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Pulmonary fibrosis is characterized by the accumulation of myofibroblasts in the lung and progressive tissue scarring. Fibroblasts exist across a spectrum of states, from quiescence in health to activated myofibroblasts in the setting of injury. Highly activated myofibroblasts have a critical role in the establishment of fibrosis as the predominant source of type 1 collagen and profibrotic mediators. Myofibroblasts are also highly contractile cells and can alter lung biomechanical properties through tissue contraction. Inhibiting signaling pathways involved in myofibroblast activation could therefore have significant therapeutic value. One of the ways myofibroblast activation occurs is through activation of the Rho/myocardin-related transcription factor (MRTF)/serum response factor (SRF) pathway, which signals through intracellular actin polymerization. However, concerns surrounding the pleiotropic and ubiquitous nature of these signaling pathways have limited the translation of inhibitory drugs. Herein, we demonstrate a novel therapeutic antifibrotic strategy using myofibroblast-targeted nanoparticles containing a MTRF/SRF pathway inhibitor (CCG-1423), which has been shown to block myofibroblast activation in vitro. Myofibroblasts were preferentially targeted via the angiotensin 2 receptor, which has been shown to be selectively upregulated in animal and human studies. These nanoparticles were nontoxic and accumulated in lung myofibroblasts in the bleomycin-induced mouse model of pulmonary fibrosis, reducing the number of these activated cells and their production of profibrotic mediators. Ultimately, in a murine model of lung fibrosis, a single injection of these drugs containing targeted nanoagents reduced fibrosis as compared with control mice. This approach has the potential to deliver personalized therapy by precisely targeting signaling pathways in a cell-specific manner, allowing increased efficacy with reduced deleterious off-target effects.

Asthma is a chronic disease most commonly associated with allergy and type 2 inflammation. However, the mechanisms that link airway inflammation to the structural changes that define asthma are incompletely understood. Using a human model of allergen-induced asthma exacerbation, we compared the lower airway mucosa in allergic asthmatics and allergic non-asthmatic controls using single-cell RNA sequencing. In response to allergen, the asthmatic airway epithelium was highly dynamic and up-regulated genes involved in matrix degradation, mucus metaplasia, and glycolysis while failing to induce injury-repair and antioxidant pathways observed in controls. IL9 -expressing pathogenic T H 2 cells were specific to asthmatic airways and were only observed after allergen challenge. Additionally, conventional type 2 dendritic cells (DC2 that express CD1C ) and CCR2 -expressing monocyte-derived cells (MCs) were uniquely enriched in asthmatics after allergen, with up-regulation of genes that sustain type 2 inflammation and promote pathologic airway remodeling. In contrast, allergic controls were enriched for macrophage-like MCs that up-regulated tissue repair programs after allergen challenge, suggesting that these populations may protect against asthmatic airway remodeling. Cellular interaction analyses revealed a T H 2–mononuclear phagocyte–basal cell interactome unique to asthmatics. These pathogenic cellular circuits were characterized by type 2 programming of immune and structural cells and additional pathways that may sustain and amplify type 2 signals, including TNF family signaling, altered cellular metabolism, failure to engage antioxidant responses, and loss of growth factor signaling. Our findings therefore suggest that pathogenic effector circuits and the absence of proresolution programs drive structural airway disease in response to type 2 inflammation.

Idiopathic pulmonary fibrosis is a progressive lung disease with limited therapeutic options that is characterized by pathological fibroblast activation and aberrant lung remodeling with scar formation. YAP (Yes-associated protein) is a transcriptional coactivator that mediates mechanical and biochemical signals controlling fibroblast activation. We previously identified HMG-CoA (3-hydroxy-3-methylglutaryl coenzyme A) reductase inhibitors (statins) as YAP inhibitors based on a high-throughput small-molecule screen in primary human lung fibroblasts. Here we report that several Aurora kinase inhibitors were also identified from the top hits of this screen. MK-5108, a highly selective inhibitor for AURKA (Aurora kinase A), induced YAP phosphorylation and cytoplasmic retention and significantly reduced profibrotic gene expression in human lung fibroblasts. The inhibitory effect on YAP nuclear translocation and profibrotic gene expression is specific to inhibition of AURKA, but not Aurora kinase B or C, and is independent of the Hippo pathway kinases LATS1 and LATS2 (Large Tumor Suppressor 1 and 2). Further characterization of the effects of MK-5108 demonstrate that it inhibits YAP nuclear localization indirectly via effects on actin polymerization and TGFβ (Transforming Growth Factor β) signaling. In addition, MK-5108 treatment reduced lung collagen deposition in the bleomycin mouse model of pulmonary fibrosis. Our results reveal a novel role for AURKA in YAP-mediated profibrotic activity in fibroblasts and highlight the potential of small-molecule screens for YAP inhibitors for identification of novel agents with antifibrotic activity.

