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BackgroundConsiderable clinical heterogeneity in idiopathic pulmonary fibrosis (IPF) suggests the existence of multiple disease endotypes. Identifying these endotypes would improve our understanding of the pathogenesis of IPF and could allow for a biomarker-driven personalised medicine approach. We aimed to identify clinically distinct groups of patients with IPF that could represent distinct disease endotypes.MethodsWe co-normalised, pooled and clustered three publicly available blood transcriptomic datasets (total 220 IPF cases). We compared clinical traits across clusters and used gene enrichment analysis to identify biological pathways and processes that were over-represented among the genes that were differentially expressed across clusters. A gene-based classifier was developed and validated using three additional independent datasets (total 194 IPF cases).FindingsWe identified three clusters of patients with IPF with statistically significant differences in lung function (p=0.009) and mortality (p=0.009) between groups. Gene enrichment analysis implicated mitochondrial homeostasis, apoptosis, cell cycle and innate and adaptive immunity in the pathogenesis underlying these groups. We developed and validated a 13-gene cluster classifier that predicted mortality in IPF (high-risk clusters vs low-risk cluster: HR 4.25, 95% CI 2.14 to 8.46, p=3.7×10−5).InterpretationWe have identified blood gene expression signatures capable of discerning groups of patients with IPF with significant differences in survival. These clusters could be representative of distinct pathophysiological states, which would support the theory of multiple endotypes of IPF. Although more work must be done to confirm the existence of these endotypes, our classifier could be a useful tool in patient stratification and outcome prediction in IPF.

IntroductionPersisting respiratory symptoms in COVID-19 survivors may be related to development of pulmonary fibrosis. We assessed the proportion of chest CT scans and pulmonary function tests consistent with parenchymal lung disease in the follow-up of people hospitalised with COVID-19 and viral pneumonitis.MethodsSystematic review and random effects meta-analysis of proportions using studies of adults hospitalised with SARS-CoV-2, SARS-CoV, MERS-CoV or influenza pneumonia and followed up within 12 months. Searches performed in MEDLINE and Embase. Primary outcomes were proportion of radiological sequelae on CT scans; restrictive impairment; impaired gas transfer. Heterogeneity was explored in meta-regression.ResultsNinety-five studies (98.9% observational) were included in qualitative synthesis, 70 were suitable for meta-analysis including 60 SARS-CoV-2 studies with a median follow-up of 3 months. In SARS-CoV-2, the overall estimated proportion of inflammatory sequelae was 50% during follow-up (0.50; 95% CI 0.41 to 0.58; I2=95%), fibrotic sequelae were estimated in 29% (0.29; 95% CI 0.22 to 0.37; I2=94.1%). Follow-up time was significantly associated with estimates of inflammatory sequelae (−0.036; 95% CI −0.068 to –0.004; p=0.029), associations with fibrotic sequelae did not reach significance (−0.021; 95% CI −0.051 to 0.009; p=0.176). Impaired gas transfer was estimated at 38% of lung function tests (0.38 95% CI 0.32 to 0.44; I2=92.1%), which was greater than restrictive impairment (0.17; 95% CI 0.13 to 0.23; I2=92.5%), neither were associated with follow-up time (p=0.207; p=0.864).DiscussionSequelae consistent with parenchymal lung disease were observed following COVID-19 and other viral pneumonitis. Estimates should be interpreted with caution due to high heterogeneity, differences in study casemix and initial severity.PROSPERO registration numberCRD42020183139.

Interstitial lung abnormalities (ILA) can be incidentally detected in patients undergoing low-dose CT screening for lung cancer. In this retrospective study, we explore the downstream impact of ILA detection on interstitial lung disease (ILD) diagnosis and treatment. Using a targeted approach in a lung cancer screening programme, the rate of de novo ILD diagnosis was 1.5%. The extent of abnormality on CT and severity of lung function impairment, but not symptoms were the most important factors in differentiating ILA from ILD. Disease modifying therapies were commenced in 39% of ILD cases, the majority being antifibrotic therapy for idiopathic pulmonary fibrosis.

IntroductionThe Winter Meeting of the British Thoracic Society (BTS) is a platform for the latest clinical and scientific research in respiratory medicine. This review summarises some key symposia and presentations from the BTS Winter Meeting 2018.MethodsKey symposia and research presentations from the BTS Winter Meeting 2018 were attended and reviewed by the authors.ResultsThe seminal messages from the latest clinical and scientific research covering a range of respiratory diseases, including asthma, interstitial lung disease, infection, cystic fibrosis, pulmonary vascular disease, pleural disease and occupational lung disease were summarised in this review.DiscussionThe BTS Winter Meeting 2018 brought the very best of respiratory research to an audience of scientists, physicians, nurses and allied health professionals. The Winter Meeting continues to be a highlight of the UK respiratory research calendar, and we look forward to the next meeting in December 2019.

