Publications by authors named "John A Cidlowski"

150 Publications

Glucocorticoids as Regulators of Macrophage-Mediated Tissue Homeostasis.

Front Immunol 2021 17;12:669891. Epub 2021 May 17.

Molecular Endocrinology Group, Signal Transduction Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC, United States.

Our immune system has evolved as a complex network of cells and tissues tasked with maintaining host homeostasis. This is evident during the inflammatory responses elicited during a microbial infection or traumatic tissue damage. These responses seek to eliminate foreign material or restore tissue integrity. Even during periods without explicit disturbances, the immune system plays prominent roles in tissue homeostasis. Perhaps one of the most studied cells in this regard is the macrophage. Tissue-resident macrophages are a heterogenous group of sensory cells that respond to a variety of environmental cues and are essential for organ function. Endogenously produced glucocorticoid hormones connect external environmental stress signals with the function of many cell types, producing profound changes in immune cells, including macrophages. Here, we review the current literature which demonstrates specific effects of glucocorticoids in several organ systems. We propose that tissue-resident macrophages, through glucocorticoid signaling, may play an underappreciated role as regulators of organ homeostasis.
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http://dx.doi.org/10.3389/fimmu.2021.669891DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8165320PMC
May 2021

Glucocorticoids and Androgens Protect From Gastric Metaplasia by Suppressing Group 2 Innate Lymphoid Cell Activation.

Gastroenterology 2021 May 7. Epub 2021 May 7.

Molecular Endocrinology Group, Signal Transduction Laboratory, North Carolina. Electronic address:

Background & Aims: The immune compartment is critical for maintaining tissue homeostasis. A weak immune response increases susceptibility to infection, but immune hyperactivation causes tissue damage, and chronic inflammation may lead to cancer development. In the stomach, inflammation damages the gastric glands and drives the development of potentially preneoplastic metaplasia. Glucocorticoids are potent anti-inflammatory steroid hormones that are required to suppress gastric inflammation and metaplasia. However, these hormones function differently in males and females. Here, we investigate the impact of sex on the regulation of gastric inflammation.

Methods: Endogenous glucocorticoids and male sex hormones were removed from mice using adrenalectomy and castration, respectively. Mice were treated with 5α-dihydrotestosterone (DHT) to test the effects of androgens on regulating gastric inflammation. Single-cell RNA sequencing of gastric leukocytes was used to identify the leukocyte populations that were the direct targets of androgen signaling. Type 2 innate lymphoid cells (ILC2s) were depleted by treatment with CD90.2 antibodies.

Results: We show that adrenalectomized female mice develop spontaneous gastric inflammation and spasmolytic polypeptide-expressing metaplasia (SPEM) but that the stomachs of adrenalectomized male mice remain quantitatively normal. Simultaneous depletion of glucocorticoids and sex hormones abolished the male-protective effects and triggered spontaneous pathogenic gastric inflammation and SPEM. Treatment of female mice with DHT prevented gastric inflammation and SPEM development when administered concurrent with adrenalectomy and also reversed the pathology when administered after disease onset. Single-cell RNAseq of gastric leukocytes revealed that ILC2s expressed abundant levels of both the glucocorticoid receptor (Gr) and androgen receptor (Ar). We demonstrated that DHT treatment potently suppressed the expression of the proinflammatory cytokines Il13 and Csf2 by ILC2s. Moreover, ILC2 depletion protected the stomach from SPEM development.

Conclusions: Here, we report a novel mechanism by which glucocorticoids and androgens exert overlapping effects to regulate gastric inflammation. Androgen signaling within ILC2s prevents their pathogenic activation by suppressing the transcription of proinflammatory cytokines. This work revealed a critical role for sex hormones in regulating gastric inflammation and metaplasia.
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http://dx.doi.org/10.1053/j.gastro.2021.04.075DOI Listing
May 2021

Deletion of hippocampal Glucocorticoid receptors unveils sex-biased microRNA expression and neuronal morphology alterations in mice.

Neurobiol Stress 2021 May 9;14:100306. Epub 2021 Feb 9.

Laboratory of Neuroplasticity and Neurogenetics, Faculty of Chemical and Pharmaceutical Sciences, Department of Biochemistry and Molecular Biology, Universidad de Chile, Independencia, 8380492, Santiago, Chile.

Sex differences in the brain have prompted many researchers to investigate the underlying molecular actors, such as the glucocorticoid receptor (GR). This nuclear receptor controls gene expression, including microRNAs (miRNAs), in non-neuronal cells. Here, we investigated sex-biased effects of GR on hippocampal miRNA expression and neuronal morphology by generating a neuron-specific GR knockout mouse ( ). The levels of 578 mature miRNAs were assessed using NanoString technology and, in contrast to males, female mice showed a substantially higher number of differentially expressed miRNAs, confirming a sex-biased effect of GR ablation. Based on bioinformatic analyses we identified several transcription factors potentially involved in miRNA regulation. Functional enrichment analyses of the miRNA-mRNA interactions revealed pathways related to neuronal arborization and both spine morphology and density in both sexes. Two recognized regulators of dendritic morphology, CAMKII-α and GSK-3β, increased their protein levels by GR ablation in female mice hippocampus, without changes in males. Additionally, sex-specific effects of GR deletion were observed on CA1 neuronal arborization and dendritic spine features. For instance, a reduced density of mushroom spines in apical dendrites was evidenced only in females, while a decreased length in basal dendrites was noted only in males. However, length and arborization of apical dendrites were reduced by GR ablation irrespective of the sex. Overall, our study provides new insights into the sex-biased GR actions, especially in terms of miRNAs expression and neuronal morphology in the hippocampus.
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http://dx.doi.org/10.1016/j.ynstr.2021.100306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7906897PMC
May 2021

Ions, the Movement of Water and the Apoptotic Volume Decrease.

Front Cell Dev Biol 2020 25;8:611211. Epub 2020 Nov 25.

Signal Transduction Laboratory, Department of Health and Human Services, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC, United States.

The movement of water across the cell membrane is a natural biological process that occurs during growth, cell division, and cell death. Many cells are known to regulate changes in their cell volume through inherent compensatory regulatory mechanisms. Cells can sense an increase or decrease in their cell volume, and compensate through mechanisms known as a regulatory volume increase (RVI) or decrease (RVD) response, respectively. The transport of sodium, potassium along with other ions and osmolytes allows the movement of water in and out of the cell. These compensatory volume regulatory mechanisms maintain a cell at near constant volume. A hallmark of the physiological cell death process known as apoptosis is the loss of cell volume or cell shrinkage. This loss of cell volume is in stark contrast to what occurs during the accidental cell death process known as necrosis. During necrosis, cells swell or gain water, eventually resulting in cell lysis. Thus, whether a cell gains or loses water after injury is a defining feature of the specific mode of cell death. Cell shrinkage or the loss of cell volume during apoptosis has been termed apoptotic volume decrease or AVD. Over the years, this distinguishing feature of apoptosis has been largely ignored and thought to be a passive occurrence or simply a consequence of the cell death process. However, studies on AVD have defined an underlying movement of ions that result in not only the loss of cell volume, but also the activation and execution of the apoptotic process. This review explores the role ions play in controlling not only the movement of water, but the regulation of apoptosis. We will focus on what is known about specific ion channels and transporters identified to be involved in AVD, and how the movement of ions and water change the intracellular environment leading to stages of cell shrinkage and associated apoptotic characteristics. Finally, we will discuss these concepts as they apply to different cell types such as neurons, cardiomyocytes, and corneal epithelial cells.
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http://dx.doi.org/10.3389/fcell.2020.611211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7723978PMC
November 2020

After 62 years of regulating immunity, dexamethasone meets COVID-19.