Smoking and HIV-1 infection are risk factors for COPD, which is among the most common comorbid conditions in people living with HIV-1. HIV-1 infection leads to persistent expansion of CD8+ T cells, and CD8+ T cell-mediated inflammation has been implicated in COPD pathogenesis. In this study, we investigated the effects of HIV-1 infection and smoking on T cell dynamics in patients at risk of COPD. Bronchoalveolar lavage (BAL), endobronchial brushings and blood from HIV-1 infected and uninfected non-smokers and smokers were analyzed by flow cytometry, and lungs were imaged by computed tomography. Chemokines were measured in BAL fluid, and CD8+ T cell chemotaxis in the presence of cigarette smoke extract was assessed in vitro. HIV-1 infection increased CD8+ T cells in the BAL, but this increase was abrogated by smoking. Smokers had reduced BAL levels of the T cell-recruiting chemokines CXCL10 and CCL5, and cigarette smoke extract inhibited CXCL10 and CCL5 production by macrophages and CD8+ T cell transmigration in vitro. In contrast to the BAL, CD8+ T cells in endobronchial brushings were increased in HIV-1 infected smokers, driven by an accumulation of effector memory T cells in the airway mucosa and an increase in tissue resident memory T cells. Mucosal CD8+ T cell numbers inversely correlated with lung aeration, suggesting an association with inflammation and remodeling. HIV-1 infection and smoking lead to retention of CD8+ T cells within the airway mucosa.

Tobacco smoking doubles the risk of active tuberculosis (TB) and accounts for up to 20% of all active TB cases globally. How smoking promotes lung microenvironments permissive toMycobacterium tuberculosis(Mtb) growth remains incompletely understood. We investigated primary bronchoalveolar lavage cells from current- and never-smokers by performing single-cell RNA-sequencing (scRNA-seq), flow cytometry, and functional assays. We observed enrichment of immature inflammatory monocytes in the lungs of smokers compared to non-smokers. These monocytes exhibited phenotypes consistent with recent recruitment from blood, ongoing differentiation, increased activation, and states similar to those with chronic obstructive pulmonary disease (COPD). Using integrative scRNA-seq and flow cytometry, we identify CD93 as a marker for a subset of these newly recruited smoking-associated lung monocytes and further provide evidence that recruitment of monocytes into the lung is mediated by CCL11 binding to CCR2. We also show that these cells exhibit elevated inflammatory responses upon exposure toMtband accelerated intracellular growth ofMtbcompared to mature macrophages. This elevatedMtbgrowth could be inhibited with an anti-inflammatory small molecule, providing a direct connection between smoking-induced pro-inflammatory states and permissiveness toMtbgrowth. Our findings suggest a model in which smoking leads to recruitment of immature inflammatory monocytes from the periphery to the lung via CCL11-CCR2 interactions, which results in the accumulation of theseMtbpermissive cells in the airway. This work defines how smoking may lead to increased susceptibility toMtband identifies novel host-directed therapies to reduce the burden of TB among those who smoke.One Sentence SummaryInflammatory monocytes are recruited to the airways of smokers where they may contribute to more rapid growth ofMycobacterium tuberculosisin the lungs.

Rationale: Standard physiologic assessments of extubation readiness in patients with acute hypoxemic respiratory failure (AHRF) may not reflect lung injury resolution and could adversely affect cli...

BACKGROUND: Vaping, including the use of electronic cigarettes (e-cigarettes), has become increasingly prevalent, yet the associated long-term health risks are largely unknown. Given the prevalence of use, particularly among adolescents early in their lifespan, it is vital to understand the potential chronic pathologic sequelae of vaping. METHODS: We present the cases of four patients with chronic lung disease associated with e-cigarette use characterized by clinical evaluation, with pulmonary function tests (PFTs), chest high-resolution computed tomography (HRCT), endobronchial optical coherence tomography (EB-OCT) imaging, and histopathologic assessment. RESULTS: Each patient presented with shortness of breath and chest pain in association with a 3- to 8-year history of e-cigarette use, with mild progressive airway obstruction on PFTs and/or chest HRCT findings demonstrating evidence of air trapping and bronchial wall thickening. EB-OCT imaging performed in two patients showed small airway–centered fibrosis with bronchiolar narrowing and lumen irregularities. The predominant histopathologic feature on surgical lung biopsy was small airway–centered fibrosis, including constrictive bronchiolitis and MUC5AC overexpression in all patients. Patients who ceased vaping had a partial, but not complete, reversal of disease over 1 to 4 years. CONCLUSIONS: After thorough evaluation for other potential etiologies, vaping was considered to be the most likely common causal etiology for all patients due to the temporal association of symptomatic chronic lung disease with e-cigarette use and partial improvement in symptoms after e-cigarette cessation. In this series, we associate the histopathologic pattern of small airway–centered fibrosis, including constrictive bronchiolitis, with vaping, potentially defining a clinical and pathologic entity associated with e-cigarette use. (Funded in part by the National Institutes of Health.)

