Microscopic Small Airway Abnormalities Identified in Early Idiopathic Pulmonary Fibrosis In Vivo Using Endobronchial Optical Coherence Tomography.

RATIONALE: Idiopathic pulmonary fibrosis (IPF) affects subpleural lung, but is considered to spare small airways. Micro-CT studies demonstrated small airway reduction in end-stage IPF explanted lungs, raising questions about small airway involvement in early-stage disease. Endobronchial optical coherence tomography (EB-OCT) is a volumetric imaging modality that detects microscopic features from subpleural to proximal airways. We use EB-OCT to evaluate small airways in early IPF and control subjects in vivo. METHODS: EB-OCT was performed in 12 IPF and 5 control subjects (matched by age, sex, smoking-history, height, BMI). IPF subjects had early disease with mild restriction (FVC: 83.5% predicted), diagnosed per current guidelines and confirmed by surgical biopsy. EB-OCT volumetric imaging was acquired bronchoscopically in multiple, distinct, bilateral lung locations (total: 97 sites). IPF imaging sites were classified by severity into affected (all criteria for UIP present) and less affected (some but not all criteria for UIP present) sites. Bronchiole count and small airway stereology metrics were measured for each EB-OCT imaging site. RESULTS: Compared to control subjects (mean: 11.2 bronchioles/cm3; SD: 6.2), there was significant bronchiole reduction in IPF subjects (42% loss; mean: 6.5/cm3; SD: 3.4; p=0.0039), including in IPF affected (48% loss; mean: 5.8/cm3; SD: 2.8; p<0.00001) and IPF less affected (33% loss; mean: 7.5/cm3; SD: 4.1; p=0.024) sites. Stereology metrics showed IPF affected small airways were significantly larger and more distorted/irregular than in IPF less affected sites and control subjects. IPF less affected and control airways were statistically indistinguishable for all stereology parameters (p=0.36-1.0). CONCLUSION: EB-OCT demonstrated marked bronchiolar loss in early IPF (between 30 and 50%), even in areas minimally affected by disease, compared to matched controls. These findings support small airway disease as a feature of early IPF, providing novel insight into pathogenesis and potential therapeutic targets.

Ablation of lysophosphatidic acid receptor 1 attenuates hypertrophic cardiomyopathy in a mouse model.

Myocardial fibrosis is a key pathologic feature of hypertrophic cardiomyopathy (HCM). However, the fibrotic pathways activated by HCM-causing sarcomere protein gene mutations are poorly defined. Because lysophosphatidic acid is a mediator of fibrosis in multiple organs and diseases, we tested the role of the lysophosphatidic acid pathway in HCM. Lysphosphatidic acid receptor 1 (LPAR1), a cell surface receptor, is required for lysophosphatidic acid mediation of fibrosis. We bred HCM mice carrying a pathogenic myosin heavy-chain variant (403+/-) with Lpar1-ablated mice to create mice carrying both genetic changes (403+/- LPAR1 -/-) and assessed development of cardiac hypertrophy and fibrosis. Compared with 403+/- LPAR1WT, 403+/- LPAR1 -/- mice developed significantly less hypertrophy and fibrosis. Single-nucleus RNA sequencing of left ventricular tissue demonstrated that Lpar1 was predominantly expressed by lymphatic endothelial cells (LECs) and cardiac fibroblasts. Lpar1 ablation reduced the population of LECs, confirmed by immunofluorescence staining of the LEC markers Lyve1 and Ccl21a and, by in situ hybridization, for Reln and Ccl21a. Lpar1 ablation also altered the distribution of fibroblast cell states. FB1 and FB2 fibroblasts decreased while FB0 and FB3 fibroblasts increased. Our findings indicate that Lpar1 is expressed predominantly by LECs and fibroblasts in the heart and is required for development of hypertrophy and fibrosis in an HCM mouse model. LPAR1 antagonism, including agents in clinical trials for other fibrotic diseases, may be beneficial for HCM.

