Keratinocytes-Derived Reactive Oxygen Species Play an Active Role to Induce Type 2 Inflammation of the Skin: A Pathogenic Role of Reactive Oxygen Species at the Early Phase of Atopic Dermatitis.

BACKGROUND: Atopic dermatitis (AD) is characterized by chronic, relapsing skin inflammation (eczema) with itchy sensation. Keratinocytes, which are located at the outermost part of our body, are supposed to play important roles at the early phase of type 2 inflammation including AD pathogenesis. OBJECTIVE: The purpose of this study was to evaluate whether keratinocytes-derived reactive oxygen species (ROS) could be produced by the allergens or non-allergens, and the keratinocytes-derived ROS could modulate a set of biomarkers for type 2 inflammation of the skin. METHODS: Normal human epidermal keratinocytes (NHEKs) were treated with an allergen of house dust mites (HDM) or a non-allergen of compound 48/80 (C48/80). Then, biomarkers for type 2 inflammation of the skin including those for neurogenic inflammation were checked by reverse transcriptase-polymerase chain reaction and western immunoblot experiments. RESULTS: HDM or C48/80 was found to upregulate expression levels of our tested biomarkers, including type 2 T helper-driving pathway (KLK5, PAR2, and NFκB), epithelial-cell-derived cytokines (thymic stromal lymphopoietin, interleukin [IL]-25, IL-33), and neurogenic inflammation (NGF, CGRP). The HDM- or C-48/80-induced expression levels of the biomarkers could be blocked by an antioxidant treatment with 5 mM N-acetyl-cysteine. In contrast, pro-oxidant treatment with 1 mM H2O2 could upregulate expression levels of the tested biomarkers in NHEKs. CONCLUSION: Our results reveal that keratinocytes-derived ROS, irrespective to their origins from allergens or non-allergens, have a potential to induce type 2 inflammation of AD skin.

Bee venom-derived antimicrobial peptide melectin has broad-spectrum potency, cell selectivity, and salt-resistant properties.

Antimicrobial peptides have attracted attention as alternatives to conventional antibiotics. Previously, a novel antimicrobial peptide, melectin, consisting of 18 amino acids was isolated from the venom of a bee, Melecta albifrons. Here, we investigated the antibacterial activity of melectin against drug-resistant bacteria. Melectin showed broad-spectrum antimicrobial activity but low cytotoxicity and no hemolytic activity. Melectin maintained its antimicrobial activity at physiological salt concentrations. Melectin is an α-helical structure that binds to the bacterial membrane via electrostatic interactions and kills bacteria in a short time by bacterial membrane targeting. Collectively, our results suggest that melectin has antibacterial activity and anti-inflammatory activity.

Global, Regional, and National Cancer Incidence, Mortality, Years of Life Lost, Years Lived With Disability, and Disability-Adjusted Life-Years for 29 Cancer Groups, 1990 to 2016: A Systematic Analysis for the Global Burden of Disease Study.

Importance: The increasing burden due to cancer and other noncommunicable diseases poses a threat to human development, which has resulted in global political commitments reflected in the Sustainable Development Goals as well as the World Health Organization (WHO) Global Action Plan on Non-Communicable Diseases. To determine if these commitments have resulted in improved cancer control, quantitative assessments of the cancer burden are required. Objective: To assess the burden for 29 cancer groups over time to provide a framework for policy discussion, resource allocation, and research focus. Evidence Review: Cancer incidence, mortality, years lived with disability, years of life lost, and disability-adjusted life-years (DALYs) were evaluated for 195 countries and territories by age and sex using the Global Burden of Disease study estimation methods. Levels and trends were analyzed over time, as well as by the Sociodemographic Index (SDI). Changes in incident cases were categorized by changes due to epidemiological vs demographic transition. Findings: In 2016, there were 17.2 million cancer cases worldwide and 8.9 million deaths. Cancer cases increased by 28% between 2006 and 2016. The smallest increase was seen in high SDI countries. Globally, population aging contributed 17%; population growth, 12%; and changes in age-specific rates, -1% to this change. The most common incident cancer globally for men was prostate cancer (1.4 million cases). The leading cause of cancer deaths and DALYs was tracheal, bronchus, and lung cancer (1.2 million deaths and 25.4 million DALYs). For women, the most common incident cancer and the leading cause of cancer deaths and DALYs was breast cancer (1.7 million incident cases, 535 000 deaths, and 14.9 million DALYs). In 2016, cancer caused 213.2 million DALYs globally for both sexes combined. Between 2006 and 2016, the average annual age-standardized incidence rates for all cancers combined increased in 130 of 195 countries or territories, and the average annual age-standardized death rates decreased within that timeframe in 143 of 195 countries or territories. Conclusions and Relevance: Large disparities exist between countries in cancer incidence, deaths, and associated disability. Scaling up cancer prevention and ensuring universal access to cancer care are required for health equity and to fulfill the global commitments for noncommunicable disease and cancer control.