Background The aim of this study was to characterize severe immune-related adverse events (irAEs) seen among hospitalized patients and to examine risk factors for irAE admissions and clinically relevant outcomes, including length of stay, immune checkpoint inhibitor (ICI) discontinuation, readmission, and death. Methods Patients who received ICI therapy (ipilimumab, pembrolizumab, nivolumab, atezolizumab, durvalumab, avelumab, or any ICI combination) at Massachusetts General Hospital (MGH) and were hospitalized at MGH following ICI initiation between January 1, 2011, and October 24, 2018, were identified using pharmacy and hospital admission databases. Medical records of all irAE admissions were reviewed, and specialist review with defined criteria was performed. Demographic data, relevant clinical history (malignancy type and most recent ICI regimen), and key admission characteristics, including dates of admission and discharge, immunosuppressive management, ICI discontinuation, readmission, and death, were collected. Results In total, 450 admissions were classified as irAE admissions and represent the study's cohort. Alongside the increasing use of ICIs at our institution, the number of patients admitted to MGH for irAEs has gradually increased every year from 9 in 2011 to 92 in 2018. The hospitalization rate per ICI recipient has declined over that same time period (25.0% in 2011 to 8.5% in 2018). The most common toxicities leading to hospitalization in our cohort were gastrointestinal (30.7%; n = 138), pulmonary (15.8%; n = 71), hepatic (14.2%; n = 64), endocrine (12.2%; n = 55), neurologic (8.4%; n = 38), cardiac (6.7%; n = 30), and dermatologic (4.4%; n = 20). Multivariable logistic regression revealed statistically significant increases in irAE admission risk for CTLA-4 monotherapy recipients (odds ratio [OR], 2.02; p < .001) and CTLA-4 plus PD-1 combination therapy recipients (OR, 1.88; p < .001), relative to PD-1/PD-L1 monotherapy recipients, and patients with multiple toxicity had a 5-fold increase in inpatient mortality. Conclusion This study illustrates that cancer centers must be prepared to manage a wide variety of irAE types and that CTLA-4 and combination ICI regimens are more likely to cause irAE admissions, and earlier. In addition, admissions for patients with multi-organ involvement is common and those patients are at highest risk of inpatient mortality. Implications for Practice The number of patients admitted to Massachusetts General Hospital for immune-related adverse events (irAEs) has gradually increased every year and the most common admissions are for gastrointestinal (30.7%), pulmonary (15/8%), and hepatic (14.2%) events. Readmission rates are high (29% at 30 days, 49% at 180 days) and 64.2% have to permanently discontinue immune checkpoint inhibitor therapy. Importantly, multiple concurrent toxicities were seen in 21.6% (97/450) of irAE admissions and these patients have a fivefold increased risk of inpatient death.

Alveolar macrophages (AMs) are critical for defense against airborne pathogens and AM dysfunction is thought to contribute to the increased burden of pulmonary infections observed in individuals living with HIV-1 (HIV). While HIV nucleic acids have been detected in AMs early in infection, circulating HIV during acute and chronic infection is usually CCR5 T cell-tropic (T-tropic) and enters macrophages inefficiently in vitro. The mechanism by which T-tropic viruses infect AMs remains unknown. We collected AMs by bronchoscopy performed in HIV-infected, antiretroviral therapy (ART)-naive and uninfected subjects. We found that viral constructs made with primary HIV envelope sequences isolated from both AMs and plasma were T-tropic and inefficiently infected macrophages. However, these isolates productively infected macrophages when co-cultured with HIV-infected CD4+ T cells. In addition, we provide evidence that T-tropic HIV is transmitted from infected CD4+ T cells to the AM cytosol. We conclude that AM-derived HIV isolates are T-tropic and can enter macrophages through contact with an infected CD4+ T cell, which results in productive infection of AMs. CD4+ T cell-dependent entry of HIV into AMs helps explain the presence of HIV in AMs despite inefficient cell-free infection, and may contribute to AM dysfunction in people living with HIV.

Foxp3+ regulatory T (Treg) cells expressing the interleukin (IL)-33 receptor ST2 mediate tissue repair in response to IL-33. Whether Treg cells also respond to the alarmin IL-33 to regulate specific aspects of the immune response is not known. Here we describe an unexpected function of ST2+ Treg cells in suppressing the innate immune response in the lung to environmental allergens without altering the adaptive immune response. Following allergen exposure, ST2+ Treg cells were activated by IL-33 to suppress IL-17-producing γδ T cells. ST2 signaling in Treg cells induced Ebi3, a component of the heterodimeric cytokine IL-35 that was required for Treg cell-mediated suppression of γδ T cells. This response resulted in fewer eosinophil-attracting chemokines and reduced eosinophil recruitment into the lung, which was beneficial to the host in reducing allergen-induced inflammation. Thus, we define a fundamental role for ST2+ Treg cells in the lung as a negative regulator of the early innate γδ T cell response to mucosal injury. Luster and colleagues show that Treg cells that reside in lung mucosa can respond to IL-33 upon allergen exposure and suppress innate cell responses. IL-33-activated ST2+ Treg cells secrete IL-35, which suppresses IL-17 production by γδ T cells and lessens eosinophil recruitment into the lung.

