Pulmonology
Abstract
Abnormal airway smooth muscle function can contribute to cystic fibrosis (CF) airway disease. We previously found that airway smooth muscle from newborn CF pigs had increased basal tone, an increased bronchodilator response, and abnormal calcium handling. Since CF pigs lack airway infection and inflammation at birth, these findings suggest intrinsic airway smooth muscle dysfunction in CF. In this study, we tested the hypothesis that CFTR loss in airway smooth muscle would produce a distinct set of changes in the airway smooth muscle transcriptome that we could use to develop novel therapeutic targets. Total RNA sequencing of newborn wild-type and CF airway smooth muscle revealed changes in muscle contraction–related genes, ontologies, and pathways. Using connectivity mapping, we identified several small molecules that elicit transcriptional signatures opposite of CF airway smooth muscle, including NVP-TAE684, an inhibitor of proline-rich tyrosine kinase 2 (PYK2). In CF airway smooth muscle tissue, PYK2 phosphorylation was increased and PYK2 inhibition decreased smooth muscle contraction. In vivo NVP-TAE684 treatment of wild-type mice reduced methacholine-induced airway smooth muscle contraction. These findings suggest that studies in the newborn CF pig may provide an important approach to enhance our understanding of airway smooth muscle biology and for discovery of novel airway smooth muscle therapeutics for CF and other diseases of airway hyperreactivity.
Authors
Daniel P. Cook, Ryan J. Adam, Keyan Zarei, Benjamin Deonovic, Mallory R. Stroik, Nicholas D. Gansemer, David K. Meyerholz, Kin Fai Au, David A. Stoltz
Abstract
The pathogenesis of primary Sjogren’s syndrome (SS), an autoimmune disease that targets the mucosa of exocrine tissues, is poorly understood. Although several mouse models have been developed that display features of SS, most of these are within the larger context of a lupus-like presentation. Immunity-related GTPase family M protein 1 (Irgm1) is an interferon-inducible cytoplasmic GTPase that is reported to regulate autophagy and mitochondrial homeostasis. Here, we report that naive Irgm1–/– mice display lymphocytic infiltration of multiple mucosal tissues including the lung in a manner reminiscent of SS, together with IgA class–predominant autoantibodies including anti-Ro and anti-La. This phenotype persists in the germ-free state, but is abolished by deletion of Irgm3. Irgm1–/– mice have increased local production in the lung of TECP15-idiotype IgA, a natural antibody with dual reactivity against host and pneumococcal phosphorylcholine. Associated with this, Irgm1–/– mice display enhanced opsonization and clearance of Streptococcus pneumoniae from the lung and increased survival from pneumococcal pneumonia. Taken together, our results identify Irgm1 as a master regulator of mucosal immunity that dually modulates evolutionarily conserved self- and other-directed immune responses at the interface of host with environment.
Authors
Kathleen M. Azzam, Jennifer H. Madenspacher, Derek W. Cain, Lihua Lai, Kymberly M. Gowdy, Prashant Rai, Kyathanahalli Janardhan, Natasha Clayton, Willie Cunningham, Heather Jensen, Preeyam S. Patel, John F. Kearney, Gregory A. Taylor, Michael B. Fessler
Abstract
The chronic progressive decline in lung function observed in idiopathic pulmonary fibrosis (IPF) appears to result from persistent nonresolving injury to the epithelium, impaired restitution of the epithelial barrier in the lung, and enhanced fibroblast activation. Thus, understanding these key mechanisms and pathways modulating both is essential to greater understanding of IPF pathogenesis. We examined the association of VEGF with the IPF disease state and preclinical models in vivo and in vitro. Tissue and circulating levels of VEGF were significantly reduced in patients with IPF, particularly in those with a rapidly progressive phenotype, compared with healthy controls. Lung-specific overexpression of VEGF significantly protected mice following intratracheal bleomycin challenge, with a decrease in fibrosis and bleomycin-induced cell death observed in the VEGF transgenic mice. In vitro, apoptotic endothelial cell–derived mediators enhanced epithelial cell injury and reduced epithelial wound closure. This process was rescued by VEGF pretreatment of the endothelial cells via a mechanism involving thrombospondin-1 (TSP1). Taken together, these data indicate beneficial roles for VEGF during lung fibrosis via modulating epithelial homeostasis through a previously unrecognized mechanism involving the endothelium.