Nat Rev Immunol 2020 10;20(10):587-588

The Signal Transduction Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, NC, USA.

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http://dx.doi.org/10.1038/s41577-020-00421-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7416654PMC
October 2020

Protein phosphatase 1 alpha enhances glucocorticoid receptor activity by a mechanism involving phosphorylation of serine-211.

Mol Cell Endocrinol 2020 12 22;518:110873. Epub 2020 Jun 22.

Swiss Centre for Applied Human Toxicology (SCAHT), University of Basel, Missionsstrasse 64, 4055, Basel, Switzerland; Division of Molecular and Systems Toxicology, Department of Pharmaceutical Sciences, University of Basel, Klingelbergstrasse 50, 4056, Basel, Switzerland. Electronic address:

By acting as a ligand-dependent transcription factor the glucocorticoid receptor (GR) mediates the actions of glucocorticoids and regulates many physiological processes. An impaired regulation of glucocorticoid action has been associated with numerous disorders. Thus, the elucidation of underlying signaling pathways is essential to understand mechanisms of disrupted glucocorticoid function and contribution to diseases. This study found increased GR transcriptional activity upon overexpression of protein phosphatase 1 alpha (PP1α) in HEK-293 cells and decreased expression levels of GR-responsive genes following PP1α knockdown in the endogenous A549 cell model. Mechanistic investigations revealed reduced phosphorylation of GR-Ser211 following PP1α silencing and provided a first indication for an involvement of glycogen synthase kinase 3 (GSK-3). Thus, the present study identified PP1α as a novel post-translational activator of GR signaling, suggesting that disruption of PP1α function could lead to impaired glucocorticoid action and thereby contribute to diseases.
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http://dx.doi.org/10.1016/j.mce.2020.110873DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7606615PMC
December 2020

Murine Glucocorticoid Receptors Orchestrate B Cell Migration Selectively between Bone Marrow and Blood.

J Immunol 2020 08 22;205(3):619-629. Epub 2020 Jun 22.

Signal Transduction Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, U.S. Department of Health and Human Services, Research Triangle Park, NC 27709

Glucocorticoids promote CXCR4 expression by T cells, monocytes, macrophages, and eosinophils, but it is not known if glucocorticoids regulate CXCR4 in B cells. Considering the important contributions of CXCR4 to B cell development and function, we investigated the glucocorticoid/CXCR4 axis in mice. We demonstrate that glucocorticoids upregulate CXCR4 mRNA and protein in mouse B cells. Using a novel strain of mice lacking glucocorticoid receptors (GRs) specifically in B cells, we show that reduced CXCR4 expression associated with GR deficiency results in impaired homing of mature B cells to bone marrow, whereas migration to other lymphoid tissues is independent of B cell GRs. The exchange of mature B cells between blood and bone marrow is sensitive to small, physiologic changes in glucocorticoid activity, as evidenced by the lack of circadian rhythmicity in GR-deficient B cell counts normally associated with diurnal patterns of glucocorticoid secretion. B cell mice mounted normal humoral responses to immunizations with T-dependent and T-independent (Type 1) Ags, but Ab responses to a multivalent T-independent (Type 2) Ag were impaired, a surprise finding considering the immunosuppressive properties commonly attributed to glucocorticoids. We propose that endogenous glucocorticoids regulate a dynamic mode of B cell migration specialized for rapid exchange between bone marrow and blood, perhaps as a means to optimize humoral immunity during diurnal periods of activity.
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http://dx.doi.org/10.4049/jimmunol.1901135DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7369236PMC
August 2020

Coordinate expression loss of and in gastric cancer via impairment of a glucocorticoid-responsive enhancer.

Am J Physiol Gastrointest Liver Physiol 2020 08 15;319(2):G175-G188. Epub 2020 Jun 15.

Murdoch Children's Research Institute, Melbourne, Victoria, Australia.

Gastrokines (GKNs) are anti-inflammatory proteins secreted by gastric epithelial (surface mucous and pit) cells, with their aberrant loss of expression causally linked to premalignant inflammation and gastric cancer (GC). Transcriptional mechanisms accounting for GKN expression loss have not been elucidated. Using human clinical cohorts, mouse transgenics, bioinformatics, and transfection/reporter assays, we report a novel mechanism of GKN gene transcriptional regulation and its impairment in GC. / loss is highly coordinated, with both genes showing parallel downregulation during human and mouse GC development, suggesting joint transcriptional control. In BAC transgenic studies, we defined a 152-kb genomic region surrounding the human / genes sufficient to direct their tissue- and lineage-restricted expression. A screen of the 152-kb region for candidate regulatory elements identified a DNase I hypersensitive site (CR2) located 4 kb upstream of the gene. CR2 showed overlapping enrichment of enhancer-related histone marks (H3K27Ac), a consensus binding site (GRE) for the glucocorticoid receptor (GR), strong GR occupancy in ChIP-seq data sets and, critically, exhibited dexamethasone-sensitive enhancer activity in reporter assays. Strikingly, GR showed progressive expression loss, paralleling that of /, in human and mouse GC, suggesting desensitized glucocorticoid signaling as a mechanism underlying GKN loss. Finally, mouse adrenalectomy studies revealed a critical role for endogenous glucocorticoids in sustaining correct expression (and anti-inflammatory restraint) of GKNs in vivo. Together, these data link the coordinate expression of GKNs to a glucocorticoid-responsive and likely shared transcriptional enhancer mechanism, with its compromised activation contributing to dual GKN loss during GC progression. Gastrokine 2 (GKN2) is an anti-inflammatory protein produced by the gastric epithelium. GKN2 expression is progressively lost during gastric cancer (GC), which is believed to play a casual role in GC development. Here, we use bacterial artificial chromosome transgenic studies to identify a glucocorticoid-responsive enhancer element that likely governs expression of , which, via parallel expression loss of the anti-inflammatory glucocorticoid receptor, reveals a novel mechanism to explain the loss of GKN2 during GC pathogenesis.
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http://dx.doi.org/10.1152/ajpgi.00019.2020DOI Listing
August 2020

Glucocorticoid Signaling and the Aging Heart.

Front Endocrinol (Lausanne) 2020 27;11:347. Epub 2020 May 27.