This article reviews the British Thoracic Society Winter Meeting 2017 and summarises the new developments in scientific and clinical research across the breadth of respiratory medicine. The article discusses a number of symposia and selected abstract presentations from the meeting.

At the end of January 2020, the WHO declared the SARS-CoV-2 outbreak a public health emergency of international concern, its highest level of alarm. Although measures in the UK to reduce spread and ‘protect the NHS’ did prevent a complete collapse of the acute healthcare system, delay in the implementation of a lockdown until 23 March led to tens of thousands of excess deaths and, by December, more than 220,000 COVID-19 hospital admissions, with effects for many individuals persisting beyond hospital discharge (https://coronavirus.data.gov.uk/healthcare).1–5 The SARS-CoV-2 targets the respiratory epithelium and has a myriad of clinical consequences ranging from asymptomatic illness, through to mild, moderate and severe disease. Although the initial illness leads primarily to respiratory symptoms, many patients will have physical, cognitive and psychological disability that will require long-term management. In this edition of Thorax , four Brief Communications from the UK and Canada report the short-term to medium-term post-discharge outcome of hospitalised COVID-19 patients, identifying a pattern of five key persisting symptoms: breathlessness, cough, fatigue, muscle and joint pain and poor sleep quality.6–9 Mandal and colleagues recruited 384 patients from three large London hospitals.6 A median 54 days after discharge, approximately half of the patients reported persistent breathlessness with a third reporting an ongoing cough. Two-thirds of patients described persistent fatigue and poor sleep with 15% reporting scores consistent with depression. Biomarkers of ongoing inflammation (D-dimer and CRP) were commonly elevated. Only 11% of patients reported an absence of all four of these key symptoms. Of note, although respiratory disease was a common comorbidity in hospitalised patients, the MRC Dyspnoea Score at follow-up was not different between those with and …

> Ignorance, allied with power, is the most ferocious enemy justice can have. James Baldwin On the eve of President Trump’s inauguration in 2017, we published an editorial setting out the threat that climate change poses to lung health. We urged the incoming President to face up to his responsibilities and take steps to address this growing existential threat.1 We drew a parallel with the actions of two conservative leaders in the 1980s, Ronald Reagan and Margaret Thatcher who, faced with unequivocal scientific evidence, led the decisive steps necessary to avert the destruction of the ozone layer.2 3 Rather than being contrary to his stated goals, acting on climate change, we wrote ‘is one way in which President Trump can make good on his promises to improve the wellbeing of Americans, increase America’s energy independence, and act with fiscal prudence. Investing in green infrastructure creates jobs. It reduces healthcare costs, as 1/4 - 1/3 of the costs of decarbonising come straight back as health economic gains’.4 Sadly, 2 years on our worst fears have been realised. The USA is experiencing an unprecedented wave of natural disasters attributable to climate change, including widespread wildfires, destructive storms and temperature swings. However, rather than providing global leadership, the Trump administration has not merely abdicated responsibility but is actively sabotaging efforts to mitigate climate change by announcing plans to withdraw from the Paris Climate accords.5 Clear evidence in the most recent Intergovernmental Panel on Climate Change report sets …

This article reviews the British Thoracic Society Winter Meeting 2013, the annual scientific meeting attended by over 2000 delegates and representing the depth and breadth of UK respiratory medicine. This year's meeting from 4 to 6 December in London featured cutting-edge research alongside keynote symposia from international experts in respiratory science, epidemiology and clinical trials. This article reviews the key symposia and selected abstract sessions from the 2013 meeting.

IntroductionThe Its Not JUST Idiopathic pulmonary fibrosis Study (INJUSTIS) is a multicentre, prospective, observational cohort study. The aims of this study are to identify genetic, serum and other biomarkers that may identify specific molecular mechanisms, reflecting disease endotypes that are shared among patients with progressive pulmonary fibrosis regardless of aetiology. Furthermore, it is anticipated that these biomarkers will help predict fibrotic activity that may identify patterns of disease behaviour with greater accuracy than current clinical phenotyping.Methods and analysis200 participants with the multidisciplinary team confirmed fibrotic lung disease (50 each of rheumatoid-interstitial lung disease (ILD), asbestosis, chronic hypersensitivity pneumonitis and unclassifiable ILD) and 50 idiopathic pulmonary fibrosis participants, recruited as positive controls, will be followed up for 2 years. Participants will have blood samples, lung function tests, quality of life questionnaires and a subgroup will be offered bronchoscopy. Participants will also be given the option of undertaking blinded home handheld spirometry for the first 3 months of the study. The primary end point will be identification of a biomarker that predicts disease progression, defined as 10% relative change in forced vital capacity (FVC) or death at 12 months.Ethics and disseminationThe trial has received ethical approval from the National Research Ethics Committee Nottingham (18/EM/0139). All participants must provide written informed consent. The trial will be overseen by the INJUSTIS steering group that will include a patient representative, and an independent chairperson. The results from this study will be submitted for publication in peer-reviewed journals and disseminated at regional and national conferences.Trial registration numberNCT03670576.

