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.
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
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.
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
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.
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
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
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
Fibrotic lung disease, most notably idiopathic pulmonary fibrosis (IPF), is thought to result from aberrant wound-healing responses to repetitive lung injury. Increased vascular permeability is a cardinal response to tissue injury, but whether it is mechanistically linked to lung fibrosis is unknown. We previously described a model in which exaggeration of vascular leak after lung injury shifts the outcome of wound-healing responses from normal repair to pathological fibrosis. Here we report that the fibrosis produced in this model is highly dependent on thrombin activity and its downstream signaling pathways. Direct thrombin inhibition with dabigatran significantly inhibited protease-activated receptor-1 (PAR1) activation, integrin αvβ6 induction, TGF-β activation, and the development of pulmonary fibrosis in this vascular leak–dependent model. We used a potentially novel imaging method — ultashort echo time (UTE) lung magnetic resonance imaging (MRI) with the gadolinium-based, fibrin-specific probe EP-2104R — to directly visualize fibrin accumulation in injured mouse lungs, and to correlate the antifibrotic effects of dabigatran with attenuation of fibrin deposition. We found that inhibition of the profibrotic effects of thrombin can be uncoupled from inhibition of hemostasis, as therapeutic anticoagulation with warfarin failed to downregulate the PAR1/αvβ6/TGF-β axis or significantly protect against fibrosis. These findings have direct and important clinical implications, given recent findings that warfarin treatment is not beneficial in IPF, and the clinical availability of direct thrombin inhibitors that our data suggest could benefit these patients.
Barry S. Shea, Clemens K. Probst, Patricia L. Brazee, Nicholas J. Rotile, Francesco Blasi, Paul H. Weinreb, Katharine E. Black, David E. Sosnovik, Elizabeth M. Van Cott, Shelia M. Violette, Peter Caravan, Andrew M. Tager
Idiopathic pulmonary fibrosis (IPF) is a life-threatening disease without effective treatment, highlighting the need for identifying new targets and treatment modalities. The pathogenesis of IPF is complex, and engaging multiple targets simultaneously might improve therapeutic efficacy. To assess the role of the endocannabinoid/cannabinoid receptor 1 (endocannabinoid/CB1R) system in IPF and its interaction with inducible nitric oxide synthase (iNOS) as dual therapeutic targets, we analyzed lung fibrosis and the status of the endocannabinoid/CB1R system and iNOS in mice with bleomycin-induced pulmonary fibrosis (PF) and in lung tissue and bronchoalveolar lavage fluid (BALF) from patients with IPF, as well as controls. In addition, we investigated the antifibrotic efficacy in the mouse PF model of an orally bioavailable and peripherally restricted CB1R/iNOS hybrid inhibitor. We report that increased activity of the endocannabinoid/CB1R system parallels disease progression in the lungs of patients with idiopathic PF and in mice with bleomycin-induced PF and is associated with increased tissue levels of interferon regulatory factor-5. Furthermore, we demonstrate that simultaneous engagement of the secondary target iNOS by the hybrid CB1R/iNOS inhibitor has greater antifibrotic efficacy than inhibition of CB1R alone. This hybrid antagonist also arrests the progression of established fibrosis in mice, thus making it a viable candidate for future translational studies in IPF.
