Pulmonology
Abstract
BACKGROUND. Information about the size, airway location, and longitudinal behavior of mucus plugs in asthma is needed to understand their role in mechanisms of airflow obstruction and to rationally design muco-active treatments. METHODS. Computed tomography (CT) lung scans from 57 asthma patients were analyzed to quantify mucus plug size and airway location, and paired CT scans obtained 3 years apart were analyzed to determine plug behavior over time. Radiologist annotations of mucus plugs were incorporated in an image-processing pipeline to generate size and location information that was related to measures of airflow. RESULTS. The length distribution of 778 annotated mucus plugs was multimodal and a 12 mm length defined short (“stubby”, ≤12 mm) and long (“stringy”, >12 mm) plug phenotypes. High mucus plug burden was disproportionately attributable to stringy mucus plugs. Mucus plugs localized predominantly to airway generations 6 to 9, and 47% of plugs in baseline scans, persisted in the same airway for three years, and fluctuated in length and volume. Mucus plugs in larger proximal generations had greater effects on spirometry measures than plugs in smaller distal generations, and a model of airflow that estimates the increased airway resistance attributable to plugs predicted higher impact for proximal and more numerous mucus plugs. CONCLUSIONS. Persistent mucus plugs in proximal airway generations occur in asthma and demonstrate a stochastic process of formation and resolution over time. Proximal airway mucus plugs are consequential for airflow and are in locations amenable to treatment by inhaled muco-active drugs or bronchoscopy. TRIAL REGISTRATION. Clinicaltrials.gov NCT01718197, NCT01606826, NCT01750411, NCT01761058, NCT01761630, NCT01759186, NCT01716494, and NCT01760915 FUNDING. NIH Grants: R01 HL080414, UG1 HL139106, P01 HL107202, U01 HL146002, U10 HL109172, U10 HL109168, U10 HL109152, U10 HL109257, U10 HL109146, U10 HL109250, U10 HL109164, U10 109086, and T32 HL007185, F32 HL162422. The following companies provided financial support for study activities at the Coordinating and Clinical Centers beyond the third year of patient follow-up: AstraZeneca, Boehringer-Ingelheim, Genentech, GlaxoSmithKline, Sanofi–Genzyme– Regeneron, and TEVA. These companies had no role in study design or data analysis, and the only restriction on the funds was that they be used to support the SARP initiative.
Authors
Brendan K. Huang, Brett M. Elicker, Travis S. Henry, Kimberly G. Kallianos, Lewis D. Hahn, Monica Tang, Franklin Heng, Charles E. McCulloch, Nirav R. Bhakta, Sharmila Majumdar, Jiwoong Choi, Loren C. Denlinger, Sean B. Fain, Annette T. Hastie, Eric A. Hoffman, Elliot Israel, Nizar N. Jarjour, Bruce D. Levy, David T. Mauger, Kaharu Sumino, Sally E. Wenzel, Mario Castro, Prescott G. Woodruff, John V. Fahy
Abstract
Infection of immature mice with rhinovirus (RV) induces an asthma-like phenotype consisting of type 2 inflammation, mucous metaplasia, eosinophilic inflammation and airways hyperresponsiveness which is dependent on IL-25 and type 2 innate lymphoid cells (ILC2s). Doublecortin-like kinase (DCLK)-1+ tuft cells are a major source of IL-25. We sought to determine the requirement of tuft cells for the RV-induced asthma phenotype in wild-type mice and mice deficient in Pou2f3, a transcription factor required for tuft cell development. C57Bl/6 mice infected with RV-A1B on day 6 of life and RV-A2 on day 13 of life showed increased DCLK1+ positive tuft cells in the large airways. Compared to wild-type mice, RV-infected Pou2f3–/– mice showed reductions in IL-25 mRNA and protein expression, ILC2 expansion, type 2 cytokine expression, mucous metaplasia, lung eosinophils and airway methacholine responsiveness. We conclude that airway tuft cells are required for the asthma phenotype observed in immature mice undergoing repeated RV infections. Furthermore, RV-induced tuft cell development provides a mechanism by which early life viral infections could potentiate type 2 inflammatory responses to future infections.