Signal Transduction Laboratory, Department of Health and Human Services, National Institute of Environmental Health Sciences, National Institutes of Health, Durham, NC, United States.

A decline in normal physiological functions characterizes the aging process. While some of these changes are benign, the decrease in the function of the cardiovascular system that occurs during aging leads to the activation of pathological processes associated with an increased risk for heart disease and its complications. Imbalances in endocrine function are also common occurrences during the aging process. Glucocorticoids are primary stress hormones and are critical regulators of energy metabolism, inflammation, and cardiac function. Glucocorticoids exert their actions by binding the glucocorticoid receptor (GR) and, in some instances, to the mineralocorticoid receptor (MR). GR and MR are members of the nuclear receptor family of ligand-activated transcription factors. There is strong evidence that imbalances in GR and MR signaling in the heart have a causal role in cardiac disease. The extent to which glucocorticoids play a role in the aging heart, however, remains unclear. This review will summarize the positive and negative direct and indirect effects of glucocorticoids on the heart and the latest molecular and physiological evidence on how alterations in glucocorticoid signaling lead to changes in cardiac structure and function. We also briefly discuss the effects of other hormones systems such as estrogens and GH/IGF-1 on different cardiovascular cells during aging. We will also review the link between imbalances in glucocorticoid levels and the molecular processes responsible for promoting cardiomyocyte dysfunction in aging. Finally, we will discuss the potential for selectively manipulating glucocorticoid signaling in cardiomyocytes, which may represent an improved therapeutic approach for preventing and treating age-related heart disease.
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http://dx.doi.org/10.3389/fendo.2020.00347DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7266971PMC
May 2021

Glucocorticoid receptors are required effectors of TGFβ1-induced p38 MAPK signaling to advanced cancer phenotypes in triple-negative breast cancer.

Breast Cancer Res 2020 05 1;22(1):39. Epub 2020 May 1.

Departments of Medicine (Division of Hematology, Oncology, and Transplantation) and Pharmacology, University of Minnesota Masonic Cancer Center, Delivery Code 2812 Cancer and Cardiovascular Research Building; Suite 3-126 2231 6th St SE, Minneapolis, MN, 55455, USA.

Background: Altered signaling pathways typify breast cancer and serve as direct inputs to steroid hormone receptor sensors. We previously reported that phospho-Ser134-GR (pS134-GR) species are elevated in triple-negative breast cancer (TNBC) and cooperate with hypoxia-inducible factors, providing a novel avenue for activation of GR in response to local or cellular stress.

Methods: We probed GR regulation by factors (cytokines, growth factors) that are rich within the tumor microenvironment (TME). TNBC cells harboring endogenous wild-type (wt) or S134A-GR species were created by CRISPR/Cas knock-in and subjected to transwell migration, invasion, soft-agar colony formation, and tumorsphere assays. RNA-seq was employed to identify pS134-GR target genes that are regulated both basally (intrinsic) or by TGFβ1 in the absence of exogenously added GR ligands. Regulation of selected basal and TGFβ1-induced pS134-GR target genes was validated by qRT-PCR and chromatin immunoprecipitation assays. Bioinformatics tools were used to probe public data sets for expression of pS134-GR 24-gene signatures.

Results: In the absence of GR ligands, GR is transcriptionally activated via p38-dependent phosphorylation of Ser134 as a mechanism of homeostatic stress-sensing and regulated upon exposure of TNBC cells to TME-derived agents. The ligand-independent pS134-GR transcriptome encompasses TGFβ1 and MAPK signaling gene sets associated with TNBC cell survival and migration/invasion. Accordingly, pS134-GR was essential for TNBC cell anchorage-independent growth in soft-agar, migration, invasion, and tumorsphere formation, an in vitro readout of cancer stemness properties. Both pS134-GR and expression of the MAPK-scaffolding molecule 14-3-3ζ were essential for a functionally intact p38 MAPK signaling pathway downstream of MAP3K5/ASK1, indicative of a feedforward signaling loop wherein self-perpetuated GR phosphorylation enables cancer cell autonomy. A 24-gene pS134-GR-dependent signature induced by TGFβ1 predicts shortened overall survival in breast cancer patients.

Conclusions: Phospho-S134-GR is a critical downstream effector of p38 MAPK signaling and TNBC migration/invasion, survival, and stemness properties. Our studies define a ligand-independent role for GR as a homeostatic "sensor" of intrinsic stimuli as well as extrinsic factors rich within the TME (TGFβ1) that enable potent activation of the p38 MAPK stress-sensing pathway and nominate pS134-GR as a therapeutic target in aggressive TNBC.
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http://dx.doi.org/10.1186/s13058-020-01277-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7193415PMC
May 2020

Glucocorticoids mobilize macrophages by transcriptionally up-regulating the exopeptidase DPP4.

J Biol Chem 2020 03 27;295(10):3213-3227. Epub 2020 Jan 27.

Molecular Endocrinology Group, Signal Transduction Laboratory, NIEHS, National Institutes of Health, Research Triangle Park, North Carolina 27709. Electronic address:

Glucocorticoids are potent endogenous anti-inflammatory molecules, and their cognate receptor, glucocorticoid receptor (GR), is expressed in nearly all immune cells. Macrophages are heterogeneous immune cells having a central role in both tissue homeostasis and inflammation and also play a role in the pathogenesis of some inflammatory diseases. Paradoxically, glucocorticoids have only a limited efficacy in controlling the resolution of these macrophage-related diseases. Here, we report that the transcriptomes of monocyte-like THP-1 cells and macrophage-like THP-1 cells (THP1-MΦ) have largely conserved gene expression patterns. In contrast, the differentiation to THP1-MΦ significantly altered the sensitivity of gene transcription to glucocorticoids. Among glucocorticoid-regulated genes, we identified the exopeptidase dipeptidyl peptidase-4 () as a critical glucocorticoid-responsive gene in THP1-MΦ. We found that GR directly induces DPP4 gene expression by binding to two glucocorticoid-responsive elements (GREs) within the promoter. Additionally, we show that glucocorticoid-induced expression is blocked by the GR antagonist RU-486 and by GR siRNA transfection and that DPP4 enzyme activity is reduced by DPP4 inhibitors. Of note, glucocorticoids highly stimulated macrophage mobility; unexpectedly, DPP4 mediated the glucocorticoid-induced macrophage migration, and siRNA-mediated knockdowns of GR and DPP4 blocked dexamethasone-induced THP1-MΦ migration. Moreover, glucocorticoid-induced activation was also observed in proinflammatory M1-polarized murine macrophages, as well as peritoneal macrophages, and was associated with increased macrophage migration. Our results indicate that glucocorticoids directly up-regulate DPP4 expression and thereby induce migration in macrophages, potentially explaining why glucocorticoid therapy is less effective in controlling macrophage-dominated inflammatory disorders.
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http://dx.doi.org/10.1074/jbc.RA119.010894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7062181PMC
March 2020

Inhibition of miR-378a-3p by Inflammation Enhances IL-33 Levels: A Novel Mechanism of Alarmin Modulation in Ulcerative Colitis.