Resat Cinar, Bernadette R. Gochuico, Malliga R. Iyer, Tony Jourdan, Tadafumi Yokoyama, Joshua K. Park, Nathan J. Coffey, Hadass Pri-Chen, Gergő Szanda, Ziyi Liu, Ken Mackie, William A. Gahl, George Kunos
Influenza A virus (IAV) infections lead to severe inflammation in the airways. Patients with chronic obstructive pulmonary disease (COPD) characteristically have exaggerated airway inflammation and are more susceptible to infections with severe symptoms and increased mortality. The mechanisms that control inflammation during IAV infection and the mechanisms of immune dysregulation in COPD are unclear. We found that IAV infections lead to increased inflammatory and antiviral responses in primary bronchial epithelial cells (pBECs) from healthy nonsmoking and smoking subjects. In pBECs from COPD patients, infections resulted in exaggerated inflammatory but deficient antiviral responses. A20 is an important negative regulator of NF-κB–mediated inflammatory but not antiviral responses, and A20 expression was reduced in COPD. IAV infection increased the expression of miR-125a or -b, which directly reduced the expression of A20 and mitochondrial antiviral signaling (MAVS), and caused exaggerated inflammation and impaired antiviral responses. These events were replicated in vivo in a mouse model of experimental COPD. Thus, miR-125a or -b and A20 may be targeted therapeutically to inhibit excessive inflammatory responses and enhance antiviral immunity in IAV infections and in COPD.
Alan C-Y. Hsu, Kamal Dua, Malcolm R. Starkey, Tatt-Jhong Haw, Prema M. Nair, Kristy Nichol, Nathan Zammit, Shane T. Grey, Katherine J. Baines, Paul S. Foster, Philip M. Hansbro, Peter A. Wark
In cystic fibrosis (CF), airway mucus becomes thick and viscous, and its clearance from the airways is impaired. The gel-forming mucins undergo an ordered “unpacking/maturation” process after granular release that requires an optimum postsecretory environment, including hydration and pH. We hypothesized that this unpacking process is compromised in the CF lung due to abnormal transepithelial fluid transport that reduces airway surface hydration and alters ionic composition. Using human tracheobronchial epithelial cells derived from non-CF and CF donors and mucus samples from human subjects and domestic pigs, we investigated the process of postsecretory mucin unfolding/maturation, how these processes are defective in CF airways, and the probable mechanism underlying defective unfolding. First, we found that mucins released into a normal lung environment transform from a compact granular form to a linear form. Second, we demonstrated that this maturation process is defective in the CF airway environment. Finally, we demonstrated that independent of HCO3− and pH levels, airway surface dehydration was the major determinant of this abnormal unfolding process. This defective unfolding/maturation process after granular release suggests that the CF extracellular environment is ion/water depleted and likely contributes to abnormal mucus properties in CF airways prior to infection and inflammation.
Lubna H. Abdullah, Jessica R. Evans, T. Tiffany Wang, Amina A. Ford, Alexander M. Makhov, Kristine Nguyen, Raymond D. Coakley, Jack D. Griffith, C. William Davis, Stephen T. Ballard, Mehmet Kesimer
Accumulating evidence suggests that altered cellular metabolism is systemic in pulmonary hypertension (PH) and central to disease pathogenesis. However, bioenergetic changes in PH patients and their association with disease severity remain unclear. Here, we hypothesize that alteration in bioenergetic function is present in platelets from PH patients and correlates with clinical parameters of PH. Platelets isolated from controls and PH patients (
Quyen L. Nguyen, Catherine Corey, Pamela White, Annie Watson, Mark T. Gladwin, Marc A. Simon, Sruti Shiva
Ciliary motion defects cause defective mucociliary transport (MCT) in primary ciliary dyskinesia (PCD). Current diagnostic tests do not assess how MCT is affected by perturbation of ciliary motion. In this study, we sought to use micro-optical coherence tomography (μOCT) to delineate the mechanistic basis of cilia motion defects of PCD genes by functional categorization of cilia motion. Tracheae from three PCD mouse models were analyzed using μOCT to characterize ciliary motion and measure MCT. We developed multiple measures of ciliary activity, integrated these measures, and quantified dyskinesia by the angular range of the cilia effective stroke (ARC).
George M. Solomon, Richard Francis, Kengyeh K. Chu, Susan E. Birket, George Gabriel, John E. Trombley, Kristi L. Lemke, Nikolai Klena, Brett Turner, Guillermo J. Tearney, Cecilia W. Lo, Steven M. Rowe
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