Global, Regional, and National Burden of Cardiovascular Diseases for 10 Causes, 1990 to 2015.

BACKGROUND: The burden of cardiovascular diseases (CVDs) remains unclear in many regions of the world. OBJECTIVES: The GBD (Global Burden of Disease) 2015 study integrated data on disease incidence, prevalence, and mortality to produce consistent, up-to-date estimates for cardiovascular burden. METHODS: CVD mortality was estimated from vital registration and verbal autopsy data. CVD prevalence was estimated using modeling software and data from health surveys, prospective cohorts, health system administrative data, and registries. Years lived with disability (YLD) were estimated by multiplying prevalence by disability weights. Years of life lost (YLL) were estimated by multiplying age-specific CVD deaths by a reference life expectancy. A sociodemographic index (SDI) was created for each location based on income per capita, educational attainment, and fertility. RESULTS: In 2015, there were an estimated 422.7 million cases of CVD (95% uncertainty interval: 415.53 to 427.87 million cases) and 17.92 million CVD deaths (95% uncertainty interval: 17.59 to 18.28 million CVD deaths). Declines in the age-standardized CVD death rate occurred between 1990 and 2015 in all high-income and some middle-income countries. Ischemic heart disease was the leading cause of CVD health lost globally, as well as in each world region, followed by stroke. As SDI increased beyond 0.25, the highest CVD mortality shifted from women to men. CVD mortality decreased sharply for both sexes in countries with an SDI >0.75. CONCLUSIONS: CVDs remain a major cause of health loss for all regions of the world. Sociodemographic change over the past 25 years has been associated with dramatic declines in CVD in regions with very high SDI, but only a gradual decrease or no change in most regions. Future updates of the GBD study can be used to guide policymakers who are focused on reducing the overall burden of noncommunicable disease and achieving specific global health targets for CVD.

CSP0510 Lotion as a Novel Moisturizer Containing Citric Acid and Trisodium Phosphate Relieves Objective and Subjective Symptoms of Atopic Dermatitis.

BACKGROUND: Moisturizers with anti-inflammatory or anti-itch activity should be developed for the safe and effective management of atopic dermatitis (AD). OBJECTIVE: This study evaluated the efficacy of a newly developed moisturizer, CSP0510 lotion (Twolines Inc., Korea), containing citric acid (CA) and trisodium phosphate (TSP) as active ingredients, in mild to moderate AD. METHODS AND RESULTS: CSP0510 lotion applied twice daily for 4 weeks to eczematous lesions improved objective and subjective (itch) symptoms of AD. The physician's global assessment (PGA) score for objective symptoms decreased from 2.5±0.6 before application to 1.3±0.5 after application in the CSP0510-treated group (n=42, p<0.001). Also, the PGA score decreased from 2.3±0.6 to 1.9±0.5 by vehicle-treated (without CA and TSP) control group (p=0.001), but there was no statistical difference between CSP0510-treated and vehicle-treated groups (p=0.089). The visual analogue scale score for itch decreased from 4.8±1.3 to 2.0±0.9 in the CSP0510-treated group (p<0.001), and from 4.6±1.1 to 3.5±0.9 in the control group (p=0.075), showing a statistical significance between two groups (p=0.002). Our results in humans were further supported by in vitro and animal experiments. In HaCaT cells treated with compound 48/80 (7.5 µg/ml), CA:TSP (1:1, vol:vol) synergistically suppressed the compound 48/80-induced upregulation of thymic stromal lymphopoietin, nerve grow factor, and calcitonin gene-related peptide. Application of CSP0510 to the dorsal skin of hairless mice for 3 weeks suppressed the oxazolone-induced allergic skin inflammation. CONCLUSION: In conclusion, CSP0510 lotion has anti-itch and anti-inflammatory activity in the skin, which improves both objective and subjective symptoms of AD.