Front Immunol 2019 20;10:2449. Epub 2019 Nov 20.

Innate Immunity Laboratory, Immunology Program, Faculty of Medicine, Biomedical Sciences Institute, Universidad de Chile, Santiago, Chile.

Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) characterized by mucosa damage associated with an uncontrolled inflammatory response. This immunological impairment leads to altered inflammatory mediators such as IL-33, which is shown to increase in the mucosa of active UC (aUC) patients. MicroRNAs present a distorted feature in inflamed colonic mucosa and are potential IL-33 regulating candidates in UC. Therefore, we studied the microRNA and mRNA profiles in inflamed colonic samples of UC patients, evaluating the effect of a microRNA (selected by analysis and its expression in UC patients), on IL-33 under inflammatory conditions. We found that inflamed mucosa ( = 8) showed increased expression of 40 microRNAs and 2,120 mRNAs, while 49 microRNAs and 1,734 mRNAs were decreased, as determined by microarrays. In particular, IL-33 mRNA showed a 3.8-fold increase and eight members of a microRNA family (miR-378), which targets IL-33 mRNA in the 3'UTR, were decreased (-3.9 to -3.0 times). We selected three members of the miR-378 family (miR-378a-3p, miR-422a, and miR-378c) according to background information and interaction energy analysis, for further correlation analyses with IL-33 expression through qPCR and ELISA, respectively. We determined that aUC ( = 24) showed high IL-33 levels, and decreased expression of miR-378a-3p and miR-422a compared to inactive UC ( = 10) and controls ( = 6). Moreover, both microRNAs were inversely correlated with IL-33 expression, while miR-378c does not show a significant difference. To evaluate the effect of TNFα on the studied microRNAs, aUC patients with anti-TNF therapy were compared to aUC receiving other treatments. The levels of miR-378a-3p and miR-378c were higher in aUC patients with anti-TNF. Based on these findings, we selected miR-378a-3p to exploring the molecular mechanism involved by assays, showing that over-expression of miR-378a-3p decreased the levels of an IL-33 target sequence β-gal-reporter gene in HEK293 cells. Stable miR-378a-3p over-expression/inhibition inversely modulated IL-33 content and altered viability of HT-29 cells. Additionally, in an inflammatory context, TNFα decreased miR-378a-3p levels in HT-29 cells enhancing IL-33 expression. Together, our results propose a regulatory mechanism of IL-33 expression exerted by miR-378a-3p in an inflammatory environment, contributing to the understanding of UC pathogenesis.
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http://dx.doi.org/10.3389/fimmu.2019.02449DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6879552PMC
November 2020

THE CONCISE GUIDE TO PHARMACOLOGY 2019/20: Introduction and Other Protein Targets.

Br J Pharmacol 2019 12;176 Suppl 1:S1-S20

Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh, EH8 9XD, UK.

The Concise Guide to PHARMACOLOGY 2019/20 is the fourth in this series of biennial publications. The Concise Guide provides concise overviews of the key properties of nearly 1800 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide represents approximately 400 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.14747. In addition to this overview, in which are identified Other protein targets which fall outside of the subsequent categorisation, there are six areas of focus: G protein-coupled receptors, ion channels, nuclear hormone receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2019, and supersedes data presented in the 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the International Union of Basic and Clinical Pharmacology Committee on Receptor Nomenclature and Drug Classification (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.
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http://dx.doi.org/10.1111/bph.14747DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6844537PMC
December 2019

THE CONCISE GUIDE TO PHARMACOLOGY 2019/20: Nuclear hormone receptors.

Br J Pharmacol 2019 12;176 Suppl 1:S229-S246

Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh, EH8 9XD, UK.

The Concise Guide to PHARMACOLOGY 2019/20 is the fourth in this series of biennial publications. The Concise Guide provides concise overviews of the key properties of nearly 1800 human drug targets with an emphasis on selective pharmacology (where available), plus links to the open access knowledgebase source of drug targets and their ligands (www.guidetopharmacology.org), which provides more detailed views of target and ligand properties. Although the Concise Guide represents approximately 400 pages, the material presented is substantially reduced compared to information and links presented on the website. It provides a permanent, citable, point-in-time record that will survive database updates. The full contents of this section can be found at http://onlinelibrary.wiley.com/doi/10.1111/bph.14750. Nuclear hormone receptors are one of the six major pharmacological targets into which the Guide is divided, with the others being: G protein-coupled receptors, catalytic receptors, enzymes and transporters. These are presented with nomenclature guidance and summary information on the best available pharmacological tools, alongside key references and suggestions for further reading. The landscape format of the Concise Guide is designed to facilitate comparison of related targets from material contemporary to mid-2019, and supersedes data presented in the 2017/18, 2015/16 and 2013/14 Concise Guides and previous Guides to Receptors and Channels. It is produced in close conjunction with the International Union of Basic and Clinical Pharmacology Committee on Receptor Nomenclature and Drug Classification (NC-IUPHAR), therefore, providing official IUPHAR classification and nomenclature for human drug targets, where appropriate.
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http://dx.doi.org/10.1111/bph.14750DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6844575PMC
December 2019

Glucocorticoids preserve the t-tubular system in ventricular cardiomyocytes by upregulation of autophagic flux.

Basic Res Cardiol 2019 10 31;114(6):47. Epub 2019 Oct 31.

Institute of Cellular and Molecular Physiology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Waldstr. 6, 91054, Erlangen, Germany.

A major contributor to contractile dysfunction in heart failure is remodelling and loss of the cardiomyocyte transverse tubular system (t-system), but underlying mechanisms and signalling pathways remain elusive. It has been shown that dexamethasone promotes t-tubule development in stem cell-derived cardiomyocytes and that cardiomyocyte-specific glucocorticoid receptor (GR) knockout (GRKO) leads to heart failure. Here, we studied if the t-system is altered in GRKO hearts and if GR signalling is required for t-system preservation in adult cardiomyocytes. Confocal and 3D STED microscopy of myocardium from cardiomyocyte-specific GRKO mice revealed decreased t-system density and increased distances between ryanodine receptors (RyR) and L-type Ca channels (LTCC). Because t-system remodelling and heart failure are intertwined, we investigated the underlying mechanisms in vitro. Ventricular cardiomyocytes from failing human and healthy adult rat hearts cultured in the absence of glucocorticoids (CTRL) showed distinctively lower t-system density than cells treated with dexamethasone (EC 1.1 nM) or corticosterone. The GR antagonist mifepristone abrogated the effect of dexamethasone. Dexamethasone improved RyR-LTCC coupling and synchrony of intracellular Ca release, but did not alter expression levels of t-system-associated proteins junctophilin-2 (JPH2), bridging integrator-1 (BIN1) or caveolin-3 (CAV3). Rather, dexamethasone upregulated LC3B and increased autophagic flux. The broad-spectrum protein kinase inhibitor staurosporine prevented dexamethasone-induced upregulation of autophagy and t-system preservation, and autophagy inhibitors bafilomycin A and chloroquine accelerated t-system loss. Conversely, induction of autophagy by rapamycin or amino acid starvation preserved the t-system. These findings suggest that GR signalling and autophagy are critically involved in t-system preservation and remodelling in the heart.
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http://dx.doi.org/10.1007/s00395-019-0758-6DOI Listing
October 2019

Deletion of the Cardiomyocyte Glucocorticoid Receptor Leads to Sexually Dimorphic Changes in Cardiac Gene Expression and Progression to Heart Failure.

