1, and alters collagen homeostasis in human skin fibroblasts.

Intracellular ROS levels determine the apoptotic potential of keratinocyte by Quantum Dot via blockade of AKT Phosphorylation

Quantum dots (QDs) have shown great potential for biomedical use in a broad range including diagnostic agents. However, the regulatory mechanism of dermal toxicity is poorly understood. In this study, we investigated how QDs‐induced apoptosis is regulated in human keratinocytes. We also examined the effect of carboxylic acid‐coated QDs (QD 565 and QD 655) on reactive oxygen species (ROS) production and apoptosis‐related cellular signalling. The viability of keratinocyte was inhibited by two types of QDs in a concentration‐dependent manner. QDs induce ROS production and blockade of AKT phosphorylation. Moreover, the cleavage of AKT‐dependent pro‐apoptotic proteins such as poly (ADP‐ribose) polymerase, caspases‐3 and caspases‐9 was significantly increased. We also found that a decrease in cellular ROS level by ROS scavenger, N‐acetylcysteine (NAC), resulting in the abolishment of QDs‐induced AKT de‐phosphorylation and cellular apoptosis. Interestingly, QD 655 had a more cytotoxic effect including oxidative stress and AKT‐dependent apoptosis than QD 565. In addition, QD 655 had the cytotoxic potential in the human skin equivalent model (HSEM). These data show that QD‐induced intracellular ROS levels may be an important parameter in QD‐induced apoptosis. These findings from this study indicate that intracellular ROS levels might determine the apoptotic potential of keratinocyte by QD via blockade of AKT phosphorylation.

IL-33 down-regulates filaggrin expression by inducing STAT3 and ERK phosphorylation in human keratinocytes.

Atopic dermatitis (AD) is characterized by immune dysregulation and skin barrier disruption. Filaggrin (FLG) is an important structural protein in corneocytes that form the outermost layer of skin and provide skin barrier function, and its intracellular metabolite functions as a hydrator of the stratum corneum and a regulator of skin pH. AD is thought to develop in persons with genetic risk factors for skin barrier dysfunction and immune dysregulation. Eventually, it can be considered to be a reciprocal action in which immune dysregulation by increasing inflammatory cytokines cause skin barrier dysfunction and immune dysregulation. It follows the outside-inside-outside pathogenic loop in AD which was suggested by Elias. Elias et al. [1] suggested that barrier disruption makes allergen and bacterial penetration easy in atopic dermatitis and this causes an increase in cytokines and subsequently these cytokines in turn compromise barrier function. AD is characterized by defective barrier function that is associated with filaggrin deficit. Since Palmar et al. [2] demonstrated that AD is related to filaggrin mutation, there have been many reports showing the relationship between FLG mutation and severe AD. However, there are many AD patients without FLG null mutation who have decreased expression of FLG. Therefore, acquired factors are thought to be involved in the process of barrier formation. Gutowskia-Owsiak and Ogg [3] demonstrated several cytokines involved in down-regulation of FLG expression. Interleukin (IL)-4, IL-13, tumor necrosis factor (TNF)-a, IL-17, IL-22, IL-25, and IL-31 are associated with decreased FLG production. Recent report mentioned IL-33 as an important cytokine in the pathogenesis of AD, and IL-33 is also thought to be related to decreased production of FLG [4]. IL-33 plays a role as an ‘alarmin’, when is released in tissue damage, and also activate T-helper type 2 (Th2) immune responses [5]. Nonetheless, the IL-33 pathway responsible for decreasing FLG production has not yet been elucidated. IL-33 is a well-known IL-1 family cytokine which can activates mast cell and Th2 lymphocytes. Also, IL-33 activates innate immune response immediately in tissue damage [5]. IL-33 is suggested to play a role in this pathogenic mechanism of outside– inside–outside loop in AD. However, the regulatory mechanism of IL-33 in AD patients is not fully understood in keratinocytes yet.

Chloroform upregulates early growth response-1-dependent thymic stromal lymphopoietin expression via the JNK and ERK pathways in human keratinocytes

Exposure to volatile organic compounds (VOCs) in the environment has been reported to exacerbate allergic inflammatory diseases, such as atopic dermatitis (AD). However, the exact mechanism by which VOCs induce an inflammatory response in the skin is poorly understood. Thymic stromal lymphopoietin (TSLP) is known to be an important factor in the initiation and maintenance of allergic inflammatory diseases, including AD.