Ethanol Extract of Peanut Sprout Exhibits a Potent Anti-Inflammatory Activity in Both an Oxazolone-Induced Contact Dermatitis Mouse Model and Compound 48/80-Treated HaCaT Cells.

BACKGROUND: We developed an ethanol extract of peanut sprouts (EPS), a peanut sprout-derived natural product, which contains a high level of trans-resveratrol (176.75 µg/ml) and was shown to have potent antioxidant activity. OBJECTIVE: We evaluated the potential anti-inflammatory activity of EPS by measuring its antioxidant potential in skin. METHODS: The anti-inflammatory activity of EPS was tested using two models of skin inflammation: oxazolone (OX)-induced contact dermatitis in mice and compound 48/80-treated HaCaT cells. As biomarkers of skin inflammation, cyclooxygenase-2 (COX-2) and nerve growth factor (NGF) levels were measured. RESULTS: OX-induced contact dermatitis was suppressed markedly in mice that were treated with an ointment containing 5% EPS as evidenced by a decrease in the extent of scaling and thickening (p<0.05) and supported by a histological study. COX-2 (messenger RNA [mRNA] and protein) and NGF (mRNA) levels, which were upregulated in the skin of OX-treated mice, were suppressed markedly in the skin of OX+EPS-treated mice. Consistent with this, compound 48/80-induced expression of COX-2 (mRNA and protein) and NGF (mRNA) in HaCaT cells were suppressed by EPS treatment in a dose-dependent manner. As an inhibitor of NF-κB, IκB protein levels were dose-dependently upregulated by EPS. Fluorescence-activated cell sorting (FACS) analysis revealed that EPS scavenged compound 48/80-induced reactive oxygen species (ROS) in HaCaT cells. CONCLUSION: EPS exerts a potent anti-inflammatory activity via its anti-oxidant activity in both mouse skin and compound 48/80-treated HaCaT cells in vitro. Compound 48/80-treated HaCaT cells are a useful new in vitro model of skin inflammation.

Methyl-ß-cyclodextrin up-regulates collagen I expression in chronologically-aged skin via its anti-caveolin-1 activity.

Caveolin-1 (Cav-1) is one of the key molecules to modulate collagen metabolism in the skin. This study aimed to unravel the relationship between Cav-1 and collagen levels in the aged skin, and also to evaluate a new role of anti-Cav-1 agent as a collagen-modulating agent. A negative correlation between Cav-1 and collagen I (COL I) was detected in chronologically aged skin of humans and mice, which was further confirmed by Cav-1 knock-down or knock-out experiments. Next, we tested whether methyl-ß-cyclodextrin (MßCD) as a chemical Cav-1 inhibitor could be developed as a collagen-modulating agent in the skin. Testing different conditions of MßCD injection via the intra-dermal route revealed that 2.5% MßCD administered twice per week for two months showed a potent COL I-up-regulating activity, leading to the increase of skin thickness (P < 0.05) without adverse reactions such as skin fibrosis. In human dermal fibroblasts, MßCD treatment induced up-regulated COL I and down-regulated Cav-1, supporting the results of mouse experiments. Collectively, MßCD has a COL I-enhancing activity in chronologically-aged skin, where Cav-1 acts as a brake in COL I expression, suggesting its potential role for an anti-aging agent.

