Pulmonology
Abstract
Authors
Kaushik Parthasarathi, Hideo Ichimura, Eiji Monma, Jens Lindert, Sadiqa Quadri, Andrew Issekutz, Jahar Bhattacharya
Abstract
Acute lung injury (ALI), which is associated with a mortality of 30–40%, is attributable to inflammation that develops rapidly across the lung’s vast vascular surface, involving an entire lung or even both lungs. No specific mechanism explains this extensive inflammatory spread, probably because of the lack of approaches for detecting signal conduction in lung capillaries. Here, we addressed this question by applying the photolytic uncaging approach to induce focal increases in Ca2+ levels in targeted endothelial cells of alveolar capillaries. Uncaging caused Ca2+ levels to increase not only in the targeted cell, but also in vascular locations up to 150 μm from the target site, indicating that Ca2+ was conducted from the capillary to adjacent vessels. No such conduction was evident in mouse lungs lacking endothelial connexin 43 (Cx43), or in rat lungs in which we pretreated vessels with peptide inhibitors of Cx43. These findings provide the first direct evidence to our knowledge that interendothelial Ca2+ conduction occurs in the lung capillary bed and that Cx43-containing gap junctions mediate the conduction. A proinflammatory effect was evident in that induction of increases in Ca2+ levels in the capillary activated expression of the leukocyte adherence receptor P-selectin in venules. Further, peptide inhibitors of Cx43 completely blocked thrombin-induced microvascular permeability increases. Together, our findings reveal a novel role for Cx43-mediated gap junctions, namely as conduits for the spread of proinflammatory signals in the lung capillary bed. Gap junctional mechanisms require further consideration in the understanding of ALI.
Authors
Kaushik Parthasarathi, Hideo Ichimura, Eiji Monma, Jens Lindert, Sadiqa Quadri, Andrew Issekutz, Jahar Bhattacharya
Abstract
Adenosine has been implicated in the pathogenesis of chronic lung diseases such as asthma and chronic obstructive pulmonary disease. In vitro studies suggest that activation of the A2B adenosine receptor (A2BAR) results in proinflammatory and profibrotic effects relevant to the progression of lung diseases; however, in vivo data supporting these observations are lacking. Adenosine deaminase–deficient (ADA-deficient) mice develop pulmonary inflammation and injury that are dependent on increased lung adenosine levels. To investigate the role of the A2BAR in vivo, ADA-deficient mice were treated with the selective A2BAR antagonist CVT-6883, and pulmonary inflammation, fibrosis, and airspace integrity were assessed. Untreated and vehicle-treated ADA-deficient mice developed pulmonary inflammation, fibrosis, and enlargement of alveolar airspaces; conversely, CVT-6883–treated ADA-deficient mice showed less pulmonary inflammation, fibrosis, and alveolar airspace enlargement. A2BAR antagonism significantly reduced elevations in proinflammatory cytokines and chemokines as well as mediators of fibrosis and airway destruction. In addition, treatment with CVT-6883 attenuated pulmonary inflammation and fibrosis in wild-type mice subjected to bleomycin-induced lung injury. These findings suggest that A2BAR signaling influences pathways critical for pulmonary inflammation and injury in vivo. Thus in chronic lung diseases associated with increased adenosine, antagonism of A2BAR-mediated responses may prove to be a beneficial therapy.
Authors
Chun-Xiao Sun, Hongyan Zhong, Amir Mohsenin, Eva Morschl, Janci L. Chunn, Jose G. Molina, Luiz Belardinelli, Dewan Zeng, Michael R. Blackburn
Abstract
Bronchial asthma, the most prevalent cause of significant respiratory morbidity in the developed world, typically is a chronic disorder associated with long-term changes in the airways. We developed a mouse model of chronic asthma that results in markedly increased numbers of airway mast cells, enhanced airway responses to methacholine or antigen, chronic inflammation including infiltration with eosinophils and lymphocytes, airway epithelial goblet cell hyperplasia, enhanced expression of the mucin genes Muc5ac and Muc5b, and increased levels of lung collagen. Using mast cell–deficient (KitW-sh/W-sh and/or KitW/W-v) mice engrafted with FcRγ+/+ or FcRγ–/– mast cells, we found that mast cells were required for the full development of each of these features of the model. However, some features also were expressed, although usually at less than wild-type levels, in mice whose mast cells lacked FcRγ and therefore could not be activated by either antigen- and IgE-dependent aggregation of FcεRI or the binding of antigen-IgG1 immune complexes to FcγRIII. These findings demonstrate that mast cells can contribute to the development of multiple features of chronic asthma in mice and identify both FcRγ-dependent and FcRγ-independent pathways of mast cell activation as important for the expression of key features of this asthma model.