This is a story from recent history that we believe incoming President Trump urgently needs to hear. In 1985, a huge and growing hole in the planet's ozone layer was identified. Ozone in the stratosphere blocks some of the sun's ultraviolet radiation from reaching the Earth's surface, thus protecting its biosphere (which includes humans) from DNA damage that would otherwise occur. The hole was caused by the action of chlorofluorocarbons (CFCs) used in refrigeration and as aerosol propellants.1 Margaret Thatcher is admired by many on both sides of the Atlantic, including President Trump2 as a strong politician, a person with clear beliefs on which she acted; the ‘Iron Lady’ as Donald Trump has described her on Twitter. A scientist by background, Thatcher appreciated the magnitude of the threat immediately, throwing her weight behind international efforts to address this. Working with Ronald Reagan, within 2 years the 1987 Montreal Protocol was signed to phase out the production and use of CFCs. Despite this, because CFCs persist in the atmosphere for more than a century, the ozone layer will not recover completely until 2060. A failure of leadership at that time would have been catastrophic. CFCs are also powerful greenhouse gases and without the Montreal agreement the world would already …

Pirfenidone works. There have been four randomised placebo-control trials of pirfenidone for the treatment of idiopathic pulmonary fibrosis (IPF); a phase II and phase III study in Japan, and two international multicentre phase III studies.1–3 In all four studies, patients treated with pirfenidone had slower rates of decline in lung volume (vital capacity (VC) in the Japanese studies and forced vital capacity (FVC) in the international studies) than placebo, and in three studies, the results were statistically significant. National Institute of Clinical Excellence (NICE) agrees that pirfenidone has a ‘modest but measurable effect on slowing the decline in lung function’.4 Therefore, whether patients receive pirfenidone depends on whether pirfenidone works enough to justify its cost. IPF is a chronic progressive disease of unknown aetiology, it is fatal with a median survival of approximately 3 years,5 or less than one electoral cycle. No therapy has been proven to improve survival. Despite the absence of good evidence, various combinations of N-acetylcysteine (NAC), prednisolone and azathioprine have been used for many years to treat patients with IPF. In the early 1990s, a small study suggested that prednisolone and azathioprine might be beneficial,6 and the initial British Thoracic Society (BTS) guidelines for the treatment of cryptogenic fibrosing alveolitis (as IPF was then known) recommended this therapy.7 In 2005, the Idiopathic Pulmonary Fibrosis International Group Exploring N-Acetylcysteine I Annual (IFIGENIA) study demonstrated that NAC might be beneficial in IPF.8 However, this study was criticised at the time for including azathioprine and prednisolone in both the placebo and treatment arms. The inclusion of this faux placebo lead some, possibly correctly, to suggest that NAC was inhibiting the adverse effects of prednisolone and azathioprine, rather than possessing any direct antifibrotic effect. The 2008 BTS IPF guidelines9 implicitly recommended treatment with triple …

Background We have previously demonstrated that people with idiopathic pulmonary fibrosis (IPF) are more likely to have a prothrombotic state and that people with IPF and a prothrombotic state have a higher risk of death at a year’s follow up. The aim of this study was to establish the impact of clotting abnormalities on the natural history of IPF with respect to median survival and lung function (forced vital capacity (FVC)). Methods We recruited 211 incident cases of radiologically diagnosed definite or probable IPF and collected longitudinal information on pulmonary function tests done as part of routine care. All participants were tagged with the NHS Information Centre to enable us to collect data on mortality. Blood samples were tested for a prothrombotic state defined as at least one inherited or acquired clotting defect or marker of fibrinolytic dysfunction. Kaplan-Meir methods were used to calculate median survival. Random effects linear regression modelling was used to estimate decline in FVC. Results Median follow-up was four years, during which 148 (70.1%) people died. Median survival in those with and without prothrombotic state was 2.7 and 3.7 years respectively (see Figure 1). We found evidence of effect modification between risk of death and follow-up time (p = 0.031). There was more than a three-fold increase risk of death in individuals with IPF and a prothrombotic state in the first half of follow-up (HR 3.36, 95% CI: 1.35 to 8.36), but this was reduced (HR 1.79, 95% CI: 1.08 to 2.94) in the second half. The estimated decline in FVC was 288mls (95% CI: 184 to 392mls) in those with normal clotting and 328mls (95% CI: 269 to 387mls) in those with one or more clotting defects. Conclusions Coagulation dysfunction has an adverse impact on the natural history of IPF, both in terms of median survival and lung function decline. Our findings suggest that a prothombotic state may be a useful biomarker to predict prognosis as part of routine care.

