Hye Eun Lee

Expression of StMYB1R-1, a Novel Potato Single MYB-like domain Transcription Factor, Increases Drought Tolerance

Potato (Solanum tuberosum) is relatively vulnerable to abiotic stress conditions such as drought, but the tolerance mechanisms for such stresses in potato are largely unknown. To identify stress-related factors in potato, we previously carried out a genetic screen of potato plants exposed to abiotic environmental stress conditions using reverse northern-blot analysis. A cDNA encoding a putative R1-type MYB-like transcription factor (StMYB1R-1) was identified as a putative stress-response gene. Here, the transcript levels of StMYB1R-1 were enhanced in response to several environmental stresses in addition to drought but were unaffected by biotic stresses. The results of intracellular targeting and quadruple 9-mer protein-binding microarray analysis indicated that StMYB1R-1 localizes to the nucleus and binds to the DNA sequence G/AGATAA. Overexpression of a StMYB1R-1 transgene in potato plants improved plant tolerance to drought stress while having no significant effects on other agricultural traits. Transgenic plants exhibited reduced rates of water loss and more rapid stomatal closing than wild-type plants under drought stress conditions. In addition, overexpression of StMYB1R-1 enhanced the expression of drought-regulated genes such as AtHB-7, RD28, ALDH22a1, and ERD1-like. Thus, the expression of StMYB1R-1 in potato enhanced drought tolerance via regulation of water loss. These results indicated that StMYB1R-1 functions as a transcription factor involved in the activation of drought-related genes.

Association between Blood Cadmium Levels and 10-Year Coronary Heart Disease Risk in the General Korean Population: The Korean National Health and Nutrition Examination Survey 2008–2010

Background Non-occupational heavy metals are considered risk factors for coronary heart disease (CHD). Several recent epidemiologic studies have evaluated the relationship between non-occupational cadmium exposure and risk factors for cardiovascular disease (CVD). This study was designed to investigate the relationship between non-occupational cadmium exposure and risk factors for CHD using the Framingham estimate of 10 year CHD risk. Methods The heavy metal dataset of the Korean National Health and Nutrition Examination Survey for 2008 through 2010, a cross-sectional survey of a representative sample of 4,668 non-institutionalized Koreans, was analyzed. Subjects were stratified into seven age groups to minimize the effects of age. The log-transformed blood cadmium concentrations were compared with the Framingham estimate of 10 year CHD risk in each age stratum. Results The Framingham estimate of 10 year CHD risk was significantly associated with the log-transformed blood cadmium concentrations (p<0.05) in all age groups of Korean men, with the lowest regression coefficient (0.254) for men aged 20 to <35 years and the highest (3.354) for men aged 55 to <60 years; similar results, however, were not observed in Korean women. After adjusting for survey year, age, and urinary cotinine concentration, the log-transformed blood cadmium levels among men aged 20 to <35, 40 to <45, 50 to <55, and 60 to <65 years were significantly associated with systolic blood pressure (p<0.05), but not with total and high density lipoprotein (HDL) cholesterol concentrations. Conclusions Cadmium exposure, even at non-occupational levels, may be associated with CHD risk in men. Despite the declines in non-occupational cadmium exposure over the past several decades, more efforts are needed.

Oxidized phosphatidylcholine induces the activation of NLRP3 inflammasome in macrophages

