The mechanisms underlying the development and natural progression of the airway mucus defect in cystic fibrosis (CF) remain largely unclear. New animal models of CF, coupled with imaging using micro-optical coherence tomography, can lead to insights regarding these questions. The Cftr–/– (KO) rat allows for longitudinal examination of the development and progression of airway mucus abnormalities. The KO rat exhibits decreased periciliary depth, hyperacidic pH, and increased mucus solid content percentage; however, the transport rates and viscoelastic properties of the mucus are unaffected until the KO rat ages. Airway submucosal gland hypertrophy develops in the KO rat by 6 months of age. Only then does it induce increased mucus viscosity, collapse of the periciliary layer, and delayed mucociliary transport; stimulation of gland secretion potentiates this evolution. These findings could be reversed by bicarbonate repletion but not pH correction without counterion donation. These studies demonstrate that abnormal surface epithelium in CF does not cause delayed mucus transport in the absence of functional gland secretions. Furthermore, abnormal bicarbonate transport represents a specific target for restoring mucus clearance, independent of effects on periciliary collapse. Thus, mature airway secretions are required to manifest the CF defect primed by airway dehydration and bicarbonate deficiency.
Susan E. Birket, Joy M. Davis, Courtney M. Fernandez, Katherine L. Tuggle, Ashley M. Oden, Kengyeh K. Chu, Guillermo J. Tearney, Michelle V. Fanucchi, Eric J. Sorscher, Steven M. Rowe
Adaptation to air breathing after birth is dependent upon the synthesis and secretion of pulmonary surfactant by alveolar type 2 (AT2) cells. Surfactant, a complex mixture of phospholipids and proteins, is secreted into the alveolus, where it reduces collapsing forces at the air-liquid interface to maintain lung volumes during the ventilatory cycle. ABCA3, an ATP-dependent Walker domain containing transport protein, is required for surfactant synthesis and lung function at birth. Mutations in ABCA3 cause severe surfactant deficiency and respiratory failure in newborn infants. We conditionally deleted the Abca3 gene in AT2 cells in the mature mouse lung. Loss of ABCA3 caused alveolar cell injury and respiratory failure. ABCA3-related lung dysfunction was associated with surfactant deficiency, inflammation, and alveolar-capillary leak. Extensive but incomplete deletion of ABCA3 caused alveolar injury and inflammation, and it initiated proliferation of progenitor cells, restoring ABCA3 expression, lung structure, and function. M2-like macrophages were recruited to sites of AT2 cell proliferation during the regenerative process and were present in lung tissue from patients with severe lung disease caused by mutations in ABCA3. The remarkable and selective regeneration of ABCA3-sufficient AT2 progenitor cells provides plausible approaches for future correction of ABCA3 and other genetic disorders associated with surfactant deficiency and acute interstitial lung disease.
Tara N. Rindler, Courtney A. Stockman, Alyssa L. Filuta, Kari M. Brown, John M. Snowball, Wenjia Zhou, Ruud Veldhuizen, Erika M. Zink, Sydney E. Dautel, Geremy Clair, Charles Ansong, Yan Xu, James P. Bridges, Jeffrey A. Whitsett
The neutrophil chemoattractant proline-glycine-proline (PGP) is generated from collagen by matrix metalloproteinase-8/9 (MMP-8/9) and prolyl endopeptidase (PE), and it is concomitantly degraded by extracellular leukotriene A4 hydrolase (LTA4H) to limit neutrophilia. Components of cigarette smoke can acetylate PGP, yielding a species (AcPGP) that is resistant to LTA4H-mediated degradation and can, thus, support a sustained neutrophilia. In this study, we sought to elucidate if an antiinflammatory system existed to degrade AcPGP that is analogous to the PGP-LTA4H axis. We demonstrate that AcPGP is degraded through a previously unidentified action of the enzyme angiotensin-converting enzyme (ACE). Pulmonary ACE is elevated during episodes of acute inflammation, as a consequence of enhanced vascular permeability, to ensure the efficient degradation of AcPGP. Conversely, we suggest that this pathway is aberrant in chronic obstructive pulmonary disease (COPD) enabling the accumulation of AcPGP. Consequently, we identify a potentially novel protective role for AcPGP in limiting pulmonary fibrosis and suggest the pathogenic function attributed to ACE in idiopathic pulmonary fibrosis (IPF) to be a consequence of overzealous AcPGP degradation. Thus, AcPGP seemingly has very divergent roles: it is pathogenic in its capacity to drive neutrophilic inflammation and matrix degradation in the context of COPD, but it is protective in its capacity to limit fibrosis in IPF.
Philip J. O’Reilly, Qiang Ding, Samia Akthar, Guoqiang Cai, Kristopher R. Genschmer, Dhiren F. Patel, Patricia L. Jackson, Liliana Viera, Mojtaba Roda, Morgan L. Locy, Ellen A. Bernstein, Clare M. Lloyd, Kenneth E. Bernstein, Robert J. Snelgrove, J. Edwin Blalock
Expansion of novel therapeutics to all patients with cystic fibrosis (CF) requires personalized CFTR modulator therapy. We have developed nasospheroids, a primary cell culture–based model derived from individual CF patients and healthy subjects by a minimally invasive nasal biopsy. Confocal microscopy was utilized to measure CFTR activity by analyzing changes in cross-sectional area over time that resulted from CFTR-mediated ion and fluid movement. Both the rate of change over time and AUC were calculated. Non-CF nasospheroids with active CFTR-mediated ion and fluid movement showed a reduction in cross-sectional area, whereas no changes were observed in CF spheroids. Non-CF spheroids treated with CFTR inhibitor lost responsiveness for CFTR activation. However, nasospheroids from F508del CF homozygotes that were treated with lumacaftor and ivacaftor showed a significant reduction in cross-sectional area, indicating pharmacologic rescue of CFTR function. This model employs a simple measurement of size corresponding to changes in CFTR activity and is applicable for detection of small changes in CFTR activity from individual patients in vitro. Advancements of this technique will provide a robust model for individualized prediction of CFTR modulator efficacy.