Introduction and Objectives: Idiopathic Pulmonary Fibrosis (IPF) is a chronic, progressive interstitial lung disease that is refractory to current treatment options. Transforming growth factor (TGF)-β1 is a key pro-fibrotic cytokine that plays a crucial role in IPF pathogenesis. Our group previously demonstrated distinct epigenetic modifications involved in repression of the antifibrotic gene cyclooxygenase-2 (COX-2) in fibroblasts from IPF (F-IPF) lungs compared with fibroblasts from non-fibrotic lungs (F-NL). Epigenetic drugs capable of inhibiting DNA and histone modifications may, therefore, represent a putative novel therapy. The aim of this study was to investigate the ability of 4 epigenetic inhibitors to regulate TGF-β-driven fibrosis in ex vivo mouse lung. Methods: A precision-cut lung slice (PCLS) model of fibrosis was established using the previously described [1] CC10-tTS-rtTA-TGFβ1 transgenic (tgTGF-β1) mouse. The model was first assessed by investigating PCLS overexpression of TGF-β1 in response to stimulation of the transgene by doxycycline treatment. Gene expression of COX-2 and fibrotic markers including collagen were assessed after 4 days of treatment. The anti-fibrotic potential of 4 epigenetic inhibitors; BIX01294 (BIX, inhibitor of G9a histone methyltransferase), 3-deazaneplanocin A (DZNep, inhibitor of EZH2 histone methyltransferase), SAHA (inhibitor of histone deacetylases, HDACs) and Decitabine (DAC, DNA demethylating agent) was investigated. Viability of PCLS was assessed by MTT and Prestoblue® viability assay. Results: Treatment of PCLS from tgTGF-β1 mice with doxycycline induced a concentration-dependent increase in global TGF-β1, pro-fibrotic markers including collagen and pro-inflammatory COX-2, which was comparable to recombinant TGF-β1 treatment. Treatment with three of the epigenetic inhibitors BIX01294, DZNep and DAC did not reduce the pro-fibrotic response following doxycycline treatment. However SAHA demonstrated a significant suppressive effect on COX-2 and collagen expression, while not directly affecting TGF-β1 transgene expression. Conclusions: The data suggests that SAHA has the potential to reduce fibrosis in a TGF-β1 driven model of pulmonary fibrosis. Further work is currently underway to assess the anti-fibrotic potential of this drug in tgTGF-β1 animals.

The first few years of the current millennium has seen a paradigm shift in the way people view idiopathic pulmonary fibrosis (IPF). In 2004, Raghu et al 1published the first multicentre, double blind, randomised, placebo-controlled trial in IPF. Although a negative study, it taught the field much about the natural history and conduct of clinical trials in IPF. Ultimately, this study paved the way for the Clinical Studies Assessing Pirfenidone in idiopathic pulmonary fibrosis: Research of Efficacy and Safety Outcomes (CAPACITY) studies2 that lead to the approval of Pirfenidone in Europe and eventually the Assessment of Pirfenidone to Confirm Efficacy and Safety in Idiopathic Pulmonary Fibrosis (ASCEND)3 and Prospective Observation of Fibrosis in the Lung: Clinical Endpoints (IMPULSIS)4 studies, which might lead to approval for the use of two disease-modifying drugs for the treatment of IPF in both the USA and Europe. Any thought at the turn of the century that progressive fibrotic lung disease, in the absence of inflammation, might be modifiable would have been regarded with a fair degree of scepticism. The field has come far. However, the journey has only just begun. Extrapolating data from recent epidemiological studies would suggest there has been approximately a doubling of the incidence of IPF and associated healthcare requirements since the turn of the century.5 ,6 Furthermore, patients enrolled into clinical trials represent a unique subpopulation of patients with IPF, itself a subpopulation of patients …

Introduction Idiopathic pulmonary fibrosis (IPF) is a restrictive lung disease of unknown cause. The median survival time after diagnosis is 3–5 years and there are limited treatments available. MUC5AC , MUC5B and TOLLIP in the 11p15.5 region have been shown to be associated with susceptibility to IPF. A variant in TOLLIP (rs5743890) has also been shown to be associated with both susceptibility and survival time however the effects were in opposite directions, i.e. the allele associated with increased susceptibility was also associated with increased survival time. Methods We performed survival analysis on 612 European IPF cases passing QC using a Cox proportional hazards model adjusting for age, sex, first 10 principal components and study centre. 134 variants were genotyped in a 200 kb region on chromosome 11 covering MUC5AC , MUC5B and TOLLIP . Results No SNPs in this region reached genome-wide significance (p –8 ). The most significant variant was rs56367042 (Hazard Ratio 3.38, 95% CI [1.91, 5.96]; p = 2.74 × 10 –5 ) which is located downstream of MUC5B . The next significant SNP was rs5743894 (Hazard Ratio 0.67, 95% CI [0.54, 0.83]; p = 2.24 × 10 –4 ) located in TOLLIP . For this SNP the allele we found associated with increased survival time has previously been reported as associated with increased susceptibility (showing a similar pattern to that reported for rs5743890) however this SNP has also been previously reported as not being associated with survival time. We found a proxy of rs5743890 (R 2 = 0.698) not to be associated with survival time (p = 0.881). Conclusions Our results did not replicate those previously reported; however they may support the hypothesis that variants in TOLLIP that increase susceptibility may also increase survival time. In future we will perform a GWAS for susceptibility and survival time and try and replicate the findings.