CRTH2 Mediates Profibrotic Macrophage Differentiation and Promotes Lung Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a particularly deadly form of pulmonary fibrosis of unknown cause. In patients with IPF, high serum and lung concentrations of CHI3L1 (chitinase 3 like 1) can be detected and are associated with poor survival. However, the roles of CHI3L1 in these diseases have not been fully elucidated. We hypothesize that CHI3L1 interacts with CRTH2 (chemoattractant receptor-homologous molecule expressed on T-helper type 2 cells) to stimulate profibrotic macrophage differentiation and the development of pulmonary fibrosis and that circulating blood monocytes from patients with IPF are hyperresponsive to CHI3L1-CRTH2 signaling. We used murine pulmonary fibrosis models to investigate the role of CRTH2 in profibrotic macrophage differentiation and fibrosis development and primary human peripheral blood mononuclear cell culture to detect the difference of monocytes in the responses to CHI3L1 stimulation and CRTH2 inhibition between patients with IPF and normal control subjects. Our results showed that null mutation or small-molecule inhibition of CRTH2 prevents the development of pulmonary fibrosis in murine models. Furthermore, CHI3L1 stimulation induces a greater increase in CD206 expression in IPF monocytes than control monocytes. These results demonstrated that monocytes from patients with IPF appear to be hyperresponsive to CHI3L1 stimulation. These studies support targeting the CHI3L1-CRTH2 pathway as a promising therapeutic approach for IPF and that the sensitivity of blood monocytes to CHI3L1-induced profibrotic differentiation may serve as a biomarker that predicts responsiveness to CHI3L1- or CRTH2-based interventions.

Endothelial-Specific Loss of Sphingosine-1-Phosphate Receptor 1 Increases Vascular Permeability and Exacerbates Bleomycin-induced Pulmonary Fibrosis.

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.

Interstitial lung disease is a dominant feature in patients with circulating myositis-specific antibodies.

BACKGROUND: Many patients with polymyositis (PM) or dermatomyositis (DM) have circulating myositis-specific antibodies (MSAs). Interstitial lung disease (ILD) is a common manifestation of PM/DM, and it can even precede the onset of characteristic muscle or skin manifestations. Furthermore, there appear to be some patients with ILD and circulating MSAs who do not develop muscle or skin disease even after prolonged follow-up. We sought to determine whether ILD is equally or more common than myositis or dermatitis at the time of initial detection of MSAs. METHODS: We identified all patients found to have circulating MSAs at our institution over a 4-year period and assessed for the presence of lung, muscle, and skin disease at the time of initial detection of MSAs. Among those found to have ILD, we compared demographic and clinical features, chest CT scan findings, and outcomes between those with PM/DM-associated ILD and those with ILD but no muscle or skin disease. RESULTS: A total of 3078 patients were tested for MSAs, and of these 40 were positive. Nine different MSAs were detected, with anti-histidyl tRNA synthetase (anti-Jo-1) being the most common (35% of MSAs). Among patients with positive MSAs, 86% were found to have ILD, compared to 39% and 28% with muscle and skin involvement, respectively (p < 0.001). Fifty percent of all MSA-positive patients had isolated ILD, with no evidence of muscle or skin disease. Those with isolated ILD were more likely to be older and have fibrotic changes on chest CT, less likely to receive immunomodulatory therapy, and had worse overall survival. CONCLUSIONS: In this study we found that individuals with circulating MSAs were more likely to have ILD than classic muscle or skin manifestations of PM/DM at the time of initial detection of MSAs. Our findings suggest that the presence of ILD should be considered a disease-defining manifestation in the presence of MSAs and incorporated into classification criteria for PM/DM.

Diagnosis and Management of Idiopathic Pulmonary Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is the most common of the idiopathic interstitial pneumonias. Its signs and symptoms are relatively non-specific, and patients often present with chronic cough, progressive dyspnea, resting or exertional hypoxemia, and inspiratory crackles on lung auscultation. Definitive diagnosis requires the exclusion of known causes of pulmonary fibrosis and identification of the usual interstitial pneumonia (UIP) pattern of disease either on high-resolution computed tomography (HRCT) scan of the chest or on surgical lung biopsy. Multidisciplinary discussion involving pulmonologists, radiologists, and pathologists with expertise in the diagnosis of IPF and other forms of interstitial lung disease is recommended and often required. Management focuses on anti-fibrotic therapy and early referral to lung transplant centers for those who are candidates. This review will discuss the current recommendations for the diagnosis, prognostication, and management of patients with IPF.

The Role of Surgical Lung Biopsy in the Diagnosis of Fibrotic Interstitial Lung Disease: Perspective from the Pulmonary Fibrosis Foundation.