Authors
Lynne A. Murray, David M. Habiel, Miriam Hohmann, Ana Camelo, Huilan Shang, Yang Zhou, Ana Lucia Coelho, Xueyan Peng, Mridu Gulati, Bruno Crestani, Matthew A. Sleeman, Tomas Mustelin, Meagan W. Moore, Changwan Ryu, Awo D. Osafo-Addo, Jack A. Elias, Chun G. Lee, Buqu Hu, Jose D. Herazo-Maya, Darryl A. Knight, Cory M. Hogaboam, Erica L. Herzog
Abstract
BACKGROUND. Right-sided heart failure is the leading cause of death in pulmonary arterial hypertension (PAH). Similar to left heart failure, sympathetic overactivation and β-adrenoreceptor (βAR) abnormalities are found in PAH. Based on successful therapy of left heart failure with β-blockade, the safety and benefits of the nonselective β-blocker/vasodilator carvedilol were evaluated in PAH. METHODS. PAH Treatment with Carvedilol for Heart Failure (PAHTCH) is a single-center, double-blind, randomized, controlled trial. Following 1-week run-in, 30 participants were randomized to 1 of 3 arms for 24 weeks: placebo, low-fixed-dose, or dose-escalating carvedilol. Outcomes included clinical measures and mechanistic biomarkers. RESULTS. Decreases in heart rate and blood pressure with carvedilol were well tolerated; heart rate correlated with carvedilol dose. Carvedilol-treated groups had no decrease in exercise capacity measured by 6-minute walk, but had lower heart rates at peak and after exercise, and faster heart rate recovery. Dose-escalating carvedilol was associated with reduction in right ventricular (RV) glycolytic rate and increase in βAR levels. There was no evidence of RV functional deterioration; rather, cardiac output was maintained. CONCLUSIONS. Carvedilol is likely safe in PAH over 6 months of therapy and has clinical and mechanistic benefits associated with improved outcomes. The data provide support for longer and larger studies to establish guidelines for use of β-blockers in PAH. TRIAL REGISTRATION. ClinicalTrials.gov NCT01586156 FUNDING. This project was supported by NIH R01HL115008 and R01HL60917 and in part by the National Center for Advancing Translational Sciences, UL1TR000439.
Authors
Samar Farha, Didem Saygin, Margaret M. Park, Hoi I. Cheong, Kewal Asosingh, Suzy A.A. Comhair, Olivia R. Stephens, Emir C. Roach, Jacqueline Sharp, Kristin B. Highland, Frank P. DiFilippo, Donald R. Neumann, W.H. Wilson Tang, Serpil C. Erzurum
Abstract
BACKGROUND. In health, inflammation resolution is an active process governed by specialized proresolving mediators and receptors. ALX/FPR2 receptors (ALX) are targeted by both proresolving and proinflammatory ligands for opposing signaling events, suggesting pivotal roles for ALX in the fate of inflammatory responses. Here, we determined if ALX expression and ligands were linked to severe asthma (SA). METHODS. ALX expression and levels of proresolving ligands (lipoxin A4 [LXA4], 15-epi-LXA4, and annexin A1 [ANXA1]), and a proinflammatory ligand (serum amyloid A [SAA]) were measured in bronchoscopy samples collected in Severe Asthma Research Program-3 (SA [n = 69], non-SA [NSA, n = 51] or healthy donors [HDs, n = 47]). RESULTS. Bronchoalveolar lavage (BAL) fluid LXA4 and 15-epi-LXA4 were decreased and SAA was increased in SA relative to NSA. BAL macrophage ALX expression was increased in SA. Subjects with LXA4loSAAhi levels had increased BAL neutrophils, more asthma symptoms, lower lung function, increased relative risk for asthma exacerbation, sinusitis, and gastroesophageal reflux disease, and were assigned more frequently to SA clinical clusters. SAA and aliquots of LXA4loSAAhi BAL fluid induced IL-8 production by lung epithelial cells expressing ALX receptors, which was inhibited by coincubation with 15-epi-LXA4. CONCLUSIONS. Together, these findings have established an association between select ALX receptor ligands and asthma severity that define a potentially new biochemical endotype for asthma and support a pivotal functional role for ALX signaling in the fate of lung inflammation. TRIAL REGISTRATION. Severe Asthma Research Program-3 (SARP-3; ClinicalTrials.gov NCT01606826) FUNDING Sources. National Heart, Lung and Blood Institute, the NIH, and the German Society of Pediatric Pneumology.