Jennifer S. Guimbellot, Justin M. Leach, Imron G. Chaudhry, Nancy L. Quinney, Susan E. Boyles, Michael Chua, Inmaculada Aban, Ilona Jaspers, Martina Gentzsch
Oxidative stress is important in the pathogenesis of allergic asthma. Extracellular superoxide dismutase (EC-SOD; SOD3) is the major antioxidant in lungs, but its role in allergic asthma is unknown. Here we report that asthmatics have increased SOD3 transcript levels in sputum and that a single nucleotide polymorphism (SNP) in SOD3 (R213G; rs1799895) changes lung distribution of EC-SOD, and decreases likelihood of asthma-related symptoms. Knockin mice analogous to the human R213G SNP had lower airway hyperresponsiveness, inflammation, and mucus hypersecretion with decreased interleukin-33 (IL-33) in bronchoalveolar lavage fluid and reduced type II innate lymphoid cells (ILC2s) in lungs. SOD mimetic (Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin) attenuated Alternaria-induced expression of IL-33 and IL-8 release in BEAS-2B cells. These results suggest that R213G SNP potentially benefits its carriers by resulting in high EC-SOD in airway-lining fluid, which ameliorates allergic airway inflammation by dampening the innate immune response, including IL-33/ST2–mediated changes in ILC2s.
Rohit Gaurav, Jason T. Varasteh, Michael R. Weaver, Sean R. Jacobson, Laura Hernandez-Lagunas, Qing Liu, Eva Nozik-Grayck, Hong Wei Chu, Rafeul Alam, Børge G. Nordestgaard, Camilla J. Kobylecki, Shoaib Afzal, Geoffrey L. Chupp, Russell P. Bowler
Respiratory syncytial virus (RSV) infects almost all infants by 2 years of age, and severe bronchiolitis resulting from RSV infection is the primary cause of hospitalization in the first year of life. Among infants hospitalized due to RSV-induced bronchiolitis, those with a specific mutation in the chemokine receptor CX3CR1, which severely compromises binding of its ligand CX3CL1, were at a higher risk for more severe viral bronchiolitis than those without the mutation. Here, we show that RSV infection of newborn mice deficient in CX3CR1 leads to significantly greater neutrophilic inflammation in the lungs, accompanied by an increase in mucus production compared with that induced in WT mice. Analysis of innate and adaptive immune responses revealed an early increase in the number of IL-17+ γδ T cells in CX3CR1-deficient mice that outnumbered IFN-γ+ γδ T cells as well as IFN-γ+ NK cells, IFN-γ being host protective in the context of RSV infection. This bias toward IL-17+ γδ T cells persisted at a later time. The lungs of CX3CR1-deficient mice expressed higher levels of IL-1β mRNA and protein, and blockade of IL-1β signaling using IL-1 receptor antagonist significantly reduced the number of IL-17+ γδ T cells in the lungs of infected mice. Blockade of IL-17RC abolished RSV-induced lung pathology in infected CX3CR1-deficient mice. We propose that, in infants harboring mutant CX3CR1, targeting the IL-17R may minimize disease severity and hospitalization in early life.
Sudipta Das, Mahesh Raundhal, Jie Chen, Timothy B. Oriss, Rachael Huff, John V. Williams, Anuradha Ray, Prabir Ray
BACKGROUND. In obese subjects with obstructive sleep apnea (OSA), chronic intermittent hypoxia (CIH) may be linked to systemic and adipose tissue inflammation. METHODS. We obtained abdominal subcutaneous adipose tissue biopsies from OSA and non-OSA obese (BMI > 35) subjects at baseline and after 24 weeks (T1) of weight-loss intervention plus continuous positive airway pressure (c-PAP) or weight-loss intervention alone, respectively. OSA subjects were grouped according to good (therapeutic) or poor (subtherapeutic) adherence to c-PAP. RESULTS. At baseline, anthropometric and metabolic parameters, serum cytokines, and adipose tissue mRNA levels of obesity-associated chemokines and inflammatory markers were not different in OSA and non-OSA subjects. At T1, body weight was significantly reduced in all groups. Serum concentrations of IL-2, IL-4, IL-6, MCP-1, PDGFβ, and VEGFα were reduced by therapeutic c-PAP in OSA subjects and remained unaltered in non-OSA and subtherapeutic c-PAP groups. Similarly, adipose tissue mRNA levels of macrophage-specific (CD68, CD36) and ER stress (ATF4, CHOP, ERO-1) gene markers, as well as of IL-6, PDGFβ, and VEGFα, were decreased only in the therapeutic c-PAP group. CONCLUSION. CIH does not represent an additional factor increasing systemic and adipose tissue inflammation in morbid obesity. However, in subjects with OSA, an effective c-PAP therapy improves systemic and obesity-associated inflammatory markers. FUNDING. Ministero dell’Università e della Ricerca and Progetti di Rilevante Interesse Nazionale.
Sebastio Perrini, Angelo Cignarelli, Vitaliano Nicola Quaranta, Vito Antonio Falcone, Stella Kounaki, Stefania Porro, Alessandro Ciavarella, Romina Ficarella, Maria Barbaro, Valentina Annamaria Genchi, Pasquale Nigro, Pierluigi Carratù, Annalisa Natalicchio, Luigi Laviola, Onofrio Resta, Francesco Giorgino
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.
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
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.
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
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.
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
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