The NOD‐like receptor family, pyrin domain containing 3 (NLRP3) inflammasome is a multiprotein complex consisting of a receptor, an adaptor protein, and procaspase‐1 that induces the secretion of the mature form of IL‐1β in response to microbial infection and danger signals. Activation of the NLRP3 inflammasome induced by endogenous danger signal molecules is closely linked to the development and progress of chronic inflammatory diseases. The oxidation of phospholipids occurs upon cellular stress and damage, resulting in the accumulation of oxidized phosphatidylcholines (oxPAPC) such as 1‐palmitoyl‐2‐(5‐oxovaleroyl)‐sn‐glycero‐phosphocholine (POVPC) at inflammatory sites. In this study, we investigated whether oxidized phosphatidylcholine induces the activation of NLRP3 inflammasome in macrophages, leading to the secretion of IL‐1β. POVPC induced the degradation of procaspase‐1 to caspase‐1(p10), the cleavage of pro‐IL‐1β to IL‐1β, and oligomerization of ASC in primary mouse bone marrow‐derived macrophages. POVPC‐induced production of caspase‐1, and IL‐1β was abolished in macrophages derived from NLRP3‐ or caspase‐1‐deficient mice. In an air pouch model and a peritonitis model in mice, POVPC injection resulted in the production of caspase‐1(p10), IL‐1β, and IL‐18 in wild‐type, but not in NLRP3‐deficient, mice. POVPC‐induced inflammasome activation was mediated by mitochondrial reactive oxygen species resulting from intracellular Ca2+ signaling and mitochondrial destabilization. Our results demonstrate that endogenously produced oxidized phosphatidylcholines such as POVPC induce the activation of NLRP3 inflammasome, leading to the production of IL‐1β in macrophages. The results provide an insight to understand how the oxidized lipids endogenously produced upon cellular stress and tissue damage contribute to the inflammatory reaction at pathologic sites.

Targeting ASC in NLRP3 inflammasome by caffeic acid phenethyl ester: a novel strategy to treat acute gout.

Gouty arthritis is caused by the deposition of uric acid crystals, which induce the activation of NOD-like receptor family, pyrin domain containing 3(NLRP3) inflammasome. The NLRP3 inflammasome, composed of NLRP3, the adaptor protein ASC, and caspase-1, is closely linked to the pathogenesis of various metabolic diseases including gouty arthritis. We investigated whether an orally administrable inhibitor of NLRP3 inflammasome was effective for alleviating the pathological symptoms of gouty arthritis and what was the underlying mechanism. In primary mouse macrophages, caffeic acid phenethyl ester(CAPE) blocked caspase-1 activation and IL-1β production induced by MSU crystals, showing that CAPE suppresses NLRP3 inflammasome activation. In mouse gouty arthritis models, oral administration of CAPE suppressed MSU crystals-induced caspase-1 activation and IL-1β production in the air pouch exudates and the foot tissues, correlating with attenuation of inflammatory symptoms. CAPE directly associated with ASC as shown by SPR analysis and co-precipitation, resulting in blockade of NLRP3-ASC interaction induced by MSU crystals. Our findings provide a novel regulatory mechanism by which small molecules harness the activation of NLRP3 inflammasome by presenting ASC as a new target. Furthermore, the results suggest the preventive or therapeutic strategy for NLRP3-related inflammatory diseases such as gouty arthritis using orally available small molecules.

Suppression of NLRP3 inflammasome by oral treatment with sulforaphane alleviates acute gouty inflammation

Objective The aetiology of gout is closely linked to the deposition of monosodium uric acid (MSU) crystals and the consequent activation of the NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome. In this study, we investigated whether oral administration of an NLRP3 inhibitor would be effective to attenuate the symptoms of gout. Methods The effects of oral administration with sulforaphane (SFN) were examined in two mouse models of acute gout induced by injection of MSU crystals into footpads or air pouch. The production of caspase-1 (p10) and IL-1β was examined by immunoblotting and ELISA as hallmarks of NLRP3 inflammasome activation. Results Oral administration of SFN attenuated MSU crystal-induced swelling and neutrophil recruitment in a mouse foot acute gout model, correlating with the suppression of the NLRP3 inflammasome activation in foot tissues. Consistently, oral administration of SFN blocked MSU-crystal-induced activation of the NLRP3 inflammasome in a mouse air pouch gout model. SFN suppressed NLRP3 inflammasome activation induced by MSU crystals, adenosine triphosphate and nigericin but not by poly(dA:dT) in primary mouse macrophages, independent of the reactive oxygen species pathway. SFN inhibited ligand-independent activation of the NLRP3 inflammasome, suggesting that SFN may act directly on the NLRP3 inflammasome complex. Conclusion Oral administration of SFN effectively alleviated acute gouty inflammation by suppression of the NLRP3 inflammasome. Our results provide a novel strategy in which oral treatment with SFN may be beneficial in preventing acute attacks of gout.