Authors
Yiran Li, Mingyuan Han, Shilpi Singh, Haley A. Breckenridge, Jordan E. Kreger, Claudia C. Stroupe, Daniel A. Sawicky, Shiuhyang Kuo, Adam M. Goldsmith, Fang Ke, Anukul T. Shenoy, J. Kelley Bentley, Ichiro Matsumoto, Marc B. Hershenson
Abstract
Cigarette smoking is associated with a higher risk of ICU admissions among flu patients. However, the etiological mechanism by which cigarette smoke (CS) exacerbates flu remains poorly understood. Here, we show that a mild dose of influenza A virus promotes a severe lung injury in mice pre-exposed to CS but not room air for four weeks. Real-time intravital (in vivo) lung imaging revealed that the development of acute severe respiratory dysfunction in CS and flu exposed mice was associated with the accumulation of platelet-rich neutrophil-platelet aggregates (NPAs) in the lung microcirculation within 2 days following flu infection. These platelet-rich NPAs formed in situ and grew larger over time to occlude the lung microvasculature, leading to the development of pulmonary ischemia followed by the infiltration of NPAs and vascular leakage into the alveolar air space. These findings suggest for the first time that an acute onset of platelet-driven thrombo-inflammatory response in the lung contributes to the development of CS induced severe flu.
Authors
Tomasz W. Kaminski, Tomasz Brzoska, Xiuying Li, Ravi Vats, Omika Katoch, Rikesh K. Dubey, Kamal Bagale, Simon C. Watkins, Bryan J. McVerry, Tirthadipa Pradhan-Sundd, Lianghui Zhang, Keven M. Robinson, Toru Nyunoya, Prithu Sundd
Abstract
Pulmonary fibrosis is a chronic and often fatal disease. The pathogenesis is characterized by aberrant repair of lung parenchyma resulting in loss of physiological homeostasis, respiratory failure and death. The immune response in pulmonary fibrosis is dysregulated. The gut microbiome is a key regulator of immunity. The role of the gut microbiome in regulating the pulmonary immunity in lung fibrosis is poorly understood. Here, we determine the impact of gut microbiota on pulmonary fibrosis in C57BL/6 mice derived from different vendors (C57BL/6J and C57BL/6NCrl). We use germ free models, fecal microbiota transplantation and cohousing to transmit gut microbiota. Metagenomic studies of feces establish keystone species between sub-strains. Pulmonary fibrosis is microbiota dependent in C57BL/6 mice. Gut microbiota are distinct by β diversity (PERMANOVA P<0.001) and α diversity (P<0.0001). Mortality and lung fibrosis are attenuated in C57BL/6NCrl mice. Elevated CD4+ IL-10+ T cells and lower IL-6 occur in C57BL/6NCrl mice. Horizontal transmission of microbiota by cohousing attenuates mortality in C57BL/6J mice and promotes a transcriptionally altered pulmonary immunity. Temporal changes in lung and gut microbiota demonstrates that gut microbiota contribute largely to immunological phenotype. Key regulatory gut microbiota contribute to lung fibrosis generating rationale for human studies.
Authors
Stephen J. Gurczynski, Jay H. Lipinski, Joshua Y. Strauss, Shafiul Alam, Gary B. Huffnagle, Piyush Ranjan, Lucy H. Kennedy, Bethany B. Moore, David N. O'Dwyer
Abstract
The lymphatic vasculature is the natural pathway for the resolution of inflammation, while the role of pulmonary lymphatic drainage function in sepsis-induced acute respiratory distress syndrome (ARDS) remains poorly characterized. In this study, Indocyanine green (ICG)-Near Infrared (NIR) lymphatic living imaging was performed to examine pulmonary lymphatic drainage function in septic mice models. We found that the pulmonary lymphatic drainage was impaired owing to the damaged lymphatic structure in sepsis-induced ARDS. Moreover, prior lymphatic defects by blocking vascular endothelial growth factor receptor-3 (VEGFR3), worsened sepsis-induced lymphatic dysfunction and inflammation. The post-treatment of vascular endothelial growth factor-C (Cys156Ser) (VEGF-C156S), a ligand of VEGFR3, ameliorated lymphatic drainage through rejuvenating lymphatics to reduce the pulmonary edema and promote pulmonary macrophages and neutrophils to drain to pretracheal lymph nodes (pLNs). Meanwhile, VEGF-C156S post-treatment reversed sepsis-inhibited C-C motif chemokine ligand 21 (CCL21), which co-localizes with the pulmonary lymphatic vessels. Furthermore, the advantages of VEGF-C156S on the drainage of inflammatory cells and edema fluid were abolished by blocking VEGFR3 or CCL21. These results suggest that efficient pulmonary lymphatic drainage is necessary for inflammation resolution in ARDS. Our findings offer a novel therapeutic approach to sepsis-induced ARDS by promoting lymphatic drainage function.