Background Dyspnea and exercise intolerance are commonly reported post-acute sequelae of SARS-CoV-2 infection (PASC), but routine diagnostic testing is often normal. Cardiopulmonary exercise testing (CPET) offers comprehensive assessment of dyspnea to characterize pulmonary PASC. Methods We performed a retrospective cohort study of CPET performed on patients reporting dyspnea and/or exercise intolerance following confirmed Covid-19 between August 1, 2020 and March 1, 2021, and compared them to age- and sex-matched patients with unexplained dyspnea referred for CPET at the same center in the pre-Covid-19 era. Findings Compared to matched unexplained dyspnea comparators, PASC patients shared similar medical comorbidities and subjective dyspnea at referral (mMRC score 1.6 ± 0.9 vs. 1.4 ± 0.9, P = 0.5). Fifteen (83.3%) PASC patients underwent high resolution computed tomography of the chest, of which half (46.7%) were normal, and 17 (94.4%) patients had pulmonary function testing, of which the majority (76.5%) were normal. All patients underwent CPET, and 12 (67%) had normal findings. Compared to matched comparators, PASC patients had similar peak oxygen consumption, oxygen consumption at ventilatory anaerobic threshold, and ventilatory efficiency measured by the minute ventilation to carbon dioxide production (VE/VCO2) slope. Interpretation Despite prominent dyspnea, physiological abnormalities on CPET were mild across a range of initial Covid-19 severity and similar to matched comparators referred for dyspnea without antecedent SARS-CoV-2. Funding The project was supported by the NHLBI (R01HL131029, R01HL151841, U10HL110337, T32HL116275) and a KL2 award (5KL2TR002542–02) from Harvard Catalyst.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive disease which leads to significant morbidity and mortality from respiratory failure. The two drugs currently approved for clinical use slow the rate of decline in lung function but have not been shown to halt disease progression or reverse established fibrosis. Thus, new therapeutic targets are needed. Endothelial injury and the resultant vascular permeability are critical components in the response to tissue injury and are present in patients with IPF. However, it remains unclear how vascular permeability affects lung repair and fibrosis following injury. Lipid mediators such as sphingosine-1-phosphate (S1P) are known to regulate multiple homeostatic processes in the lung including vascular permeability. We demonstrate that endothelial cell–(EC) specific deletion of the S1P receptor 1 (S1PR1) in mice (EC-S1pr1(−/−)) results in increased lung vascular permeability at baseline. Following a low-dose intratracheal bleomycin challenge, EC-S1pr1(−/−) mice had increased and persistent vascular permeability compared with wild-type mice, which was strongly correlated with the amount and localization of resulting pulmonary fibrosis. EC-S1pr1(−/−) mice also had increased immune cell infiltration and activation of the coagulation cascade within the lung. However, increased circulating S1P ligand in ApoM-overexpressing mice was insufficient to protect against bleomycin-induced pulmonary fibrosis. Overall, these data demonstrate that endothelial cell S1PR1 controls vascular permeability in the lung, is associated with changes in immune cell infiltration and extravascular coagulation, and modulates the fibrotic response to lung injury.

COVID-19 emerged as a worldwide pandemic early in 2020, and at this writing has caused over 170 million cases and 3.7 million deaths worldwide, and almost 600,000 deaths in the United States. The rapid development of several safe and highly efficacious vaccines stands as one of the most extraordinary achievements in modern medicine, but the identification and administration of efficacious therapeutics to treat patients suffering from COVID-19 has been far less successful. A major factor limiting progress in the development of effective treatments has been a lack of suitable preclinical models for the disease, currently reliant upon various animal models and in vitro culture of immortalized cell lines. Here we report the first successful demonstration of SARS-CoV-2 infection and viral replication in a human primary cell-based organ-on-chip, leveraging a recently developed tissue culture platform known as PREDICT96. This successful demonstration of SARS-CoV-2 infection in human primary airway epithelial cells derived from a living donor represents a powerful new pathway for disease modeling and an avenue for screening therapeutic candidates in a high throughput platform.

Supplemental Digital Content is available in the text., OBJECTIVES: We hypothesize that elevated soluble suppression of tumorigenicity-2 concentrations, a marker of pulmonary epithelial injury, reflect ongoing lung injury in acute hypoxemic respiratory failure due to coronavirus disease 2019 and associate with continued ventilator dependence. DESIGN: We associated serial plasma soluble suppression of tumorigenicity-2 levels and markers of systemic inflammation including d-dimer, C-reactive protein, and erythrocyte sedimentation rate with 30-day mortality and ventilator dependence. SETTING: Adult medical ICUs and general medicine wards at an academic teaching hospital in Boston, MA. PATIENTS: Adult patients with severe acute respiratory syndrome coronavirus 2 infection and acute hypoxemic respiratory failure admitted to the ICU (n = 72) and non-ICU patients managed with supplemental oxygen (n = 77). INTERVENTIONS: Observational study from April 25 to June 25, 2020. MEASUREMENTS AND MAIN RESULTS: ICU patients had a higher baseline body mass index and median soluble suppression of tumorigenicity-2, d-dimer, and C-reactive protein concentrations compared with non-ICU patients. Among ICU patients, elevated baseline modified Sequential Organ Failure Assessment score and log (soluble suppression of tumorigenicity-2) were associated with 30-day mortality, whereas initial Pao2/Fio2 and markers of systemic inflammation were similar between groups. Only log (soluble suppression of tumorigenicity-2) associated with ventilator dependence over time, with the last measured log (soluble suppression of tumorigenicity-2) concentration obtained on ICU day 11.5 (interquartile range [7–17]) higher in patients who required reintubation or tracheostomy placement compared with patients who were successfully extubated (2.10 [1.89–2.26] vs 1.87 ng/mL [1.72–2.13 ng/mL]; p = 0.03). Last measured systemic inflammatory markers, modified Sequential Organ Failure Assessment score, and Pao2/Fio2 were not different between patients who were successfully extubated compared with those with continued ventilator dependence. CONCLUSIONS: Plasma soluble suppression of tumorigenicity-2 is a biomarker readily measured in blood that can provide dynamic information about the degree of a patient’s lung injury and real-time assessment of the likelihood of extubation success. Measures of systemic inflammation, illness severity, and oxygenation did not associate with ventilator outcomes.