Diagnosis of interstitial lung disease (ILD) requires a multidisciplinary discussion approach that includes clinicians, radiologists, and pathologists. Surgical lung biopsy (SLB) is currently the recommended standard in obtaining pathologic specimens for patients with ILD requiring a tissue diagnosis. The increased diagnostic confidence and accuracy provided by microscopic pathology assessment of SLB specimens must be balanced with the associated risks in patients with ILD. This document was developed by the SLB Working Group of the Pulmonary Fibrosis Foundation, composed of a multidisciplinary group of ILD physicians, including pulmonologists, radiologists, pathologists, and thoracic surgeons. In this document, we present an up-to-date literature review of the indications, contraindications, risks, and alternatives to SLB in the diagnosis of fibrotic ILD; outline an integrated approach to the decision-making around SLB in the diagnosis of fibrotic ILD; and provide practical information to maximize the yield and safety of SLB.

A Case of Interstitial Pneumonia with Features of Autoimmunity.

We present a case of a 61-year-old woman with several months of gradually worsening shortness of breath, requiring multiple hospitalizations with acute hypoxemic respiratory failure. She was initially treated for eosinophilic pneumonia presumed to be secondary to medications or rheumatoid lung without much improvement. Her subsequent chest CT showed honeycombing and diffuse ground-glass opacities, and she was found to have elevated rheumatoid factor (RF) and anti-CCP antibody titers without extrathoracic features of rheumatoid arthritis. This clinical scenario was suggestive of an interstitial lung disease (ILD) due to occult underlying connective tissue disorder (CTD), along the lines of the recently proposed entity interstitial pneumonia with autoimmune features (IPAF). She continued to deteriorate rapidly and passed away after experiencing recurrent exacerbations. As there is limited evidence to explain the clinical course of such patients, there is a need for prospective research to develop tailored regimens to prevent progression or even reverse the disease process.

Inpatient Dialysis Planning During the COVID-19 Pandemic: A Single-Center Experience and Review of the Literature.

BACKGROUND: COVID-19 has created havoc in healthcare systems worldwide, including shortages in equipment and supplies for dialysis in the acute setting. METHODS: We compared our planning and experience at a tertiary care academic medical center to recommendations in the literature. RESULTS: Published literature and our experience underscored the need to plan for adequate dialysis equipment, particularly for continuous renal replacement therapy in the ICU setting, adequate nursing, and flexible scheduling of chronic patients to accommodate the surge in acute patients. We discovered other "shortages" not mentioned in the literature: shortages in the number of portable reverse osmosis (RO) machines needed to prepare dialysis water, inadequate number of rooms in units designated for COVID-19 patients with plumbing for dialysis, and lack of temperature blending valves on sinks that necessitated using cold water only, and damaging the RO membranes. We identified the need for cooperation between nephrology and critical care medicine, hospital-based and community nephrologists and community dialysis units as well as nephrologists at other hospitals in the region. We turned to guidance from the hospital ethics committee. CONCLUSION: Planning for an expected surge in hospitalized patients requiring RRT demands coordination between critical care, dialysis and nursing services as well as community and hospital providers to make certain there are adequate dialysis resources. Our experience suggests that continuous dialysis is in greatest demand early in the illness, and that plans to increase supplies should be put in place. But, planning should also focus on unforeseen hospital-specific infrastructure shortages that can develop over time and hamper intermittent dialysis delivery to all patients who require treatment.

Vascular permeability in the fibrotic lung.

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.

Galectin-3 Interacts with the CHI3L1 Axis and Contributes to Hermansky-Pudlak Syndrome Lung Disease.

Hermansky-Pudlak syndrome (HPS) comprises a group of inherited disorders caused by mutations that alter the function of lysosome-related organelles. Pulmonary fibrosis is the major cause of morbidity and mortality in HPS-1 and HPS-4 patients. However, the mechanisms that underlie the exaggerated injury and fibroproliferative repair responses in HPS have not been adequately defined. In particular, although Galectin-3 (Gal-3) is dysregulated in HPS, its roles in the pathogenesis of HPS have not been adequately defined. In addition, although chitinase 3-like 1 (CHI3L1) and its receptors play major roles in the injury and repair responses in HPS, the ability of Gal-3 to interact with or alter the function of these moieties has not been evaluated. In this article, we demonstrate that Gal-3 accumulates in exaggerated quantities in bronchoalveolar lavage fluids, and traffics abnormally and accumulates intracellularly in lung fibroblasts and macrophages from bleomycin-treated pale ear, HPS-1-deficient mice. We also demonstrate that Gal-3 drives epithelial apoptosis when in the extracellular space, and stimulates cell proliferation and myofibroblast differentiation when accumulated in fibroblasts and M2-like differentiation when accumulated in macrophages. Biophysical and signaling evaluations also demonstrated that Gal-3 physically interacts with IL-13Rα2 and CHI3L1, and competes with TMEM219 for IL-13Rα2 binding. By doing so, Gal-3 diminishes the antiapoptotic effects of and the antiapoptotic signaling induced by CHI3L1 in epithelial cells while augmenting macrophage Wnt/ß-catenin signaling. Thus, Gal-3 contributes to the exaggerated injury and fibroproliferative repair responses in HPS by altering the antiapoptotic and fibroproliferative effects of CHI3L1 and its receptor complex in a tissue compartment-specific manner.