J Am Heart Assoc 2019 08 17;8(15):e011012. Epub 2019 Jul 17.

Signal Transduction Laboratory National Institute of Environmental Health Sciences National Institutes of Health Department of Health and Human Services Research Triangle Park NC.

Background The contribution of glucocorticoids to sexual dimorphism in the heart is essentially unknown. Therefore, we sought to determine the sexually dimorphic actions of glucocorticoid signaling in cardiac function and gene expression. To accomplish this goal, we conducted studies on mice lacking glucocorticoid receptors (GR) in cardiomyocytes (cardioGRKO mouse model). Methods and Results Deletion of cardiomyocyte GR leads to an increase in mortality because of the development of spontaneous cardiac pathology in both male and female mice; however, females are more resistant to GR signaling inactivation in the heart. Male cardioGRKO mice had a median survival age of 6 months. In contrast, females had a median survival age of 10 months. Transthoracic echocardiography data showed phenotypic differences between male and female cardioGRKO hearts. By 3 months of age, male cardioGRKO mice exhibited left ventricular systolic dysfunction. Conversely, no significant functional deficits were observed in female cardioGRKO mice at the same time point. Functional sensitivity of male hearts to the loss of cardiomyocyte GR was reversed following gonadectomy. RNA-Seq analysis showed that deleting GR in the male hearts leads to a more profound dysregulation in the expression of genes implicated in heart rate regulation (calcium handling). In agreement with these gene expression data, cardiomyocytes isolated from male cardioGRKO hearts displayed altered intracellular calcium responses. In contrast, female GR-deficient cardiomyocytes presented a response comparable with controls. Conclusions These data suggest that GR regulates calcium responses in a sex-biased manner, leading to sexually distinct responses to stress in male and female mice hearts, which may contribute to sex differences in heart disease, including the development of ventricular arrhythmias that contribute to heart failure and sudden death.
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http://dx.doi.org/10.1161/JAHA.118.011012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6761632PMC
August 2019

β-Arrestin-1 inhibits glucocorticoid receptor turnover and alters glucocorticoid signaling.

J Biol Chem 2019 07 5;294(29):11225-11239. Epub 2019 Jun 5.

Signal Transduction Laboratory, NIEHS, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina 27709

Glucocorticoids are among the most widely used drugs to treat many autoimmune and inflammatory diseases. Although much research has been focused on investigating glucocorticoid activity, it remains unclear how glucocorticoids regulate distinct processes in different cells. Glucocorticoids exert their effects through the glucocorticoid receptor (GR), which, upon glucocorticoid binding, interacts with regulatory proteins, affecting its activity and function. These protein-protein interactions are necessary for the resolution of glucocorticoid-dependent physiological and pharmacological processes. In this study, we discovered a novel protein interaction between the glucocorticoid receptor and β-arrestin-1, a scaffold protein with a well-established role in G protein-coupled receptor signaling. Using co-immunoprecipitation and proximity ligation assays in A549 cells, we observed that β-arrestin-1 and unliganded GR interact in the cytoplasm and that, following glucocorticoid binding, the protein complex is found in the nucleus. We show that siRNA-mediated β-arrestin-1 knockdown alters GR protein turnover by up-regulating the E3 ubiquitin ligase Pellino-1, which catalyzes GR ubiquitination and thereby marks the receptor for proteasomal degradation. The enhanced GR turnover observed in β-arrestin-1-deficient cells limits the duration of the glucocorticoid response on GR target genes. These results demonstrate that β-arrestin-1 is a crucial player for the stability of the glucocorticoid receptor. The GR/β-arrestin-1 interaction uncovered here may help unravel mechanisms that contribute to the cell type-specific activities of glucocorticoids.
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http://dx.doi.org/10.1074/jbc.RA118.007150DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6643030PMC
July 2019

Cardiomyocyte glucocorticoid and mineralocorticoid receptors directly and antagonistically regulate heart disease in mice.

Sci Signal 2019 04 16;12(577). Epub 2019 Apr 16.

Signal Transduction Laboratory, NIEHS, NIH, DHHS, 111 T.W. Alexander Drive, Research Triangle Park, NC 27709, USA.

Stress is increasingly associated with heart dysfunction and is linked to higher mortality rates in patients with cardiometabolic disease. Glucocorticoids are primary stress hormones that regulate homeostasis through two nuclear receptors, the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR), both of which are present in cardiomyocytes. To examine the specific and coordinated roles that these receptors play in mediating the direct effects of stress on the heart, we generated mice with cardiomyocyte-specific deletion of GR (cardioGRKO), MR (cardioMRKO), or both GR and MR (cardioGRMRdKO). The cardioGRKO mice spontaneously developed cardiac hypertrophy and left ventricular systolic dysfunction and died prematurely from heart failure. In contrast, the cardioMRKO mice exhibited normal heart morphology and function. Despite the presence of myocardial stress, the cardioGRMRdKO mice were resistant to the cardiac remodeling, left ventricular dysfunction, and early death observed in the cardioGRKO mice. Gene expression analysis revealed the loss of gene changes associated with impaired Ca handling, increased oxidative stress, and enhanced cell death and the presence of gene changes that limited the hypertrophic response and promoted cardiomyocyte survival in the double knockout hearts. Reexpression of MR in cardioGRMRdKO hearts reversed many of the cardioprotective gene changes and resulted in cardiac failure. These findings reveal a critical role for balanced cardiomyocyte GR and MR stress signaling in cardiovascular health. Therapies that shift stress signaling in the heart to favor more GR and less MR activity may provide an improved approach for treating heart disease.
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http://dx.doi.org/10.1126/scisignal.aau9685DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7082727PMC
April 2019

Silencing of maternal hepatic glucocorticoid receptor is essential for normal fetal development in mice.

Commun Biol 2019 15;2:104. Epub 2019 Mar 15.

Signal Transduction Laboratory, Research Triangle Park, North Carolina USA.

Excessive or chronic stress can lead to a variety of diseases due to aberrant activation of the glucocorticoid receptor (GR), a ligand activated transcription factor. Pregnancy represents a particular window of sensitivity in which excessive stress can have adverse outcomes, particularly on the developing fetus. Here we show maternal hepatic stress hormone responsiveness is diminished via epigenetic silencing of the glucocorticoid receptor during pregnancy. Provocatively, reinstallation of GR to hepatocytes during pregnancy by adeno-associated viral transduction dysregulates genes involved in proliferation, resulting in impaired pregnancy-induced hepatomegaly. Disruption of the maternal hepatic adaptation to pregnancy results in in utero growth restriction (IUGR). These data demonstrate pregnancy antagonizes the liver-specific effects of stress hormone signaling in the maternal compartment to ultimately support the healthy development of embryos.
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http://dx.doi.org/10.1038/s42003-019-0344-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6420645PMC
April 2020

Endogenous glucocorticoids prevent gastric metaplasia by suppressing spontaneous inflammation.