Authors
Isabell Ricklefs, Ioanna Barkas, Melody G. Duvall, Manuela Cernadas, Nicole L. Grossman, Elliot Israel, Eugene R. Bleecker, Mario Castro, Serpil C. Erzurum, John V. Fahy, Benjamin M. Gaston, Loren C. Denlinger, David T. Mauger, Sally E. Wenzel, Suzy A. Comhair, Andrea M. Coverstone, Merritt L. Fajt, Annette T. Hastie, Mats W. Johansson, Michael C. Peters, Brenda R. Phillips, Bruce D. Levy, the National Heart Lung and Blood Institute’s Severe Asthma Research Program-3 Investigators
Abstract
We previously showed that Th1/type 1 inflammation marked by increased IFN-γ levels in the airways can be appreciated in 50% of patients with severe asthma, despite high dose corticosteroid (CS) treatment. We hypothesized that a downstream target of IFN-γ, CXCL10, which recruits Th1 cells via the cognate receptor CXCR3, is an important contributor to Th1high asthma and CS unresponsiveness. We show high levels of CXCL10 mRNA closely associated with IFNG levels in the BAL cells of 50% of severe asthmatics and also in the airways of mice subjected to a severe asthma model, both in the context of high-dose CS treatment. The inability of CS to dampen IFNG or CXCL10 expression was not because of impaired nuclear translocation of the glucocorticoid receptor (GR) or its transactivational functions. Rather, in the presence of CS and IFN-γ, STAT1 and GR were recruited on critical regulatory elements in the endogenous CXCL10 promoter in monocytes, albeit without any abatement of CXCL10 gene expression. High CXCL10 gene expression was also associated with a mast cell signature in both humans and mice, CXCR3 being also expressed by mast cells. These findings suggest that the IFN-γ–CXCL10 axis plays a central role in persistent type 1 inflammation that may be facilitated by CS therapy through GR-STAT1 cooperation converging on the CXCL10 promoter.
Authors
Marc Gauthier, Krishnendu Chakraborty, Timothy B. Oriss, Mahesh Raundhal, Sudipta Das, Jie Chen, Rachael Huff, Ayan Sinha, Merritt Fajt, Prabir Ray, Sally E. Wenzel, Anuradha Ray
Abstract
Mechanical ventilation is necessary to support patients with acute lung injury, but also exacerbates injury through mechanical stress–activated signaling pathways. We show that stretch applied to cultured human cells, and to mouse lungs in vivo, induces robust expression of metallothionein, a potent antioxidant and cytoprotective molecule critical for cellular zinc homeostasis. Furthermore, genetic deficiency of murine metallothionein genes exacerbated lung injury caused by high tidal volume mechanical ventilation, identifying an adaptive role for these genes in limiting lung injury. Stretch induction of metallothionein required zinc and the zinc-binding transcription factor MTF1. We further show that mouse dietary zinc deficiency potentiates ventilator-induced lung injury, and that plasma zinc levels are significantly reduced in human patients who go on to develop acute respiratory distress syndrome (ARDS) compared with healthy and non-ARDS intensive care unit (ICU) controls, as well as with other ICU patients without ARDS. Taken together, our findings identify a potentially novel adaptive response of the lung to stretch and a critical role for zinc in defining the lung’s tolerance for mechanical ventilation. These results demonstrate that failure of stretch-adaptive responses play an important role in exacerbating mechanical ventilator–induced lung injury, and identify zinc and metallothionein as targets for lung-protective interventions in patients requiring mechanical ventilation.