Authors
Mang Yu, Mindy Tsai, See-Ying Tam, Carol Jones, James Zehnder, Stephen J. Galli
Abstract
Activation of latent TGF-β by the αvβ6 integrin is a critical step in the development of acute lung injury. However, the mechanism by which αvβ6-mediated TGF-β activation is regulated has not been identified. We show that thrombin, and other agonists of protease-activated receptor 1 (PAR1), activate TGF-β in an αvβ6 integrin–specific manner. This effect is PAR1 specific and is mediated by RhoA and Rho kinase. Intratracheal instillation of the PAR1-specific peptide TFLLRN increases lung edema during high-tidal-volume ventilation, and this effect is completely inhibited by a blocking antibody against the αvβ6 integrin. Instillation of TFLLRN during high-tidal-volume ventilation is associated with increased pulmonary TGF-β activation; however, this is not observed in Itgb6–/– mice. Furthermore, Itgb6–/– mice are also protected from ventilator-induced lung edema. We also demonstrate that pulmonary edema and TGF-β activity are similarly reduced in Par1–/– mice following bleomycin-induced lung injury. These results suggest that PAR1-mediated enhancement of αvβ6-dependent TGF-β activation could be one mechanism by which activation of the coagulation cascade contributes to the development of acute lung injury, and they identify PAR1 and the αvβ6 integrin as potential therapeutic targets in this condition.
Authors
R. Gisli Jenkins, Xiao Su, George Su, Christopher J. Scotton, Eric Camerer, Geoffrey J. Laurent, George E. Davis, Rachel C. Chambers, Michael A. Matthay, Dean Sheppard
Abstract
Transfusion-related acute lung injury (TRALI) is the most common cause of transfusion-related mortality. To explore the pathogenesis of TRALI, we developed an in vivo mouse model based on the passive transfusion of an MHC class I (MHC I) mAb (H2Kd) to mice with the cognate antigen. Transfusion of the MHC I mAb to BALB/c mice produced acute lung injury with increased excess lung water, increased lung vascular and lung epithelial permeability to protein, and decreased alveolar fluid clearance. There was 50% mortality at a 2-hour time point after Ab administration. Pulmonary histology and immunohistochemistry revealed prominent neutrophil sequestration in the lung microvasculature that occurred concomitantly with acute peripheral blood neutropenia, all within 2 hours of administration of the mAb. Depletion of neutrophils by injection of anti-granulocyte mAb Gr-1 protected mice from lung injury following MHC I mAb challenge. FcRγ–/– mice were resistant to MHC I mAb–induced lung injury, while adoptive transfer of wild-type neutrophils into the FcRγ–/– animals restored lung injury following MHC I mAb challenge. In conclusion, in a clinically relevant in vivo mouse model of TRALI using an MHC I mAb, the mechanism of lung injury was dependent on neutrophils and their Fcγ receptors.
Authors
Mark R. Looney, Xiao Su, Jessica A. Van Ziffle, Clifford A. Lowell, Michael A. Matthay
Abstract
To define the factors that control the tissue effects of IL-4, we compared the effects of Tg IL-4 in Balb/c and C57BL/6 mice. In the former, IL-4 caused modest eosinophilic inflammation and mild airway fibrosis and did not shorten survival. In C57BL/6 mice, IL-4 caused profound eosinophilic inflammation, airway fibrosis, emphysematous alveolar destruction, and premature death. These differences could not be accounted for by changes in Th2 or Th1 cytokines, receptor components, STAT6 activation, MMPs, or cathepsins. In contrast, in C57BL/6 mice, alveolar remodeling was associated with decreased levels of tissue inhibitors of metalloproteinase 2, -3, and -4 and α1-antitrypsin, and fibrosis was associated with increased levels of total and bioactive TGF-β1. Impressive differences in adenosine metabolism were also appreciated, with increased tissue adenosine levels and A1, A2B, and A3 adenosine receptor expression and decreased adenosine deaminase (ADA) activity in C57BL/6 animals. Treatment with ADA also reduced the inflammation, fibrosis, and emphysematous destruction and improved the survival of C57BL/6 Tg animals. These studies demonstrate that genetic influences control IL-4 effector pathways in the murine lung. They also demonstrate that IL-4 has different effects on adenosine metabolism in Balb/c and C57BL/6 mice and that these differences contribute to the different responses that IL-4 induces in these inbred animals.