J Clin Invest 2019 03 18;129(3):1345-1358. Epub 2019 Feb 18.

Molecular Endocrinology Group, Signal Transduction Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA.

In the stomach, chronic inflammation causes metaplasia and creates a favorable environment for the evolution of gastric cancer. Glucocorticoids are steroid hormones that repress proinflammatory stimuli, but their role in the stomach is unknown. In this study, we show that endogenous glucocorticoids are required to maintain gastric homeostasis. Removal of circulating glucocorticoids in mice by adrenalectomy resulted in the rapid onset of spontaneous gastric inflammation, oxyntic atrophy, and spasmolytic polypeptide-expressing metaplasia (SPEM), a putative precursor of gastric cancer. SPEM and oxyntic atrophy occurred independently of lymphocytes. However, depletion of monocytes and macrophages by clodronate treatment or inhibition of gastric monocyte infiltration using the Cx3cr1 knockout mouse model prevented SPEM development. Our results highlight the requirement for endogenous glucocorticoid signaling within the stomach to prevent spontaneous gastric inflammation and metaplasia, and suggest that glucocorticoid deficiency may lead to gastric cancer development.
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http://dx.doi.org/10.1172/JCI123233DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391099PMC
March 2019

Muscle-specific regulation of right ventricular transcriptional responses to chronic hypoxia-induced hypertrophy by the muscle ring finger-1 (MuRF1) ubiquitin ligase in mice.

BMC Med Genet 2018 09 21;19(1):175. Epub 2018 Sep 21.

McAllister Heart Institute, University of North Carolina, Chapel Hill, NC, USA.

Background: We recently identified a role for the muscle-specific ubiquitin ligase MuRF1 in right-sided heart failure secondary to pulmonary hypertension induced by chronic hypoxia (CH). MuRF1-/- mice exposed to CH are resistant to right ventricular (RV) dysfunction whereas MuRF1 Tg + mice exhibit impaired function indicative of heart failure. The present study was undertaken to understand the underlying transcriptional alterations in the RV of MuRF1-/- and MuRF1 Tg + mice.

Methods: Microarray analysis was performed on RNA isolated from the RV of MuRF1-/-, MuRF1 Tg+, and wild-type control mice exposed to CH.

Results: MuRF1-/- RV differentially expressed 590 genes in response to CH. Analysis of the top 66 genes (> 2-fold or < - 2-fold) revealed significant associations with oxidoreductase, transcription regulation, and transmembrane component annotations. The significant genes had promoters enriched for HOXD12, HOXC13, and RREB-1 protein transcription factor binding sites. MuRF1 Tg + RV differentially expressed 150 genes in response to CH. Analysis of the top 45 genes (> 3-fold or < - 3-fold) revealed significant associations with oxidoreductase-metabolic, glycoprotein-transmembrane-integral proteins, and alternative splicing/splice variant annotations. The significant genes were enriched for promoters with ZIC1 protein transcription factor binding sites.

Conclusions: The differentially expressed genes in MuRF1-/- and MuRF1 Tg + RV after CH have common functional annotations related to oxidoreductase (including antioxidant) and transmembrane component functions. Moreover, the functionally-enhanced MuRF1-/- hearts regulate genes related to transcription, homeobox proteins, and kinases/phosphorylation. These studies also reveal potential indirect effects of MuRF1 through regulating Rreb-1, and they reveal mechanisms by which MuRF1 may transcriptionally regulate anti-oxidant systems in the face of right heart failure.
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http://dx.doi.org/10.1186/s12881-018-0670-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6150973PMC
September 2018

MicroRNA Profiling and Bioinformatics Target Analysis in Dorsal Hippocampus of Chronically Stressed Rats: Relevance to Depression Pathophysiology.

Front Mol Neurosci 2018 6;11:251. Epub 2018 Aug 6.

Laboratory of Neuroplasticity and Neurogenetics, Faculty of Chemical and Pharmaceutical Sciences, Department of Biochemistry and Molecular Biology, Universidad de Chile, Santiago, Chile.

Studies conducted in rodents subjected to chronic stress and some observations in humans after psychosocial stress, have allowed to establish a link between stress and the susceptibility to many complex diseases, including mood disorders. The studies in rodents have revealed that chronic exposure to stress negatively affects synaptic plasticity by triggering changes in the production of trophic factors, subunit levels of glutamate ionotropic receptors, neuron morphology, and neurogenesis in the adult hippocampus. These modifications may account for the impairment in learning and memory processes observed in chronically stressed animals. It is plausible then, that stress modifies the interplay between signal transduction cascades and gene expression regulation in the hippocampus, therefore leading to altered neuroplasticity and functioning of neural circuits. Considering that miRNAs play an important role in post-transcriptional-regulation of gene expression and participate in several hippocampus-dependent functions; we evaluated the consequences of chronic stress on the expression of miRNAs in dorsal (anterior) portion of the hippocampus, which participates in memory formation in rodents. Here, we show that male rats exposed to daily restraint stress (2.5 h/day) during 7 and 14 days display a differential profile of miRNA levels in dorsal hippocampus and remarkably, we found that some of these miRNAs belong to the miR-379-410 cluster. We confirmed a rise in miR-92a and miR-485 levels after 14 days of stress by qPCR, an effect that was not mimicked by chronic administration of corticosterone (14 days). Our study identified the top-10 biological functions influenced by miR-92a, nine of which were shared with miR-485: Nervous system development and function, Tissue development, Behavior, Embryonic development, Organ development, Organismal development, Organismal survival, Tissue morphology, and Organ morphology. Furthermore, our study provided a landscape of potential miRNA-92a and miR-485 targets, along with relevant canonical pathways related to axonal guidance signaling and cAMP signaling, which may influence the functioning of several neuroplastic substrates in dorsal hippocampus. Additionally, the combined effect of miR-92a and miR-485 on transcription factors, along with histone-modifying enzymes, may have a functional relevance by producing changes in gene regulatory networks that modify the neuroplastic capacity of the adult dorsal hippocampus under stress.
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http://dx.doi.org/10.3389/fnmol.2018.00251DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6088391PMC
August 2018

Glucocorticoid Receptor Mutations and Hypersensitivity to Endogenous and Exogenous Glucocorticoids.

J Clin Endocrinol Metab 2018 10;103(10):3630-3639

National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina.

Background: The glucocorticoid receptor (GR) consists of two alternatively spliced isoforms: GRα, which activates gene transcription, and GRβ, a dominant-negative receptor. Theoretically, inactivating variants of GRβ could result in glucocorticoid hypersensitivity.