Background and objective In vivo evaluation of the microstructural differences between asthmatic and non-asthmatic airways and their functional consequences is relevant to understanding and, potentially, treating asthma. In this study, we use endobronchial optical coherence tomography to investigate how allergic airways with asthma differ from allergic non-asthmatic airways in baseline microstructure and in response to allergen challenge. Methods A total of 45 subjects completed the study, including 20 allergic, mildly asthmatic individuals, 22 non-asthmatic allergic controls and 3 healthy controls. A 3-cm airway segment in the right middle and right upper lobe were imaged in each subject immediately before and 24 h following segmental allergen challenge to the right middle lobe. Relationships between optical airway measurements (epithelial and mucosal thicknesses, mucosal buckling and mucus) and airway obstruction (FEV1 /FVC (forced expiratory volume in 1 s/forced vital capacity) and FEV1 % (FEV1 as a percentage of predictive value)) were investigated. Results Significant increases at baseline and in response to allergen were observed for all four of our imaging metrics in the asthmatic airways compared to the non-asthmatic airways. Epithelial thickness and mucosal buckling exhibited a significant relationship to FEV1 /FVC in the asthmatic group. Conclusion Simultaneous assessments of airway microstructure, buckling and mucus revealed both structural and functional differences between the mildly asthmatic and control groups, with airway buckling seeming to be the most relevant factor. The results of this study demonstrate that a comprehensive, microstructural approach to assessing the airways may be important in future asthma studies as well as in the monitoring and treatment of asthma.

Rationale: Early, accurate diagnosis of interstitial lung disease (ILD) informs prognosis and therapy, especially in idiopathic pulmonary fibrosis (IPF). Current diagnostic methods are imperfect. High-resolution computed tomography has limited resolution, and surgical lung biopsy (SLB) carries risks of morbidity and mortality. Endobronchial optical coherence tomography (EB-OCT) is a low-risk, bronchoscope-compatible modality that images large lung volumes in vivo with microscopic resolution, including subpleural lung, and has the potential to improve the diagnostic accuracy of bronchoscopy for ILD diagnosis. Objectives: We performed a prospective diagnostic accuracy study of EB-OCT in patients with ILD with a low-confidence diagnosis undergoing SLB. The primary endpoints were EB-OCT sensitivity/specificity for diagnosis of the histopathologic pattern of usual interstitial pneumonia (UIP) and clinical IPF. The secondary endpoint was agreement between EB-OCT and SLB for diagnosis of the ILD fibrosis pattern. Methods: EB-OCT was performed immediately before SLB. The resulting EB-OCT images and histopathology were interpreted by blinded, independent pathologists. Clinical diagnosis was obtained from the treating pulmonologists after SLB, blinded to EB-OCT. Measurements and Main Results: We enrolled 31 patients, and 4 were excluded because of inconclusive histopathology or lack of EB-OCT data. Twenty-seven patients were included in the analysis (16 men, average age: 65.0 yr): 12 were diagnosed with UIP and 15 with non-UIP ILD. Average FVC and Dl(CO) were 75.3% (SD, 18.5) and 53.5% (SD, 16.4), respectively. Sensitivity and specificity of EB-OCT was 100% (95% confidence interval, 75.8–100.0%) and 100% (79.6–100%), respectively, for both histopathologic UIP and clinical diagnosis of IPF. There was high agreement between EB-OCT and histopathology for diagnosis of ILD fibrosis pattern (weighted κ: 0.87 [0.72–1.0]). Conclusions: EB-OCT is a safe, accurate method for microscopic ILD diagnosis, as a complement to high-resolution computed tomography and an alternative to SLB.

Early, accurate diagnosis of interstitial lung disease (ILD) is critical for clinical management and therapeutic decision-making, especially distinguishing idiopathic pulmonary fibrosis (IPF) from non-IPF ILD. Unfortunately, current diagnostic imaging methods are limited in resolution and surgical biopsy methods are invasive. In this study, we evaluate the accuracy of endobronchial optical coherence tomography (EB-OCT) as a low-risk method for in vivo ILD diagnosis in patients undergoing surgical biopsy. EB-OCT was able to distinguish diagnostic microanatomical features of IPF from non-IPF ILDs, independently compared against surgical biopsy. These findings support the potential of OCT as a minimally-invasive method for IPF diagnosis.

Idiopathic pulmonary fibrosis (IPF) is a fatal form of fibrotic interstitial lung disease (ILD). Early diagnosis of IPF is essential, but often requires invasive surgery. We conduct a prospective study evaluating the diagnostic accuracy of endobronchial optical coherence tomography (EB-OCT) for IPF diagnosis as compared to concurrent surgical lung biopsy (SLB) and clinical follow-up diagnosis. EB-OCT was performed immediately prior to SLB in 27 ILD patients. EB-OCT was 100% sensitive and 100% specific for both histologic and clinical follow up diagnosis of IPF. The results demonstrate the potential of EB-OCT as minimally-invasive, low-risk in vivo method for microscopic IPF diagnosis.

A Woman with Dyspnea on Exertion and Abnormal Findings on Chest Imaging A 70-year-old woman was evaluated because of increasing dyspnea on exertion and abnormal findings on chest imaging. Eleven ye...