Authors
Bing Ma, Michael R. Blackburn, Chun Geun Lee, Robert J. Homer, Wei Liu, Richard A. Flavell, Lynn Boyden, Richard P. Lifton, Chun-Xiao Sun, Hays W. Young, Jack A. Elias
Abstract
In models of acute lung injury, CXC chemokine receptor 2 (CXCR2) mediates migration of polymorphonuclear leukocytes (PMNs) into the lung. Since CXCR2 ligands, including CXCL1 and CXCL2/3, are chemotactic for PMNs, CXCR2 is thought to recruit PMNs by inducing chemotactic migration. In a model of PMN recruitment to the lung, aerosolized bacterial LPS inhalation induced PMN recruitment to the lung in wild-type mice, but not in littermate CXCR2–/– mice. Surprisingly, lethally irradiated wild-type mice reconstituted with CXCR2–/– BM still showed about 50% PMN recruitment into bronchoalveolar lavage fluid and into lung interstitium, but CXCR2–/– mice reconstituted with CXCR2–/– BM showed no PMN recruitment. Conversely, CXCR2–/– mice reconstituted with wild-type BM showed a surprisingly large defect in PMN recruitment, inconsistent with a role of CXCR2 on PMNs alone. Cell culture, immunohistochemistry, flow cytometry, and real-time RT-PCR were used to show expression of CXCR2 on pulmonary endothelial and bronchial epithelial cells. The LPS-induced increase in lung microvascular permeability as measured by Evans blue extravasation required CXCR2 on nonhematopoietic cells. Our data revealed what we believe to be a previously unrecognized role of endothelial and epithelial CXCR2 in LPS-induced PMN recruitment and lung injury.
Authors
Jörg Reutershan, Margaret A. Morris, Tracy L. Burcin, David F. Smith, Daniel Chang, Mary S. Saprito, Klaus Ley
Abstract
Complement component 5 (C5) has been described as either promoting or protecting against airway hyperresponsiveness (AHR) in experimental allergic asthma, suggesting pleomorphic effects of C5. Here we report that local pharmacological targeting of the C5a receptor (C5aR) prior to initial allergen sensitization in murine models of inhalation tolerance or allergic asthma resulted in either induction or marked enhancement of Th2-polarized immune responses, airway inflammation, and AHR. Importantly, C5aR-deficient mice exhibited a similar, increased allergic phenotype. Pulmonary allergen exposure in C5aR-targeted mice resulted in increased sensitization and accumulation of CD4+CD69+ T cells associated with a marked increase in pulmonary myeloid, but not plasmacytoid, DC numbers. Pulmonary DCs from C5aR-targeted mice produced large amounts of CC chemokine ligand 17 (CCL17) and CCL22 ex vivo, suggesting a negative impact of C5aR signaling on pulmonary homing of Th2 cells. In contrast, C5aR targeting in sensitized mice led to suppressed airway inflammation and AHR but was still associated with enhanced production of Th2 effector cytokines. These data suggest a dual role for C5a in allergic asthma, i.e., protection from the development of maladaptive type 2 immune responses during allergen sensitization at the DC/T cell interface but enhancement of airway inflammation and AHR in an established inflammatory environment.
Authors
Jörg Köhl, Ralf Baelder, Ian P. Lewkowich, Manoj K. Pandey, Heiko Hawlisch, Lihua Wang, Jennifer Best, Nancy S. Herman, Alyssa A. Sproles, Jörg Zwirner, Jeffrey A. Whitsett, Craig Gerard, Georgia Sfyroera, John D. Lambris, Marsha Wills-Karp
Abstract
Mucoid, mucA mutant Pseudomonas aeruginosa cause chronic lung infections in cystic fibrosis (CF) patients and are refractory to phagocytosis and antibiotics. Here we show that mucoid bacteria perish during anaerobic exposure to 15 mM nitrite (NO2–) at pH 6.5, which mimics CF airway mucus. Killing required a pH lower than 7, implicating formation of nitrous acid (HNO2) and NO, that adds NO equivalents to cellular molecules. Eighty-seven percent of CF isolates possessed mucA mutations and were killed by HNO2 (3-log reduction in 4 days). Furthermore, antibiotic-resistant strains determined were also equally sensitive to HNO2. More importantly, HNO2 killed mucoid bacteria (a) in anaerobic biofilms; (b) in vitro in ultrasupernatants of airway secretions derived from explanted CF patient lungs; and (c) in mouse lungs in vivo in a pH-dependent fashion, with no organisms remaining after daily exposure to HNO2 for 16 days. HNO2 at these levels of acidity and NO2– also had no adverse effects on cultured human airway epithelia in vitro. In summary, selective killing by HNO2 may provide novel insights into the important clinical goal of eradicating mucoid P. aeruginosa from the CF airways.
Authors
Sang Sun Yoon, Ray Coakley, Gee W. Lau, Sergei V. Lymar, Benjamin Gaston, Ahmet C. Karabulut, Robert F. Hennigan, Sung-Hei Hwang, Garry Buettner, Michael J. Schurr, Joel E. Mortensen, Jane L. Burns, David Speert, Richard C. Boucher, Daniel J. Hassett
Copyright © 2020 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)