Authors
Pu-hong Zhang, Wen-wu Zhang, Shun-shun Wang, Cheng-hua Wu, Yang-dong Ding, Xin-yi Wu, Fang Gao Smith, Yu Hao, Sheng-wei Jin
Abstract
Chronic lung allograft dysfunction (CLAD) is a major complication after lung transplantation that results from a complex interplay of innate inflammatory and alloimmune factors, culminating in parenchymal and/or obliterative airway fibrosis. Excessive IL-17A signaling and chronic inflammation have been recognized as key factors in these pathological processes. Herein, we developed a model of repeated airway inflammation in mouse minor alloantigen-mismatched single-lung transplantation. Repeated intratracheal LPS instillations augmented pulmonary IL-17A expression. LPS also increased acute rejection, airway epithelial damage, and obliterative airway fibrosis, similar to human explanted lung allografts with antecedent episodes of airway infection. We then investigated the role of donor and recipient IL-17 receptor A (IL-17RA) in this context. Donor IL-17RA deficiency significantly attenuated acute rejection and CLAD features, whereas recipient IL-17RA deficiency only slightly reduced airway obliteration in LPS allografts. IL-17RA immunofluorescence positive staining was greater in human CLAD lungs compared with control human lung specimens, with localization to fibroblasts and myofibroblasts, which was also seen in mouse LPS allografts. Taken together, repeated airway inflammation after lung transplantation caused local airway epithelial damage, with persistent elevation of IL-17A and IL-17RA expression and particular involvement of IL-17RA on donor structural cells in development of fibrosis.
Authors
Tatsuaki Watanabe, Stephen C. Juvet, Gregory Berra, Jan Havlin, Wenshan Zhong, Kristen Boonstra, Tina Daigneault, Miho Horie, Chihiro Konoeda, Grace Teskey, Zehong Guan, David M. Hwang, Mingyao Liu, Shaf Keshavjee, Tereza Martinu
Abstract
Idiopathic Pulmonary Fibrosis (IPF) is a chronic parenchymal lung disease characterized by repetitive alveolar cell injury, myofibroblast proliferation, and excessive extracellular matrix deposition for which unmet need persists for effective therapeutics. The bioactive eicosanoid, prostaglandin F2a, and its cognate receptor FPr (Ptfgr) are implicated as a TGFβ1 independent signaling hub for IPF. To assess this, we leveraged our published murine PF model (IER-SftpcI73T) expressing a disease-associated missense mutation in the surfactant protein C (Sftpc) gene. Tamoxifen treated IER-SftpcI73T mice develop an early multiphasic alveolitis and transition to spontaneous fibrotic remodeling by 28 days. IER-SftpcI73T mice crossed to a Ptgfr null (FPr-/-) line showed attenuated weight loss and gene dosage dependent rescue of mortality compared to FPr+/+ cohorts. IER-SftpcI73T/FPr-/- mice also showed reductions in multiple fibrotic endpoints for which administration of nintedanib was not additive. Single cell RNA sequencing, pseudotime analysis, and in vitro assays demonstrated Ptgfr expression predominantly within adventitial fibroblasts which were reprogrammed to an “inflammatory/transitional” cell state in a PGF2a/ FPr dependent manner. Collectively, the findings provide evidence for a role for PGF2a signaling in IPF, mechanistically identify a susceptible fibroblast subpopulation, and establish a benchmark effect size for disruption of this pathway in mitigating fibrotic lung remodeling.
Authors
Luis R. Rodriguez, Soon Yew Tang, Willy Roque Barboza, Aditi Murthy, Yaniv Tomer, Tian-Quan Cai, Swati Iyer, Katrina Chavez, Ujjalkumar Subhash Das, Soumita Ghosh, Charlotte Cooper, Thalia T. Dimopoulos, Apoorva Babu, Caitlin F. Connelly, Garret A. FitzGerald, Michael F. Beers
Abstract
Aberrant fibroblast function plays a key role in the pathogenesis of Idiopathic Pulmonary Fibrosis, a devastating disease of unrelenting extracellular matrix deposition in response to lung injury. Platelet-derived growth factor alpha-positive (PDGFRα+) lipofibroblasts (LipoFBs) are essential for lung injury response and maintenance of a functional alveolar stem cell niche. Little is known about the effects of lung injury on LipoFB function. Here, we used scRNA-Seq technology and PDGFRαGFP lineage tracing to generate a transcriptomic profile of PDGFRα+ fibroblasts in normal and injured mouse lungs 14 days after bleomycin exposure, generating eleven unique transcriptomic clusters that segregated according to treatment. While normal and injured LipoFBs shared a common gene signature, injured LipoFBs acquired fibrogenic pathway activity with an attenuation of lipogenic pathways. In a 3D organoid model, injured PDGFRα+ fibroblast- supported organoids were morphologically distinct from those cultured with normal FBs, and scRNA-Seq analysis suggested distinct transcriptomic changes in alveolar epithelia supported by injured PDGFRα+ fibroblasts. In summary, while LipoFBs in injured lung have not migrated from their niche and retain their lipogenic identity, they acquire a potentially reversible fibrogenic profile, which may alter the kinetics of epithelial regeneration and potentially contribute to dysregulated repair, leading to fibrosis.