Influenza and other respiratory viruses present a significant threat to public health, national security, and the world economy, and can lead to the emergence of global pandemics such as from COVID-19. A barrier to the development of effective therapeutics is the absence of a robust and predictive preclinical model, with most studies relying on a combination of in vitro screening with immortalized cell lines and low-throughput animal models. Here, we integrate human primary airway epithelial cells into a custom-engineered 96-device platform (PREDICT96-ALI) in which tissues are cultured in an array of microchannel-based culture chambers at an air–liquid interface, in a configuration compatible with high resolution in-situ imaging and real-time sensing. We apply this platform to influenza A virus and coronavirus infections, evaluating viral infection kinetics and antiviral agent dosing across multiple strains and donor populations of human primary cells. Human coronaviruses HCoV-NL63 and SARS-CoV-2 enter host cells via ACE2 and utilize the protease TMPRSS2 for spike protein priming, and we confirm their expression, demonstrate infection across a range of multiplicities of infection, and evaluate the efficacy of camostat mesylate, a known inhibitor of HCoV-NL63 infection. This new capability can be used to address a major gap in the rapid assessment of therapeutic efficacy of small molecules and antiviral agents against influenza and other respiratory viruses including coronaviruses.

BACKGROUND: Obesity has multifactorial effects on lung function and exercise capacity. The contributions of obesity-related inflammatory pathways to alterations in lung function remain unclear. RESEARCH QUESTION: To examine the association of obesity-related inflammatory pathways with pulmonary function, exercise capacity, and pulmonary-specific contributors to exercise intolerance. METHOD: We examined 695 patients who underwent cardiopulmonary exercise testing (CPET) with invasive hemodynamic monitoring at Massachusetts General Hospital between December 2006–June 2017. We investigated the association of adiponectin, leptin, resistin, IL-6, CRP, and insulin resistance (HOMA-IR) with pulmonary function and exercise parameters using multivariable linear regression. RESULTS: Obesity-related inflammatory pathways were associated with worse lung function. Specifically, higher CRP, IL-6, and HOMA-IR were associated with lower percent predicted FEV(1) and FVC with a preserved FEV(1)/FVC ratio suggesting a restrictive physiology pattern (P≤0.001 for all). For example, a 1-SD higher natural-logged CRP level was associated with a nearly 5% lower percent predicted FEV(1) and FVC (beta −4.8, s.e. 0.9 for FEV1; beta −4.9, s.e. 0.8 for FVC; P

Background Patients with severe coronavirus disease 2019 (COVID-19) have respiratory failure with hypoxemia and acute bilateral pulmonary infiltrates, consistent with ARDS. Respiratory failure in COVID-19 might represent a novel pathologic entity. Research Question How does the lung histopathology described in COVID-19 compare with the lung histopathology described in SARS and H1N1 influenza? Study Design and Methods We conducted a systematic review to characterize the lung histopathologic features of COVID-19 and compare them against findings of other recent viral pandemics, H1N1 influenza and SARS. We systematically searched MEDLINE and PubMed for studies published up to June 24, 2020, using search terms for COVID-19, H1N1 influenza, and SARS with keywords for pathology, biopsy, and autopsy. Using PRISMA-Individual Participant Data guidelines, our systematic review analysis included 26 articles representing 171 COVID-19 patients; 20 articles representing 287 H1N1 patients; and eight articles representing 64 SARS patients. Results In COVID-19, acute-phase diffuse alveolar damage (DAD) was reported in 88% of patients, which was similar to the proportion of cases with DAD in both H1N1 (90%) and SARS (98%). Pulmonary microthrombi were reported in 57% of COVID-19 and 58% of SARS patients, as compared with 24% of H1N1 influenza patients. Interpretation DAD, the histologic correlate of ARDS, is the predominant histopathologic pattern identified in lung pathology from patients with COVID-19, H1N1 influenza, and SARS. Microthrombi were reported more frequently in both patients with COVID-19 and SARS as compared with H1N1 influenza. Future work is needed to validate this histopathologic finding and, if confirmed, elucidate the mechanistic underpinnings and characterize any associations with clinically important outcomes.

Idiopathic pulmonary fibrosis is a lung disease with limited therapeutic options that is characterized by pathological fibroblast activation and aberrant lung remodeling with scar formation. YAP (Yes-associated protein) is a transcriptional coactivator that mediates mechanical and biochemical signals controlling fibroblast activation. In this study, we developed a high-throughput small-molecule screen for YAP inhibitors in primary human lung fibroblasts. Multiple HMG-CoA (hydroxymethylglutaryl-coenzyme A) reductase inhibitors (statins) were found to inhibit YAP nuclear localization via induction of YAP phosphorylation, cytoplasmic retention, and degradation. We further show that the mevalonate pathway regulates YAP activation, and that simvastatin treatment reduces fibrosis markers in activated human lung fibroblasts and in the bleomycin mouse model of pulmonary fibrosis. Finally, we show that simvastatin modulates YAP in vivo in mouse lung fibroblasts. Our results highlight the potential of small-molecule screens for YAP inhibitors and provide a mechanism for the antifibrotic activity of statins in idiopathic pulmonary fibrosis.