Progress in medicine and science depends on the dissemination of accurate information about health, disease and therapy. Social media are key tools to facilitate this, bringing research findings to people's attention more rapidly than ever before and also to a wider and more diverse population than previously. For information to be useful, it must be presented and interpreted in context. This is the difference between information and knowledge. We are keen to help our authors participate in this beyond the simple publication of papers to improve the quality of online debate, engaging researchers and clinicians as well as patients1 and others with a stake in healthcare. From now on, we …

Airway remodelling is a common feature of severe asthma. Transforming growth factor-s (TGF-s) is a pro-fibrotic, pleiotropic cytokine implicated in airway remodelling. TGF-s is sequestered in the extracellular matrix as a latent complex and requires activation to function. We have previously shown that contraction agonists cause aVs5-mediated TGF-s activation by human airway smooth muscle cells. The study aims were to investigate the role of the aVs5integrin in airway remodelling in vivo using two distinct mouse models of asthma. A blocking antibody directed against the aVs5 was used in the ovalbumin (OVA) model of asthma. Mice were sensitised with OVA/Alum on days 0 and 12, then challenged byoropharyngeal administration of OVA 10 times over 2 weeks. The anti-aVs5antibody or an isotype matched control antibody was administered for the duration of the OVA challenges. The second invivo model utilised itgb5 −/− mice. Aspergillous fumigatus antigen preparation was administered intra-nasally (10 μg/mice) to itgb5 −/− and wild type controls 9 times over a 21-day period. a-Smooth muscle actin (a-SMA) was quantified in lung sections from both studies by immunofluorescence. Murine airway smooth muscle cells express aVs5 integrin and can activate TGF-s in vivo in response to allergen challenge as measured by αvβ5 and phospho-Smad2 immunostaining. Treatment with both OVA and Asp.f resulted in an increase in a-SMA staining around the smaller airways. The aVs5blocking antibody significantly reduced a-SMA staining compared with the isotype control antibody. Furthermore, itgb5 −/− mice had significantly less a-SMA around their airways than wild type control mice in response to Asp.f treatment. However, both itgb5 −/− andanti-aVs5 treated mice had significantly more airway inflammation and more inflammatory cells present in the bronchoalveolar lavage compared with their matched controls. These data provide evidence that airway smooth muscle cells can activate TGF-s in vivo. Inhibition of, or genetic loss of, the aVs5 integrin significantly reduces allergen-induced increases in airway smooth muscle mass, however, peribronchial inflammation is increased consistent with the known effects of TGFβ. Targeted inhibition of the aVs5 integrin may reduce airway remodelling, but global inhibition is unlikely to be useful due to the enhanced inflammatory response.

Introduction Cough is a major symptom of Idiopathic Pulmonary fibrosis (IPF), a progressive, fatal lung disease with median survival of 3 years. The first study to suggest cough as an independent predictor of disease progression, reported it in 84% of the population. 1 The only study to quantify cough in 19 IPF patients, reported strong correlation between objective cough counts and cough related quality of life 2 There is a lack of studies investigating the prevalence, pathogenesis or treatment of cough in IPF. We set up a study to evaluate the prevalence of cough in an incident IPF and Non specific Interstitial Pneumonia (NSIP) population. Methods A prospective, multi-centre, observational, cohort study, PROFILE-Central England (September 2009 to June 2014) was set up. Patients had a diagnosis of definite or probable IPF or NSIP based on the ATS consensus. Leicester cough questionnaire (LCQ) was used to assess presence or absence of cough at baseline. Furthermore,the cohort was divided into 3 groups to assess severity of cough: Mild (17–20), Moderate (11–16.9) and Severe ( Results 312 incident patients with IPF or NSIP were enrolled. 261/312(83.6%) patients had incident IPF whilst 51/312(16.4%) had NSIP. The mean age of the cohort was 73.5 years (35–90 years). 235/312(76%) were males with mean age 73.7 years (47–90 years); 74/312(24%) were females with mean age of 72.9 years (35.8–88.8 years). 261/312(83.6%) reported cough compared with 51/312(16.4%) who reported no cough. Of the patients who reported cough, 45/261(17.2%) had severe cough, 112/261(42.9%) had moderate cough and 104/261(39.8%) had mild cough. There was no effect of gender, however, older cohort reported more cough (age >55 years; p = 0.014). Smoking may be a confounder, however the number of current smokers in the cohort is too small (p = 0.05). Interestingly both NSIP and IPF cohort reported cough; however, proportionally NSIP patients have less cough (14/51,27.4%) compared with IPF (37/261, 14%). Conclusions Cough occurs in a huge majority of patients with both IPF and NSIP. Cough appears to be a greater problem in older patients. References Ryerson CJ et al . Cough predicts prognosis in idiopathic pulmonary fibrosis. Respirology . 2011 Aug;16(6):969–75 Key AL et al . Objective cough frequency in Idiopathic Pulmonary Fibrosis. Cough . 2010;6:4