Up-regulation of cyclooxygenase 2 and matrix metalloproteinases-2 and -9 in cutaneous squamous cell carcinoma: active role of inflammation and tissue remodeling in carcinogenesis.

BACKGROUND: Tissue inflammation and remodeling have been extensively studied in various tumors in relation with their invasiveness and metastasis. OBJECTIVE: The purpose of this study was to investigate the change in tissue inflammation and remodeling markers in cutaneous squamous cell carcinoma (SCC). METHODS: Expression levels of cyclooxygenase-2 (COX-2) as an inflammatory marker and matrix metalloproteinases-2 and -9 (MMPs 2/9) as remodeling markers were studied in mouse and human SCCs. Western blot analysis and RT-PCR for COX-2 and MMPs 2/9 were performed with skin samples from SCC patients and chronic ultraviolet B (UVB)-induced SCC from hairless mice. RESULTS: mRNA and protein levels of COX-2 and MMPs 2/9 were up-regulated with the higher sensitivity for MMP-9 in mouse SCCs, which were induced by chronic UVB irradiation. Consistently, COX-2 and MMPs 2/9 were up-regulated with the higher sensitivity for MMP-9 in human SCCs. CONCLUSION: COX-2 and MMPs 2/9 are up-regulated in well-differentiated cutanous SCC. Our findings indicate that inflammatory and tissue remodeling processes are actively induced during carcinogenesis of cutaneous SCC.

Ethanol extract of peanut sprout induces Nrf2 activation and expression of antioxidant and detoxifying enzymes in human dermal fibroblasts: implication for its protection against UVB-irradiated oxidative stress.

A peanut sprout is known to contain a significant level of resveratrol, which was reported to have beneficial effects in our body due to its antioxidant activities. The purpose of this study was to evaluate the cytoprotective activity of ethanol extract of peanut sprout (EPS) from ultraviolet B (UVB)-induced oxidative stress in human dermal fibroblasts (HDF). EPS was revealed to contain 54.2 µg g(-1) of trans-resveratrol. The DCF-DA-positive reactive oxygen species level was increased by 50 mJ cm(-2) of UVB irradiation (2150 ± 450% of nonirradiated control), which was markedly suppressed by EPS treatment (180 ± 42% of control). Annexin V-positive apoptotic cell death induced by UVB irradiation (16.4 ± 4.5%) was also significantly inhibited by EPS treatment (6.7 ± 2.5%). EPS induced up-regulation and nuclear translocation of Nrf2, a transcription factor for antioxidant and detoxifying enzymes, in HDF as a dose-dependent manner. UVB irradiation up-regulated Nrf2-dependent enzymes of heme oxygenase-1, NAD(P)H:quinine oxidoreductase-1 and glutathione-S-transferase pi, and they were further stimulated by EPS treatment. Taken together, EPS is an efficient cytoprotective agent against UVB-induced oxidative stress by activation of Nrf2 and upregulation of Nrf2-relating antioxidant and detoxifying enzymes in HDF.

Nrf2-dependent and Nrf2-independent induction of phase 2 detoxifying and antioxidant enzymes during keratinocyte differentiation.

As antioxidant enzymes can be actively modulated during keratinocyte (KC) differentiation, this study was aimed to evaluate the modulation of a group of phase 2 detoxifying and antioxidant enzymes (phase 2 enzymes) during KC differentiation. In postconfluence-induced differentiation model of KC, heme oxygenase-1 (HO-1), NADP(H):quinone oxidoreductase-1 (NQO-1), and glutathione S-transferase pi (GSTpi) were up-regulated at a transcriptional level. In Western blot analysis, the phase 2 enzymes were up-regulated by H(2)O(2), but down-regulated by N-acetyl cysteine, indicating the active role of reactive oxygen species for their expression during KC differentiation. When a redox-sensitive NF-E2 related factor-2 (Nrf2), a key transcriptional factor for phase 2 enzymes, was knocked down by small interfering RNA transfection in differentiated KCs, only NQO-1 was down-regulated in both mRNA and protein levels. In human skin, expression levels of the phase 2 enzymes were up-regulated in the differentiated KC in the normal epidermis and keratotic foci in squamous cell carcinoma, further supporting the differentiation-dependent expression of phase 2 enzymes in vivo. This study demonstrates that a group of phase 2 enzymes are modulated during KC differentiation via either Nrf2-dependent (NQO-1) or Nrf2-independent (HO-1 and GSTpi) ways.