Topical Application of Dieckol Ameliorates Atopic Dermatitis in NC/Nga Mice by Suppressing Thymic Stromal Lymphopoietin Production

TO THE EDITOR Atopic dermatitis (AD) is a chronic, relapsing inflammatory skin disease that is characterized by extreme pruritus. AD is considered the first stage of atopic march (Leung et al., 2004). From an immunological aspect, AD is closely linked to the disruption of Th1/Th2 cytokine homeostasis that skews to Th2 immunity (Bieber, 2008). Immunological approaches to the treatment of AD have mainly focused on stimulating Th1 immunity to recover the Th1/Th2 balance (Leung et al., 2004). Thymic stromal lymphopoietin (TSLP) was recently shown to play a critical role in the progress of AD by inducing Th2 immune responses (Soumelis et al., 2002; Zhang et al., 2009). Increased TSLP production in keratinocytes induces the expression of the OX40 ligand (OX40L) in dendritic cells (DCs), which in turn stimulates the differentiation of naı̈ve CD4 T cells into Th2 cells to produce Th2 cytokines such as IL-4, IL-5, and IL-13 (Leyva-Castillo, Hener, Michea, et al., 2013). Thus, the TSLP-OX40L axis is considered integral to the induction of a Th2 cell-mediated allergic cascade in AD (MurakamiSatsutani et al., 2014). Indeed, high levels of TSLP expression have been observed in epidermal keratinocytes of AD skin lesions (Leyva-Castillo, Hener, Jiang, et al. 2013a). Therefore, suppressing TSLP may represent a novel therapeutic approach for treating AD by restoring Th1/Th2 balance. Here, we investigated whether dieckol, a phlorotannin from Ecklonia cava (Figure 1a), can suppress TSLP production to reduce Th2 immunity and effectively

Potentiation of skin TSLP production by a cosmetic colorant leads to aggravation of dermatitis symptoms

Certain cosmetic colorants are irritant to skin or aggravate dermatitis. Thymic stromal lymphopoietin (TSLP) plays an important role in the initiation and progress of skin inflammation and atopic dermatitis by triggering Th2 immune responses. However, the effects of cosmetic colorants on TSLP production are unknown yet. Therefore, we investigated whether cosmetic colorants regulated TSLP production and dermatitis. Lithol Rubine B (LR-B, Pigment Red 57) and its calcium salt (LR-BCA), commonly used cosmetic colorants, potentiated phorbol-12-myristate-13-acetate-induced TSLP production in keratinocytes. In addition, the topical exposure to LR-B or LR-BCA on mouse ear upregulated a TSLP inducer (MC903)-induced TSLP production and Th2 cytokine expression. Dermatitis symptoms and serum IgE and histamine levels were also aggravated by LR-B or LR-BCA, implicating the role of increased TSLP expression in acute dermatitis. LR-B or LR-BCA induced IκBα degradation and NF-κB activation in keratinocytes, leading to TSLP expression. Collectively, our results demonstrate that LR-B and LR-BCA increase TSLP expression and Th2 immune responses, thereby aggravating acute dermatitis in the compromised skin. The results further suggest that certain cosmetic colorants such as LR-B may aggravate dermatitis under pro-inflammatory conditions by upregulating TSLP production.

Licochalcone A attenuates acne symptoms mediated by suppression of NLRP3 inflammasome: Licochalcone A suppresses NLRP3 inflammasome and acne

Activation of the NACHT, LRR and PYD domains‐containing protein 3 (NLRP3) inflammasome by Propionibacterium acnes (P. acnes) is critical for inducing inflammation and aggravating the development of acne lesions. We searched for available small‐molecule inhibitors of the NLRP3 inflammasome that could be topically administered for the treatment of acne. We found that licochalcone A, a chalconoid isolated from the root of Glycyrrhiza inflate, was an effective inhibitor for P. acnes‐induced NLRP3 inflammasome activation. Licochalcone A blocked P. acnes‐induced production of caspase‐1(p10) and IL‐1β in primary mouse macrophages and human SZ95 sebocytes, indicating the suppression of NLRP3 inflammasome. Licochalcone A suppressed P. acnes‐induced ASC speck formation and mitochondrial reactive oxygen species. Topical application of licochalcone A to mouse ear skin attenuated P. acnes‐induced skin inflammation as shown by histological assessment, ear thickness measurement, and inflammatory gene expression. Licochalcone A reduced caspase‐1 activity and IL‐1β production in mouse ear injected with P. acnes. This study demonstrated that licochalcone A is effective in the control of P. acnes‐induced skin inflammation as an efficient inhibitor for NLRP3 inflammasome. Our study provides a new paradigm for the development of anti‐acne therapy via targeting NLRP3 inflammasome.