Authors
Carol S. Trempus, Brian N. Papas, Maria I. Sifre, Carl D. Bortner, Erica Scappini, Charles J. Tucker, Xin Xu, Katina L. Johnson, Leesa J. Deterding, Jason G Williams, Dylan J. Johnson, Jian-Liang Li, Deloris Sutton, Charan K. Ganta, Debabrata Mahapatra, Muhammad Arif, Abhishek Basu, Lenny Pommerolle, Resat Cinar, Anne-Karina T. Perl, Stavros Garantziotis
Abstract
Primary graft dysfunction (PGD) limits clinical benefit after lung transplantation, a life-prolonging therapy for patients with end-stage disease. PGD is the clinical syndrome resulting from pulmonary ischemia-reperfusion injury (IRI), driven by innate immune inflammation. We recently demonstrated a key role for NK cells in the airways of mouse models and human tissue samples of IRI. Here we used 2 mouse models paired with human lung transplant samples to investigate the mechanisms whereby NK cells migrate to the airways to mediate lung injury. We demonstrate that chemokine receptor ligand transcripts and proteins are increased in mouse and human disease. CCR5 ligand transcripts were correlated with NK cell gene signatures independent of NK cell CCR5 ligand secretion. NK cells expressing CCR5 were increased in the lung and airways during IRI and had increased markers of tissue residency and maturation. Allosteric CCR5 drug blockade reduced the migration of NK cells to the site of injury. CCR5 blockade also blunted quantitative measures of experimental IRI. Additionally, in human lung transplant bronchoalveolar lavage samples, we found that CCR5 ligand was associated with increased patient morbidity and that the CCR5 receptor was increased in expression on human NK cells following PGD. These data support a potential mechanism for NK cell migration during lung injury and identify a plausible preventative treatment for PGD.
Authors
Jesse Santos, Ping Wang, Avishai Shemesh, Fengchun Liu, Tasha Tsao, Oscar A. Aguilar, Simon J. Cleary, Jonathan P. Singer, Ying Gao, Steven R. Hays, Jeffrey Golden, Lorriana E. Leard, Mary Ellen Kleinhenz, Nicholas A. Kolaitis, Rupal J. Shah, Aida Venado, Jasleen Kukreja, S. Sam Weigt, John A. Belperio, Lewis L. Lanier, Mark R. Looney, John R. Greenland, Daniel R. Calabrese
Abstract
Lung contusion and gastric aspiration (LC and GA) are major risk factors for developing acute respiratory distress following trauma. Hypoxia from lung injury is mainly regulated by hypoxia-inducible factor 1α (HIF-1α). Published data from our group indicate that HIF-1α regulation in airway epithelial cells (AEC) drives the acute inflammatory response following LC and GA. Metabolomic profiling and metabolic flux of Type II AEC following LC revealed marked increases in glycolytic and TCA intermediates in vivo and in vitro that were HIF-1α dependent. GLUT-1/4 expression was also increased in HIF-1α+/+ mice, suggesting that increased glucose entry may contribute to increased intermediates. Importantly, lactate incubation in vitro on Type II cells did not significantly increase the inflammatory byproduct IL-1β. Contrastingly, succinate had a direct proinflammatory effect on human small AEC by IL-1β generation in vitro. This effect was reversed by dimethylmalonate, suggesting an important role for succinate dehydrogenase in mediating HIF-1α effects. We confirmed the presence of the only known receptor for succinate binding, SUCNR1, on Type II AEC. These results support the hypothesis that succinate drives HIF-1α–mediated airway inflammation following LC. This is the first report to our knowledge of direct proinflammatory activation of succinate in nonimmune cells such as Type II AEC in direct lung injury models.
Authors
Madathilparambil V. Suresh, Sinan Aktay, George Yalamanchili, Sumeet Solanki, Dily Thazhath Sathyarajan, Manikanta Swamy Arnipalli, Subramaniam Pennathur, Krishnan Raghavendran
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