Introduction The bleomycin mouse mode can be used as a model of pulmonary fibrosis. The Influenza A virus can infects epithelial cells leading to cell death and injury. Acute exacerbations of Idiopathic Pulmonary Fibrosis (IPF) are characterised by epithelial cell apoptosis with unknown cause. The role of infection in acute exacerbations of IPF is unclear. The aim of this study is to investigate the effect of influenza infection on bleomycin-induced pulmonary fibrosis. Materials and Methods 60 U of bleomycin was instilled into lungs of 6–8 week old male C57Bl/6 mice. After 28 days mice were exposed intranasally with 10, 20 Units of influenza virus ‘x31’ or PBS, and lungs harvested 5 or 21 days later. Lung tissue harvested for mRNA analysis, histology and hydroxyproline levels. Animal studies were ethically reviewed and carried out in accordance with Animals (Scientific Procedures) Act 1986 and the GSK Policy on the Care, Welfare and Treatment of Animals. Results Influenza infection increased in lung collagen levels: COL1 mRNA but not COL3 was increased. There was also an increase in matrix deposition on Masson’s trichrome staining. There were increased hydroxyproline levels in influenza infected mice with fibrotic lungs due to bleomycin administration, compared with mice exposed only to bleomycin. Non-fibrotic, influenza- infected mice showed apoptosis on histological TUNEL staining. CCNA2 mRNA in influenza infected mice with fibrotic lungs was increased compared to fibrotic mice alone indicating an increase in epithelial apoptosis. Conclusion These data suggest that influenza infection may enhance the fibrotic response in the lung by promoting epithelial apoptosis and fibrogenesis.

Introduction Idiopathic Pulmonary Fibrosis (IPF) is a progressive, fatal lung disease with median survival of 3 years (1). NICE has recently approved Pirfenidone, which is the first licensed treatment for IPF. Pirfenidone is licensed for mild to moderate IPF, and the current NICE recommendation is that all patients with FVC (Forced Vital capacity) between 50–80% predicted should be considered for therapy. However, there is no agreed classification of disease severity in IPF. In clinical practice a combination of both FVC and DLCO (diffusion of lung carbon monoxide) is used to monitor disease progression. We explore the practical impact of using FVC on Pirfenidone prescribing. Methods 218 incident cases of IPF have been enrolled in a prospective cohort study PROFILE in Central England from September 2009 to June 2013. All patients had a diagnosis of definite or probable IPF based on the ATS consensus 2000. The patients were stratified according to both FVC and DLco. Lung function defects were considered mild (FVC>80%, DLCO>60%), moderate (FVC 50–80%, DLCO 40–60%) and severe (FVC Results The median age was 72 years with majority cohort males (77% males’ vs 23%females). 181(84%) cases met diagnostic criteria for definite IPF. There was only a moderate correlation between FVC and TLCO with correlation coefficient = 0.54 and p Conclusion Over half our patients eligible for Pirfenidone, based on current NICE criteria, will have severely reduced DLCO. This highlights the need for an agreed classification of IPF disease severity. Follow up of PROFILE study patients may help guide a classification system. References Navaratnam et al, Thorax 2011 Jun;66(6):462–7. doi: 10.1136/thoraxjnl-2013-204457.22

Introduction Transforming Growth Factor-beta (TGFβ) promotes anti-proliferative and pro-apoptotic pathways in lung epithelial cells, both of which have been implicated in the pathogenesis of IPF. TGFβ must be activated before it can mediate these events. Acute exacerbations of IPF are characterised by widespread epithelial cell apoptosis. The precise cause of these exacerbations is not known. The Influenza A virus is a single-stranded segmented RNA virus that infects epithelial cells leading to cell death and injury, and can also activate TGFβ. The role of infection in acute exacerbations of IPF is unclear. The aim of this study is to investigate the effect of influenza infection on bleomycin-induced pulmonary fibrosis and TGFβ activation in vivo. Materials and Methods 60 U of bleomycin was instilled into the lungs of 6–8 week old male C57Bl/6 mice. After 28 days mice were exposed intranasally with 10, 20 Units of influenza virus ‘x31’ or PBS, and the lungs harvested 5 days later. Bronchoalveolar lavage (BAL) was performed and lung tissue harvested for mRNA analysis, histology and hydroxyproline levels. All animal studies were ethically reviewed and carried out in accordance with Animals (Scientific Procedures) Act 1986 and the GSK Policy on the Care, Welfare and Treatment of Animals. Results Mice exposed to bleomycin and infected with influenza lost less weight compared with saline-exposed influenza-infected animals. However, the lungs from bleomycin-exposed, influenza-infected mice showed increased lung damage with more matrix deposition on trichrome staining than saline-exposed, influenza-infected mice. Saline-exposed, influenza-infected mice demonstrated the anticipated dose dependent increase in BAL lymphocytosis as well as apoptosis staining in histological TUNEL assessment. However, in bleomycin-exposed mice, influenza infection did not promote enhanced BAL lymphocytosis or apoptosis. However, influenza appeared to enhance the fibrotic response demonstrated by an increase in matrix deposition on masson’s trichrome and increased lung hydroxyproline levels in influenza infected bleomycin exposed mice, as early as 5 days post infection. Conclusions These data suggest that influenza infection may exacerbate lung fibrosis by promoting epithelial apoptosis.