Design: A 46-year-old woman presented for evaluation of adrenal insufficiency prompted by low plasma cortisol levels and multiple unexplained symptoms but without clinical evidence of glucocorticoid insufficiency. To explain these findings, extensive clinical, genetic, and molecular studies were performed.

Methods: Standard clinical methods assessed the patient's hypothalamic-pituitary-adrenal axis. Validated molecular techniques were used for receptor sequencing, stable transfections, stimulation of candidate genes, cDNA arrays, Ingenuity Pathway Analysis, volcano analysis, and isolation and analysis of the patient's mononuclear cells.

Results: Clinical studies excluded primary or secondary adrenal insufficiency, established consistently low basal cortisol levels, and demonstrated hypersensitivity to ultra-low-dose dexamethasone. Receptor sequencing identified two variants of GR9β (A3669G and G3134T) as well as the known Bcl1 polymorphism. Reductionist studies using stable osteosarcoma cell lines transfected with the GRβ variants demonstrated glucocorticoid hypersensitivity of transcribed genes on cDNA array analysis. The patient's monocytes responded to hydrocortisone with exaggerated stimulation of the candidate genes GILZ and FKBP5.

Conclusion: Two variants of the dominant-negative GRβ, in conjunction with a common Bcl1 intron variant, resulted in hypersensitivity to endogenous and exogenous glucocorticoids and, as a reflection of severity, low circulating cortisol levels without clinical evidence of glucocorticoid insufficiency. This prismatic case exemplifies the unique effects of variants of a dominant-negative receptor.
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http://dx.doi.org/10.1210/jc.2018-00352DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6179182PMC
October 2018

Taxol Induces Brk-dependent Prosurvival Phenotypes in TNBC Cells through an AhR/GR/HIF-driven Signaling Axis.

Mol Cancer Res 2018 11 10;16(11):1761-1772. Epub 2018 Jul 10.

Division of Hematology, Oncology, and Transplantation, Departments of Medicine and Pharmacology and The Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota.

The metastatic cascade is a complex process that requires cancer cells to survive despite conditions of high physiologic stress. Previously, cooperation between the glucocorticoid receptor (GR) and hypoxia-inducible factors (HIF) was reported as a point of convergence for host and cellular stress signaling. These studies indicated p38 MAPK-dependent phosphorylation of GR on Ser134 and subsequent p-GR/HIF-dependent induction of breast tumor kinase (PTK6/Brk), as a mediator of aggressive cancer phenotypes. Herein, p-Ser134 GR was quantified in human primary breast tumors ( = 281) and the levels of p-GR were increased in triple-negative breast cancer (TNBC) relative to luminal breast cancer. Brk was robustly induced following exposure of TNBC model systems to chemotherapeutic agents (Taxol or 5-fluorouracil) and growth in suspension [ultra-low attachment (ULA)]. Notably, both Taxol and ULA resulted in upregulation of the Aryl hydrocarbon receptor (AhR), a known mediator of cancer prosurvival phenotypes. Mechanistically, AhR and GR copurified and following chemotherapy and ULA, these factors assembled at the Brk promoter and induced Brk expression in an HIF-dependent manner. Furthermore, Brk expression was upregulated in Taxol-resistant breast cancer (MCF-7) models. Ultimately, Brk was critical for TNBC cell proliferation and survival during Taxol treatment and in the context of ULA as well as for basal cancer cell migration, acquired biological phenotypes that enable cancer cells to successfully complete the metastatic cascade. These studies nominate AhR as a p-GR binding partner and reveal ways to target epigenetic events such as adaptive and stress-induced acquisition of cancer skill sets required for metastatic cancer spread. Breast cancer cells enlist intracellular stress response pathways that evade chemotherapy by increasing cancer cell survival and promoting migratory phenotypes. .
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http://dx.doi.org/10.1158/1541-7786.MCR-18-0410DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6214723PMC
November 2018

Glucocorticoid receptor isoform-specific regulation of development, circadian rhythm, and inflammation in mice.

FASEB J 2018 10 19;32(10):5258-5271. Epub 2018 Apr 19.

Signal Transduction Laboratory, Department of Health and Human Services, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA.

Glucocorticoids are primary stress hormones, and their synthetic derivatives are widely used clinically. The therapeutic efficacy of these steroids is limited by side effects and glucocorticoid resistance. Multiple glucocorticoid receptor (GR) isoforms are produced from a single gene by alternative translation initiation; however, the role individual isoforms play in tissue-specific responses to glucocorticoids is unknown. We have generated knockin mice that exclusively express the most active receptor isoform, GR-C3. GR-C3 knockin mice die at birth due to respiratory distress. Microarray analysis of fibroblasts from wild-type and GR-C3 mice indicated that most genes regulated by GR-C3 were unique to this isoform. Antenatal glucocorticoid administration rescued GR-C3 knockin mice from neonatal death. Dual-energy X-ray absorptiometry revealed no major alterations in body composition for rescued knockin mice. Rescued female, but not male, GR-C3 mice exhibited increased wheel running activity in the light portion of the day. LPS administration induced premature mortality in rescued GR-C3 knockin mice, and gene expression studies revealed a deficiency in the ability of GR-C3 to repress a large cohort of immune and inflammatory response genes. These findings demonstrate that specific GR translational isoforms can influence development, circadian rhythm, and inflammation through the regulation of distinct gene networks.-Oakley, R. H., Ramamoorthy, S., Foley, J. F., Busada, J. T., Lu, N. Z., Cidlowski, J. A. Glucocorticoid receptor isoform-specific regulation of development, circadian rhythm, and inflammation in mice.
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http://dx.doi.org/10.1096/fj.201701153RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6133704PMC
October 2018

Neonatal Genistein Exposure and Glucocorticoid Signaling in the Adult Mouse Uterus.

Environ Health Perspect 2018 04 5;126(4):047002. Epub 2018 Apr 5.

Laboratory of Signal Transduction, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, Research Triangle Park, North Carolina, USA.

Background: Female reproductive tract development is sensitive to the endocrine-disrupting potential of environmental estrogens. Early-life exposure to the dietary phytoestrogen genistein impairs fertility and persistently alters the transcriptome in the oviduct and uterus of rodents. Glucocorticoid signaling, which has recently been shown to be essential for normal fertility in the female mouse uterus, is antagonized by genistein.

Objective: Our goal was to determine whether early-life exposure to genistein disrupts glucocorticoid signaling in the mouse uterus, which may contribute to infertility.

Methods: Female C57Bl/6 mice were exposed to either 50 mg/kg per day genistein, 10 μg/kg per day estradiol, or vehicle (corn oil) on postnatal days 1-5 (PND1-5), and then treated with the synthetic glucocorticoid dexamethasone (Dex: 1 mg/kg) or vehicle (saline) on PND5, at weaning on PND21, or as adults on PND56 following adrenalectomy and ovariectomy to evaluate glucocorticoid responsiveness. Uteri were isolated following treatment for gene expression or chromatin immunoprecipitation.