Abstract A26: The metastatic potential of breast cancer cells was suppressed by NLRP3 inflammasome inhibition in macrophages

Tumor microenvironment favors tumor cells to promote tumor growth, migration, invasion, and angiogenesis. IL-1beta, one of the inflammatory cytokines associated with tumor microenvironment, plays an important role in the development and progress of tumor. The activation of inflammasome is critical to secrete mature IL-1beta through stepwise reactions to activate capspase-1. Therefore, we investigated whether the metastatic potential of breast cancer cells was regulated by NLRP3 inflammasome activation of macrophages in tumor microenvironment. The activation of NLRP3 inflammasome was induced by ATP in bone marrow-derived primary mouse macrophages. The metastatic potential of 4T1 breast cancer cells was determined by migration and invasion assays with transwell system. Conditioned media from ATP-stimulated wild-type macrophages increased the migration and invasion of breast cancer cells (4T1) while conditioned media from NLRP3 knockout macrophages stimulated with ATP failed to induce the migration and invasion of breast cancer cells. Celastrol, an inhibitor of NLRP3 inflammasome, reduced the potency of macrophages to stimulate the migration and invasion of 4T1 breast cancer cells. The suppression of NLRP3 inflammasome by celastrol was mediated by the blockade of potassium efflux. The results demonstrate that the activation of NLRP3 inflammasome in macrophages leads to the enhancement of metastatic potential of breast cancer cells. The results would provide a novel anticancer strategy to modulate tumor microenvironment by suppressing NLRP3 inflammasome and consequently reducing IL-1beta production. (This study was supported by the National Research Foundation of Korea (NRF) grants funded by the Korea government (Bio & Medical Technology Development Program, NRF-2017M3A9F5028608; Basic Research Laboratory Program, NRF-2017R1A4A1015036.) Citation Format: Jin Y. Lee, Hye E. Lee, Joo Y. Lee. The metastatic potential of breast cancer cells was suppressed by NLRP3 inflammasome inhibition in macrophages [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr A26.

Phloxine O, a Cosmetic Colorant, Suppresses the Expression of Thymic Stromal Lymphopoietin and Acute Dermatitis Symptoms in Mice

Cosmetics are primarily applied to the skin; therefore, the association of cosmetic dyes with skin diseases or inflammation is a topic of great interest. Thymic stromal lymphopoietin (TSLP) is an interleukin 7-like cytokine that activates dendritic cells to promote Th2 inflammatory immune responses. TSLP is highly expressed in keratinocytes under inflammatory conditions, which suggests that it may play a critical role in the development of skin diseases, such as atopic dermatitis. Therefore, we investigated whether cosmetic dyes influenced the production of TSLP by keratinocytes. Phloxine O, also known as D&C Red No.27, is one of the most common red synthetic pigments and is widely used in colored cosmetics. Our results showed that Phloxine O downregulated phorbol 12-myristate 13-acetate-induced production of TSLP in a murine keratinocyte cell line (PAM212). Phloxine O also suppressed TSLP expression in KCMH-1 cells, which are mouse keratinocytes that constitutively produce high levels of TSLP. To investigate the in vivo effects of Phloxine O, we induced TSLP expression in mouse ear skin by topically applying MC903, a vitamin D3 analogue that is a well-known inducer of atopic dermatitis-like symptoms. Topical application of Phloxine O prevented MC903-induced TSLP production in mouse ear skin, attenuated the acute dermatitis-like symptoms and decreased serum IgE and histamine levels in mice. Suppression of TSLP expression by Phloxine O correlated with reduced expression of OX40 ligand and Th2 cytokines in mouse ear skin. Our results showed that Phloxine O may be beneficial to prevent dermatitis by suppressing the expression of TSLP and Th2 cytokines in skin.