Chronic obstructive pulmonary disease (COPD) is the most common noninfectious pulmonary disease among people living with HIV, independent of smoking. However, the cause for this enhanced susceptibility remains unclear, and the effects of HIV on pulmonary perfusion and ventilation are unknown. Methods: We used PET/CT in 46 smokers and nonsmokers, 23 of whom had documented HIV infection. Emphysema was assessed by CT and perfusion by (13)N ((13)NN) PET scans. After removal of image noise, vertical and axial gradients in perfusion were calculated. We tested for differences in the total spatial heterogeneity of perfusion (CV(2)(Qtotal)) and its components (CV(2)(Qtotal) = CV(2)(Qvgrad) [vertical gradient] + CV(2)(Qzgrad) [axial gradient] + CV(2)(Qr) [residual heterogeneity]) among groups. Results: There were no significant differences in demographic parameters among groups, and all subjects had minimal radiographic evidence of emphysema. Compared with controls, nonsmokers living with HIV had a significantly greater CV(2)(Qr)/CV(2)(Qtotal) (0.48 vs. 0.36, P = 0.05) and reduced CV(2)(Qvgrad)/CV(2)(Qtotal) (0.46 vs. 0.65, P = 0.038). Smokers also had a reduced CV(2)(Qvgrad)/CV(2)(Qtotal), however, there was no significant difference in CV(2)(Qvgrad)/CV(2)(Qtotal) between smokers living with and without HIV (0.39 vs. 0.34, P = 0.58), despite a decreased vertical perfusion gradient (Qv(grad)) in smokers living with HIV. Conclusion: In nonsmokers living with well-controlled HIV and minimal radiographic emphysema, HIV infection contributes to pulmonary perfusion abnormalities similar to smokers. These data indicate the onset of subclinical pulmonary perfusion abnormalities that could herald the development of significant lung disease in these susceptible individuals.

Pulmonary fibrosis is thought to result from dysregulated wound repair after repetitive lung injury. Many cellular responses to injury involve rearrangements of the actin cytoskeleton mediated by the two isoforms of the Rho-associated coiled-coil-forming protein kinase (ROCK), ROCK1 and ROCK2. In addition, profibrotic mediators such as transforming growth factor-β, thrombin, and lysophosphatidic acid act through receptors that activate ROCK. Inhibition of ROCK activation may be a potent therapeutic strategy for human pulmonary fibrosis. Pharmacological inhibition of ROCK using nonselective ROCK inhibitors has been shown to prevent fibrosis in animal models; however, the specific roles of each ROCK isoform are poorly understood. Furthermore, the pleiotropic effects of this kinase have raised concerns about on-target adverse effects of ROCK inhibition such as hypotension. Selective inhibition of one isoform might be a better-tolerated strategy. In the present study, we used a genetic approach to determine the roles of ROCK1 and ROCK2 in a mouse model of bleomycin-induced pulmonary fibrosis. Using ROCK1- or ROCK2-haploinsufficient mice, we found that reduced expression of either ROCK1 or ROCK2 was sufficient to protect them from bleomycin-induced pulmonary fibrosis. In addition, we found that both isoforms contribute to the profibrotic responses of epithelial cells, endothelial cells, and fibroblasts. Interestingly, ROCK1- and ROCK2-haploinsufficient mice exhibited similar protection from bleomycin-induced vascular leak, myofibroblast differentiation, and fibrosis; however, ROCK1-haploinsufficient mice demonstrated greater attenuation of epithelial cell apoptosis. These findings suggest that selective inhibition of either ROCK isoform has the potential to be an effective therapeutic strategy for pulmonary fibrosis.

BackgroundIn 2017, Massachusetts General Hospital implemented the Severe Immunotherapy Complications (SIC) Service, a multidisciplinary care team for patients hospitalized with immune-related adverse events (irAEs), a unique spectrum of toxicities associated with immune checkpoint inhibitors (ICIs). This study’s objectives were to evaluate the intervention’s (1) effect on patient outcomes and healthcare utilization, and (2) ability to collect biological samples via a central infrastructure, in order to study the mechanisms responsible for irAEs.MethodsA hospital database was used to identify patients who received ICIs for a malignancy and were hospitalized with severe irAEs, before (April 2, 2016–October 3, 2017) and after (October 3, 2017–October 24, 2018) SIC Service initiation. The primary outcome was readmission rate after index hospitalization. Secondary outcomes included length of stay (LOS) for admissions, corticosteroid and non-steroidal second-line immunosuppression use, ICI discontinuation, and inpatient mortality.ResultsIn the pre-SIC period, 127 of 1169 patients treated with ICIs were hospitalized for irAEs; in the post-SIC period, 122 of 1159. After SIC service initiation, reductions were observed in irAE readmission rate (14.8% post-SIC vs 25.9% pre-SIC; OR 0.46; 95% CI 0.22 to 0.95; p=0.036) and readmission LOS (median 6 days post-SIC vs 7 days pre-SIC; 95% CI −16.03 to –0.14; p=0.046). No significant pre-initiation and post-initiation differences were detected in corticosteroid use, second-line immunosuppression, ICI discontinuation, or inpatient mortality rates. The SIC Service collected 789 blood and tissue samples from 234 patients with suspected irAEs.ConclusionsThis is the first study to report that establishing a highly subspecialized care team focused on irAEs is associated with improved patient outcomes and reduced healthcare utilization. Furthermore, the SIC Service successfully integrated blood and tissue collection safety into routine care.