Results: Neonatal exposure to genistein altered the uterine transcriptome of adult mice and caused substantial changes to the transcriptional response to glucocorticoids. Although expression of the glucocorticoid receptor was not affected, genistein exposure disrupted glucocorticoid receptor recruitment to specific regulatory sites in target genes. Many genes involved in chromatin remodeling were dysregulated in genistein-exposed mice, suggesting that epigenetic reprograming may contribute to the altered glucocorticoid response of the uterus following early-life exposure to genistein. These changes affected the biological activity of glucocorticoids within the uterus, as glucocorticoids antagonized the proliferative effects of estradiol in the uterus of control mice but not genistein-exposed mice.

Conclusions: Our findings suggest that disruption of glucocorticoid signaling due to early-life exposure to environmental estrogens may in part render the uterus unable to support implantation. https://doi.org/10.1289/EHP1575.
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http://dx.doi.org/10.1289/EHP1575DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6071733PMC
April 2018

Estrogen Deficiency Promotes Hepatic Steatosis via a Glucocorticoid Receptor-Dependent Mechanism in Mice.

Cell Rep 2018 03;22(10):2690-2701

Signal Transduction Laboratory, National Institute of Environmental Health Sciences, NIH, Department of Health and Human Services, Research Triangle Park, NC 27709, USA. Electronic address:

Glucocorticoids (GCs) are master regulators of systemic metabolism. Intriguingly, Cushing's syndrome, a disorder of excessive GCs, phenocopies several menopause-induced metabolic pathologies. Here, we show that the glucocorticoid receptor (GR) drives steatosis in hypogonadal female mice because hepatocyte-specific GR knockout mice are refractory to developing ovariectomy-induced steatosis. Intriguingly, transcriptional profiling revealed that ovariectomy elicits hepatic GC hypersensitivity globally. Hypogonadism-induced GC hypersensitivity results from a loss of systemic but not hepatic estrogen (E2) signaling, given that hepatocyte-specific E2 receptor deletion does not confer GC hypersensitivity. Mechanistically, enhanced chromatin recruitment and ligand-dependent hyperphosphorylation of GR underlie ovariectomy-induced glucocorticoid hypersensitivity. The dysregulated glucocorticoid-mediated signaling present in hypogonadal females is a product of increased follicle-stimulating hormone (FSH) production because FSH treatment in ovary-intact mice recapitulates glucocorticoid hypersensitivity similar to hypogonadal female mice. Our findings uncover a regulatory axis between estradiol, FSH, and hepatic glucocorticoid receptor signaling that, when disrupted, as in menopause, promotes hepatic steatosis.
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http://dx.doi.org/10.1016/j.celrep.2018.02.041DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5875726PMC
March 2018

Probing Dominant Negative Behavior of Glucocorticoid Receptor β through a Hybrid Structural and Biochemical Approach.

Mol Cell Biol 2018 04 29;38(8). Epub 2018 Mar 29.

Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, USA

Glucocorticoid receptor β (GRβ) is associated with glucocorticoid resistance via dominant negative regulation of GRα. To better understand how GRβ functions as a dominant negative inhibitor of GRα at a molecular level, we determined the crystal structure of the ligand binding domain of GRβ complexed with the antagonist RU-486. The structure reveals that GRβ binds RU-486 in the same ligand binding pocket as GRα, and the unique C-terminal amino acids of GRβ are mostly disordered. Binding energy analysis suggests that these C-terminal residues of GRβ do not contribute to RU-486 binding. Intriguingly, the GRβ/RU-486 complex binds corepressor peptide with affinity similar to that of a GRα/RU-486 complex, despite the lack of helix 12. Our biophysical and biochemical analyses reveal that in the presence of RU-486, GRβ is found in a conformation that favors corepressor binding, potentially antagonizing GRα function. This study thus presents an unexpected molecular mechanism by which GRβ could repress transcription.
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http://dx.doi.org/10.1128/MCB.00453-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5879464PMC
April 2018

Molecular mechanisms of cell death: recommendations of the Nomenclature Committee on Cell Death 2018.

Cell Death Differ 2018 03 23;25(3):486-541. Epub 2018 Jan 23.

Department of Medicine, Division of Pulmonary and Critical Care Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL, USA.

Over the past decade, the Nomenclature Committee on Cell Death (NCCD) has formulated guidelines for the definition and interpretation of cell death from morphological, biochemical, and functional perspectives. Since the field continues to expand and novel mechanisms that orchestrate multiple cell death pathways are unveiled, we propose an updated classification of cell death subroutines focusing on mechanistic and essential (as opposed to correlative and dispensable) aspects of the process. As we provide molecularly oriented definitions of terms including intrinsic apoptosis, extrinsic apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, pyroptosis, parthanatos, entotic cell death, NETotic cell death, lysosome-dependent cell death, autophagy-dependent cell death, immunogenic cell death, cellular senescence, and mitotic catastrophe, we discuss the utility of neologisms that refer to highly specialized instances of these processes. The mission of the NCCD is to provide a widely accepted nomenclature on cell death in support of the continued development of the field.
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http://dx.doi.org/10.1038/s41418-017-0012-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5864239PMC
March 2018

Pharmacology of Corticosteroids for Diabetic Macular Edema.

Invest Ophthalmol Vis Sci 2018 01;59(1):1-12

Center for Advanced Vision Science, Department of Ophthalmology, Department of Pathology, Department of Microbiology, Immunology, and Cancer Biology, University of Virginia School of Medicine, Charlottesville, Virginia, United States.

Purpose: Corticosteroids remain the mainstay of treatment for inflammatory diseases almost 80 years after their first clinical use. Topical ophthalmic formulations of corticosteroids have been available to treat disease of the anterior segment of the eye, but the approval of corticosteroids to treat vitreoretinal diseases, including vein occlusion, diabetic macular edema, and uveitis, has occurred only recently. Although most diseases respond to corticosteroid therapy, some patients are resistant to this therapy and side effects, including cataract and elevated intraocular pressure, can limit their use. The purpose of this review is to detail the basic science of corticosteroids focusing on differences in potency, drug delivery, pharmacokinetics, and gene activation, and how these differences affect safety and efficacy in the treatment of diabetic macular edema.

Methods: A review was conducted of basic science and pharmacology of the corticosteroids used to treat diabetic macular edema.

Results: Clinically available corticosteroids not only have differing potency and pharmacokinetics, but also activate different genes in different target tissues. These differences are associated with distinct efficacy, pharmacokinetic, and safety profiles. It is important to understand these differences in selecting corticosteroids to treat diabetic macular edema.

Conclusions: Recent advances in our understanding of the basic science of corticosteroids can explain clinical differences in these agents regarding efficacy and safety. Importantly, this understanding should allow the future discovery of additional novel corticosteroids to treat diabetic macular edema.
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http://dx.doi.org/10.1167/iovs.17-22259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5961100PMC
January 2018