Differentiation-dependent expression of NADP(H):quinone oxidoreductase-1 via NF-E2 related factor-2 activation in human epidermal keratinocytes.

BACKGROUND: NADP(H):quinone oxidoreductase-1 (NQO-1) is known for its protective role in skin carcinogenesis, but the expression of NQO-1 during keratinocyte (KC) differentiation has not been studied. OBJECTIVE: The purpose of the current study was to evaluate modulation of NQO-1 and NF-E2-related factor-2 (Nrf2) during KC differentiation. METHODS: Normal human epidermal keratinocytes (NHEKs) were induced to differentiation by prolonged culture after confluency (postconfluence). RESULTS: NQO-1 was induced at the late stage of differentiation of NHEKs (7th day of postconfluence). The expression of postconfluence-induced NQO-1 was stimulated by 0.1 mM H(2)O(2), but attenuated by 5 mM N-acetylcysteine, implying that reactive oxygen species (ROS) are implicated in the expression of NQO-1 in differentiated KCs. Nrf2 was up-regulated at the earlier than NQO-1 induction (3rd day of postconfluence). The Nrf2-dependent expression of NQO-1 was further supported by Nrf2-siRNA experiments. A confocal study confirmed the differentiation-dependent induction and activation of NOQ-1 and Nrf-2 in NHEKs. Immunohistochemistry showed that NQO-1 was accentuated in the upper epidermal layers, supporting the notion that differentiation-dependent NQO-1 expression is functional in human skin in vivo. CONCLUSION: These results demonstrate that NQO-1 is modulated during KC differentiation via Nrf2 pathway, suggesting the active role of NQO-1 in the differentiating epidermis.

Molecular changes following topical photodynamic therapy using methyl aminolaevulinate in mouse skin.

BACKGROUND: Photodynamic therapy (PDT) with aminolevulinic acid (ALA) or methyl aminolaevulinate (MAL) has been shown to enhance treatment of photoaged skin. However, there is little information about the molecular changes involved in dermal matrix remodeling following MAL-PDT for photorejuvenation. OBJECTIVE: We sought to analyze the molecular changes of the epidermal and dermal matrix after MAL-PDT in mouse skin. METHODS: Serial biopsy specimens were obtained at baseline and at various times after treatments with MAL-PDT, MAL alone and LED alone. To evaluate the molecular changes in the epidermal and dermal matrix, primary cytokines such as interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), transforming growth factor-beta1 (TGF-beta1), matrix metalloproteinases (MMPs), procollagen type I and III were evaluated by reverse transcriptase polymerase chain reaction, Western blot analysis, and immunohistochemistry assays. RESULTS: Elevation of primary cytokines and MMPs occurred at early points in time after one treatment with MAL-PDT based on the levels of mRNA and protein. On the other hand, procollagen type I protein increased later after MAL-PDT treatment. CONCLUSIONS: MAL-PDT activates more quantifiable alterations in the molecules associated with epidermal and dermal remodeling compared to treatment with MAL or LED alone. MAL-PDT significantly induced the epidermal and dermal matrix molecules required for photorejuvenation.

Propionibacterium acnes stimulates pro-matrix metalloproteinase-2 expression through tumor necrosis factor-alpha in human dermal fibroblasts.