TGF-β is fibrogenic cytokine implicated in the pathogenesis of idiopathic pulmonary fibrosis (IPF). The integrin αVβ6 can activate TGF-β, and dysregulation of this pathway is thought to play a role in the pathogenesis of pulmonary fibrosis. TGF-β induces αVβ6 integrin expression and aVb6 integrins are upregulated in fibrotic regions of lungs from patients with IPF. This raises the possibility that dysregulation of a self-regulating feedback loop may promote IPF. This study aims to investigate the mechanisms involved TGF-β-induced β6 expression and how this may be dysregulated in IPF. Using QPCR and flow cytometry we assessed expression of the integrin β6 (ITGB6) in lung epithelial cells. An ITGB6 promoter-luciferase construct and truncated mutants were used to identify the regulatory region of the promoter. Dominant negative (dn) constructs were used to assess the role of Smad proteins. Binding of transcription factors to the promoter was assessed by chromatin immunoprecipitation (ChIP). TGF-β caused concentration- and time-dependent increases in αVβ6 and ITGB6 mRNA, and increased activity of the ITGB6 promoter. Truncated mutants of the promoter showed that loss of 2 Smad binding sites resulted in loss of promoter activity. Co-transfection of dnSmad3 with the ITGB6 promoter reporter inhibited basal and TGF-β-induced promoter activity whereas dnSmad2 had little effect. dnSmad3 reduced TGF-β-induced αvβ6 cell surface expression. ChIP demonstrated binding of Smad3 and Smad4 to the promoter in response to TGF-β. Binding of Smad3 to the ITGB6 promoter was higher in lung tissue derived from IPF patients compared with controls. Finally, we identified a region of the ITGB6 promoter responsible for repressing transcription of the gene and demonstrate that siRNA targeting the transcription factor ELK1 increases αvβ6 expression. In conclusion, TGF-β increases expression of αVβ6 by transcriptional regulation involving Smad3. Furthermore, enhanced binding of Smad3 to the ITGB6 promoter in patients with IPF suggests dysregulated synthesis of αvβ6 integrins may promote IPF. Finally, we identified ELK1 as a potentially important negative regulator of αvβ6 expression. This highlights a positive feedback loop which could be dysregulated through hyperstimulation and impaired repression leading to amplification of αvβ6 mediated TGF-β activation that could be fundamental to IPF pathogenesis.

People with chronic lung disease are more susceptible to influenza infection which may lead to exacerbation of pre-existing conditions such as fibrosis. Transforming growth factor-β (TGFβ) is a profibrotic cytokine, but its role during influenza infection remains unclear. Toll-like-receptor 3 is located on the endosomal membrane and binds dsRNA, an intermediate product from replicating ssRNAviruses such as influenza. TLR3 activation has been shown to increase RhoA activity, and we have previously shown that RhoA is a key intermediary inactivation of TGFβ by the αVβ6-integrin. Therefore, we hypothesised that influenza infection could stimulate TLR3 leading to activation of latent TGFβ via this integrin in epithelial cells. Immortalised human bronchial epithelial cells (iHBECs) were used in all experiments. To determine whether influenza virus (A/PR/8/34 H1N1), or poly (I:C) (20µg/ml) were able to activate TGFβ the following TGFβ activation assays were used; detection of phospho-smad2/3 in nuclear extracts of cell lysates by ELISA; analysis of TGFβ activity in cells transiently transfected with a TGFβ-sensitive reporterconstruct; and a co-culture of iHBECS with a TGFβ reporter cell line (TMLCs). To confirm the involvement of TLR3, cells were dual transfected with a TGFβ-sensitive reporter and a dominant negative TLR3 construct designed to prevent TLR3 signalling. The role of the RhoA-ROCK pathway, and αVβ6-integrin were investigated using the ROCK inhibitor H1152, and the αVβ6-integrin blocking antibody 6.3G9, respectively. H1N1 infection and poly(I:C) caused an increase in luciferase in iHBECs transiently transfected with a TGFβ reporter construct. Similarly, both H1N1 and poly(I:C) caused an increase in nuclear phospho-smad2/3 which could be blocked by 6.3G9 peaking at 4h. Both agents caused an increase in TGFβ as measured by a co-culture assay and this could be blocked by H1152 and 6.3G9 suggesting the involvement of ROCK, αVβ6-integrin and the requirement for cell-to-cell contact. Finally, arole for TLR3 in this process was confirmed in cells transfected with a dnTLR3 construct which lost the ability to activate TGFβ in response to poly(I:C) orH1N1. In conclusion, these data show that both influenza A and poly (I:C) lead to increased TGFβ activity in iHBECs. This supports the hypothesis that influenza A infection activates TGFβ via TLR3 and the αVβ6 integrin. These data suggest anovel mechanism by which influenza infection of epithelial cells may promoteairway and lung fibrosis.