Authors
Francis Boudreault, Miguel Pinilla-Vera, Joshua A. Englert, Alvin T. Kho, Colleen Isabelle, Antonio J. Arciniegas, Diana Barragan-Bradford, Carolina Quintana, Diana Amador-Munoz, Jiazhen Guan, Kyoung Moo Choi, MICU Registry, Lynette Sholl, Shelley Hurwitz, Daniel J. Tschumperlin, Rebecca M. Baron
Abstract
Fibrosis results from the dysregulation of tissue repair mechanisms affecting major organ systems, leading to chronic extracellular matrix buildup, and progressive, often fatal, organ failure. Current diagnosis relies on invasive biopsies. Noninvasive methods today cannot distinguish actively progressive fibrogenesis from stable scar, and thus are insensitive for monitoring disease activity or therapeutic responses. Collagen oxidation is a universal signature of active fibrogenesis that precedes collagen crosslinking. Biochemically targeting oxidized lysine residues formed by the action of lysyl oxidase on collagen with a small-molecule gadolinium chelate enables targeted molecular magnetic resonance imaging. This noninvasive direct biochemical elucidation of the fibrotic microenvironment specifically and robustly detected and staged pulmonary and hepatic fibrosis progression, and monitored therapeutic response in animal models. Furthermore, this paradigm is translatable and generally applicable to diverse fibroproliferative disorders.
Authors
Howard H. Chen, Philip A. Waghorn, Lan Wei, Luis F. Tapias, Daniel T. Schühle, Nicholas J. Rotile, Chloe M. Jones, Richard J. Looby, Gaofeng Zhao, Justin M. Elliott, Clemens K. Probst, Mari Mino-Kenudson, Gregory Y. Lauwers, Andrew M. Tager, Kenneth K. Tanabe, Michael Lanuti, Bryan C. Fuchs, Peter Caravan
Abstract
Pulmonary function is dependent upon the precise regulation of alveolar surfactant. Alterations in pulmonary surfactant concentrations or function impair ventilation and cause tissue injury. Identification of the molecular pathways that sense and regulate endogenous alveolar surfactant concentrations, coupled with the ability to pharmacologically modulate them both positively and negatively, would be a major therapeutic advance for patients with acute and chronic lung diseases caused by disruption of surfactant homeostasis. The orphan adhesion GPCR GPR116 (also known as Adgrf5) is a critical regulator of alveolar surfactant concentrations. Here, we show that human and mouse GPR116 control surfactant secretion and reuptake in alveolar type II (AT2) cells by regulating guanine nucleotide–binding domain α q and 11 (Gq/11) signaling. Synthetic peptides derived from the ectodomain of GPR116 activated Gq/11-dependent inositol phosphate conversion, calcium mobilization, and cortical F-actin stabilization to inhibit surfactant secretion. AT2 cell–specific deletion of Gnaq and Gna11 phenocopied the accumulation of surfactant observed in Gpr116–/– mice. These data provide proof of concept that GPR116 is a plausible therapeutic target to modulate endogenous alveolar surfactant pools to treat pulmonary diseases associated with surfactant dysfunction.
Authors
Kari Brown, Alyssa Filuta, Marie-Gabrielle Ludwig, Klaus Seuwen, Julian Jaros, Solange Vidal, Kavisha Arora, Anjaparavanda P. Naren, Kathirvel Kandasamy, Kaushik Parthasarathi, Stefan Offermanns, Robert J. Mason, William E. Miller, Jeffrey A. Whitsett, James P. Bridges
Abstract
Severe asthma (SA) is a significant problem both clinically and economically, given its poor response to corticosteroids (CS). We recently reported a complex type 1–dominated (IFN-γ–dominated) immune response in more than 50% of severe asthmatics despite high-dose CS treatment. Also, IFN-γ was found to be critical for increased airway hyperreactivity (AHR) in our model of SA. The transcription factor IRF5 expressed in M1 macrophages can induce a Th1/Th17 response in cocultured human T cells. Here we show markedly higher expression of IRF5 in bronchoalveolar lavage (BAL) cells of severe asthmatics as compared with that in cells from milder asthmatics or healthy controls. Using our SA mouse model, we demonstrate that lack of IRF5 in lymph node migratory DCs severely limits their ability to stimulate the generation of IFN-γ– and IL-17–producing CD4+ T cells and
Authors
Timothy B. Oriss, Mahesh Raundhal, Christina Morse, Rachael E. Huff, Sudipta Das, Rachel Hannum, Marc C. Gauthier, Kathryn L. Scholl, Krishnendu Chakraborty, Seyed M. Nouraie, Sally E. Wenzel, Prabir Ray, Anuradha Ray
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