Propionibacterium acnes (P. acnes) is a commensal microorganism found in sebum-rich skin and plays a role in acne inflammation by stimulating keratinocyte to produce a number of proinflammatory cytokines. However, the role of P. acnes in the dermis of acne lesions, where tissue remodeling after inflammation eventually takes place, is not known. In this study, we investigated whether P. acnes induces matrix metalloproteinase (MMP), a key enzyme involved in matrix remodeling in human dermal fibroblasts (hDF). We found that P. acnes increased expression of pro-matrix metalloproteinase (proMMP)-2 mRNA/protein in hDF, but not that of proMMP-9. Concomitantly, P. acnes induced tumor necrosis factor-alpha (TNF-alpha) mRNA/protein expression in hDF, which in turn increases both proMMP-2 mRNA and protein expression. P. acnes induced such changes through the activated NF-kappaB pathway. Doxycycline was found to inhibit the expression of proMMP-2 induced either by P. acnes or TNF-alpha. These results suggest that P. acnes stimulates hDF to produce TNF-alpha, which mediates the expression of proMMP-2 through the NF-kappaB pathway. The secretion of proMMP-2 from hDF upon P. acnes stimulation may contribute to the pathogenesis of tissue remodeling in acne skin.

TGF-beta1 up-regulates transglutaminase two and fibronectin in dermal fibroblasts: a possible mechanism for the stabilization of tissue inflammation.

Transglutaminase (TGase) has been reported to stabilize tissue inflammation via the mediation of the polymerization of extracellular matrix proteins. A set of cytokines has been implicated in wound healing processes in the dermis. This study was undertaken in order to evaluate the effects of these cytokines on the expression of TGase 2 in human dermal fibroblasts (hDFs), in that TGase 2 is known to be the principal TGase in the dermis. In Western blot analysis, TGF-beta1 (1 ng/ml) treatment was found to steadily up-regulate TGase 2 expression for up to 7 days. However, such increases were not observed when the cells were treated with IL-1beta, IL-2, and TNF-alpha. In the enzyme assay, total TGase activities were closely related to the levels of TGase 2 expression. TGase 2 mRNA expression was up-regulated as the result of TGF-beta treatment in competitive RT-PCR. In the denatured SDS-PAGE, TGF-beta1 treatment resulted in marked induction of an approximately 220 kDa protein, which was revealed to be a fibronectin (FN) via western immunoblotting with an anti-FN antibody. Next, when the hDFs were treated with TGF-beta1 (1 ng/ml), FN expression was induced beginning at the third day after treatment. The immunoprecipitants generated by anti-FN antibody were positive for the anti-TGase 2 antibody, and the immune complexes were identified at molecular weights of 92 kDa. Collectively, TGF-beta1 stimulates the polymerization of FN via the action of TGase 2, which is supposed to to be an important mechanism in the stabilization of the inflammatory dermis.

Chronic ultraviolet radiation modulates epidermal differentiation as it up-regulates transglutaminase 1 and its substrates.

BACKGROUND: Ultraviolet radiation (UVR) stimulates cellular mitosis, which leads to epidermal hyperplasia. On the basis of hypothesis that chronic UVR may modulate differentiation as well as epidermal hyperplasia, we evaluated the modulation of markers of epidermal differentiation, such as transglutaminase 1 (TGase 1), filaggrin and loricrin, by chronic UVR in vivo. METHODS: Total TGase activities assay or in situ TGase activities were measured in human and mouse skin. TGase 1 expression was identified by immunohistochemical staining in human skin. In the human, the pre-auricular skin of face was used for samples of chronic UVR, and the post-auricular skin was selected as non-UVR control. The changes of filaggrin and loricrin were identified by western immunoblots. RESULTS: In human and mouse epidermis, chronic UVR induced the increase of in situ TGase activities or total TGase activities as it up-regulated TGase 1 expression in the epidermis. As the substrates of TGase 1, chronic UVR induced the up-regulation of filaggrin and loricrin in mouse epidermis as well. At the same time, chronic UVR induced the marked epidermal hyperplasia in human and mouse skin. CONCLUSION: Chronic UVR stimulates epidermal differentiation as it up-regulates TGase 1 and its substrates. The modified epidermal differentiation is balanced with epidermal hyperplasia, leading to the maintenance of epidermal homeostasis in the UV-irradiated epidermis.