Introduction and Objective Idiopathic pulmonary fibrosis (IPF) is a chronic, irreversible fibrotic disease with more than 5000 incident cases annually in UK. IPF related cough is present in 80% cases and is often refractory to current treatments. Cough has a significant impact on quality of life, causing sleep disturbance, difficulties at work and stress incontinence. A previously published prospective open label phase II trial of thalidomide for treatment of pulmonary fibrosis suggested that IPF associated cough responded well to thalidomide. We have been using thalidomideas an “off-license” indication for selected IPF patients. The objective of this study was to review our experience of using Thalidomide as treatment for cough in IPF. Methods Nine patients were referred to Nottingham Academic Interstitial Lung Disease clinic between 2009 and 2011 for assessment of their cough. A modified version of Leicester Cough Questionnaire was used, in conjunction with subjective symptoms, to clinically assess their cough. A trial of PPI (omeprazole 40 mg) and Prednisolone (10 mg) for 6 weeks was given to all subjects. Two were excluded as did not have significant cough, one patient declined thalidomide after initial screening. Results Six patients were treated with thalidomide. Four had IPF, one had Hypersensitivity Pneumonitis and one had fibrotic Cryptogenic Organising Pneumonia. 72% were males with mean age 69 years (range 51–88 years). The median pre-thalidomide cough score was 74.5 (IQR 13.25) and post treatment cough score was 51.5 (IQR 49.25). This was statistically significant (p=0.046). Three stopped thalidomide subsequent to rash. Two patients are currently stable with 50 mg once daily, and 1 with 50 mg alternate daily of thalidomide. Conclusion Our observation of a carefully selected cohort of patients suggests that thalidomide has potential for treatment of IPF associated cough. It works quickly (within days) as reflected both subjectively and objectively via questionnaire, even when prednisolone has failed. However, it does have a significant side-effect profile. Identification of thalidomide9s mechanism of action may aid the development of novel, effective anti-tussive therapy for this debilitating aspect of IPF. We will assess this in randomised open label trial comparing thalidomide vs prednisolone for treatment of IPF associated cough.

Airway remodelling is a common feature of severe asthma. Transforming growth factor-β (TGF-β) is a pro-fibrotic, pleiotropic cytokine implicated in airway remodelling. TGF-β is sequestered in the extracellular matrix as a latent complex and requires activation to function. Lysophosphatidic acid (LPA) causes TGF-β activation in airway epithelial cells. The study aims were to investigate the effect of LPA on TGF-β activation by ASM cells in asthma. TGF-β activation was assessed by a reporter cell co-culture assay, by determining expression of the TGF-β-inducible gene plasminogen activator inhibitor-1 (PAI1) and by detecting the nuclear translocation of Smad proteins. The effect of LPA on TGF-β activation in asthma was investigated by comparing the responses of ASM cells from non-asthmatic (n=3) and asthmatic (n=3) donors. TGFb activation was also assessed using a chronic ovalbumin model of airway remodelling in mice. LPA induced a time, and concentration, dependent increase in TGF-β activation by ASM cells that was abrogated by an integrin αVβ5 antibody. An inhibitor of cytoskeletal reorganisation inhibited the effects of LPA. Furthermore, the β2-agonist formoterol inhibited LPA-induced PAI1 expression. Primary asthmatic ASM cells activated more TGF-β via αVβ5 in response to LPA than control cells, however they did not express more αVβ5 on the cell surface. Phosphorylation of Smad2 and expression ofpai1 in the lungs was increased in a chronic ovalbumin model of asthmatic airway remodelling in mice. Furthermore, αVβ5 integrin staining and α-smooth muscle actin staining in the ASM layer around the airways is increased in this model. Collectively, these data show that ASM cells can activate TGF-β via the αVβ5 integrin and highlight a novel pathway of TGF-β activation in ASM cells, which may be important in development of asthmatic airway remodelling.