Trends in mortality from idiopathic pulmonary fibrosis in the European Union: an observational study of the WHO mortality database from 2001-2013.

Idiopathic pulmonary fibrosis (IPF) is the most common of the idiopathic interstitial pneumonias and is characterised by progressive accumulation of scar tissue in the lungs. The objective of this study was to describe the current mortality rates due to IPF in Europe, based on the World Health Organization (WHO) mortality database.We used country-level data for IPF mortality, identified in the WHO mortality database using International Classification of Diseases 10th Edition (ICD-10) codes, for the period 2001-2013. Joinpoint analysis was performed to describe trends throughout the observation period.The median mortality was 3.75 per 100 000 (interquartile range (IQR) 1.37-5.30) and 1.50 per 100 000 (IQR 0.65-2.02) for males and females, respectively. IPF mortality increased in the majority of the European Union (EU) countries with the exceptions of Denmark, Croatia, Austria and Romania. There was a significant disparity in rates across Europe, in the range 0.41-12.1 per 100 000 for men and 0.24-5.63 per 100 000 for women. The most notable increases were observed in the United Kingdom and Finland. Rates were also substantially higher in males, with sex disparity increasing across the period.The reported IPF mortality appears to be increasing across the EU; however, there is substantial variation in mortality trends and overall reported mortality rates between countries.

The Rho Kinase Isoforms ROCK1 and ROCK2 Each Contribute to the Development of Experimental Pulmonary Fibrosis.

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-ß, 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.

Corrigendum: ADAM10-mediated ephrin-B2 shedding promotes myofibroblast activation and organ fibrosis.

This corrects the article DOI: 10.1038/nm.4419.

ADAM10-mediated ephrin-B2 shedding promotes myofibroblast activation and organ fibrosis.

Maladaptive wound healing responses to chronic tissue injury result in organ fibrosis. Fibrosis, which entails excessive extracellular matrix (ECM) deposition and tissue remodeling by activated myofibroblasts, leads to loss of proper tissue architecture and organ function; however, the molecular mediators of myofibroblast activation have yet to be fully identified. Here we identify soluble ephrin-B2 (sEphrin-B2) as a new profibrotic mediator in lung and skin fibrosis. We provide molecular, functional and translational evidence that the ectodomain of membrane-bound ephrin-B2 is shed from fibroblasts into the alveolar airspace after lung injury. Shedding of sEphrin-B2 promotes fibroblast chemotaxis and activation via EphB3 and/or EphB4 receptor signaling. We found that mice lacking ephrin-B2 in fibroblasts are protected from skin and lung fibrosis and that a disintegrin and metalloproteinase 10 (ADAM10) is the major ephrin-B2 sheddase in fibroblasts. ADAM10 expression is increased by transforming growth factor (TGF)-ß1, and ADAM10-mediated sEphrin-B2 generation is required for TGF-ß1-induced myofibroblast activation. Pharmacological inhibition of ADAM10 reduces sEphrin-B2 levels in bronchoalveolar lavage and prevents lung fibrosis in mice. Consistent with the mouse data, ADAM10-sEphrin-B2 signaling is upregulated in fibroblasts from human subjects with idiopathic pulmonary fibrosis. These results uncover a new molecular mechanism of tissue fibrogenesis and identify sEphrin-B2, its receptors EphB3 and EphB4 and ADAM10 as potential therapeutic targets in the treatment of fibrotic diseases.

Uncoupling of the profibrotic and hemostatic effects of thrombin in lung fibrosis.

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 αvß6 induction, TGF-ß 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/αvß6/TGF-ß 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.