Solid tumours are infiltrated by effector T cells with the potential to control or reject them, as well as by regulatory T (Treg) cells that restrict the function of effector T cells and thereby promote tumour growth1. The anti-tumour activity of effector T cells can be therapeutically unleashed, and is now being exploited for the treatment of some forms of human cancer. However, weak tumour-associated inflammatory responses and the immune-suppressive function of Treg cells remain major hurdles to broader effectiveness of tumour immunotherapy2. Here we show that, after disruption of the CARMA1-BCL10-MALT1 (CBM) signalosome complex, most tumour-infiltrating Treg cells produce IFNγ, resulting in stunted tumour growth. Notably, genetic deletion of both or even just one allele of CARMA1 (also known as Card11) in only a fraction of Treg cells-which avoided systemic autoimmunity-was sufficient to produce this anti-tumour effect, showing that it is not the mere loss of suppressive function but the gain of effector activity by Treg cells that initiates tumour control. The production of IFNγ by Treg cells was accompanied by activation of macrophages and upregulation of class I molecules of the major histocompatibility complex on tumour cells. However, tumour cells also upregulated the expression of PD-L1, which indicates activation of adaptive immune resistance3. Consequently, blockade of PD-1 together with CARMA1 deletion caused rejection of tumours that otherwise do not respond to anti-PD-1 monotherapy. This effect was reproduced by pharmacological inhibition of the CBM protein MALT1. Our results demonstrate that partial disruption of the CBM complex and induction of IFNγ secretion in the preferentially self-reactive Treg cell pool does not cause systemic autoimmunity but is sufficient to prime the tumour environment for successful immune checkpoint therapy.

Mass cytometry (CyTOF), a mass spectrometry-based single cell phenotyping technology, allows utilization of over 35 antibodies in a single sample and is a promising tool for translational human immunology studies. Although several analysis tools are available to interpret the complex data sets generated, a robust method for standardization and quality control within and across studies is needed. Here we report an efficient and easily adaptable method to monitor quality of individual samples in human immunology studies and to facilitate reproducible data analysis. Samples to be assessed are spiked with a defined amount of reference peripheral blood mononuclear cells from a healthy donor, derived from a single large blood draw. The presence of known standardized numbers and phenotypic profiles of these reference cells greatly facilitates sample analysis by allowing for: 1) quality control for consistent staining of each antibody in the panel, 2) identification of potential batch effects, and 3) implementation of a robust gating strategy. We demonstrate the utility of this method using peripheral blood and bronchoalveolar lavage samples from HIV+ patients by characterizing their CD8+ T-cell phenotypes and cytokine expression, respectively. Our results indicate that this method allows quality control of experimental conditions and results in highly reproducible population frequencies through a robust gating strategy. © 2016 International Society for Advancement of Cytometry.

Idiopathic pulmonary fibrosis (IPF) is thought to result from aberrant tissue repair processes in response to chronic or repetitive lung injury. The origin and nature of the injury, as well as its cellular and molecular targets, are likely heterogeneous, which complicates accurate pre-clinical modelling of the disease and makes therapeutic targeting a challenge. Efforts are underway to identify central pathways in fibrogenesis which may allow targeting of aberrant repair processes regardless of the initial injury stimulus. Dysregulated endothelial permeability and vascular leak have long been studied for their role in acute lung injury and repair. Evidence that these processes are of importance to the pathogenesis of fibrotic lung disease is growing. Endothelial permeability is increased in non-fibrosing lung diseases, but it resolves in a self-limited fashion in conditions such as bacterial pneumonia and acute respiratory distress syndrome. In progressive fibrosing diseases such as IPF, permeability appears to persist, however, and may also predict mortality. In this hypothesis-generating review, we summarise available data on the role of endothelial permeability in IPF and focus on the deleterious consequences of sustained endothelial hyperpermeability in response to and during pulmonary inflammation and fibrosis. We propose that persistent permeability and vascular leak in the lung have the potential to establish and amplify the pro-fibrotic environment. Therapeutic interventions aimed at recognising and “plugging” the leak may therefore be of significant benefit for preventing the transition from lung injury to fibrosis and should be areas for future research.

The airway epithelial cell (AEC) response to allergens helps initiate and propagate allergic inflammation in asthma. CARMA3 is a scaffold protein that mediates G protein-coupled receptor-induced NF-κB activation in airway epithelium. In this study, we demonstrate that mice with CARMA3-deficient AECs have reduced airway inflammation, as well as reduced type 2 cytokine levels in response to Alternaria alternata. These mice also have reduced production of IL-33 and IL-25, and reduced numbers of innate lymphoid cells in the lung. We also show that CARMA3-deficient human AECs have decreased production of proasthmatic mediators in response to A. alternata. Finally, we show that CARMA3 interacts with inositol 1,4,5-trisphosphate receptors in AECs, and that inhibition of CARMA3 signaling reduces A. alternata-induced intracellular calcium release. In conclusion, we show that CARMA3 signaling in AECs helps mediate A. alternata-induced allergic airway inflammation, and that CARMA3 is an important signaling molecule for type 2 immune responses in the lung.

Solid tumours are infiltrated by effector T cells with the potential to control or reject them, as well as by regulatory T (T