Bronchoalveolar Lavage Cell Counts Research Articles

Benefits and risks of bronchoalveolar lavage in severe asthma in children.

BackgroundAlthough bronchoscopy can be part of the exploration of severe asthma in children, the benefit of bronchoalveolar lavage (BAL) is unknown. The present study aimed to decipher whether systematic BAL during a flexible bronchoscopy procedure could better specify the characteristics of severe asthma and improve asthma management.Material and methodsThe study took place in two departments of a university hospital in Paris. Children who underwent flexible bronchoscopy for the exploration of severe asthma between April 2017 and September 2019 were retrospectively included.ResultsIn total, 203 children were included, among whom 107 had a BAL. BAL cell count was normal in most cases, with an increasing number of eosinophils with age, independently from the atopic status of the patients. Compared with bronchial aspiration only, BAL increased the rate of identified bacterial infection by 1.5. Nonatopic patients had more bacterial infections (p<0.001). BAL induced a therapeutic modification only for azithromycin and omalizumab prescriptions. The practice of a BAL decreased bronchoscopy tolerance (p=0.037), especially in the presence of tracheobronchial malacia (p<0.01) and when performed in a symptomatic patient (p=0.019).Discussion and conclusionAlthough BAL may provide interesting information in characterising severe asthma, in most cases its impact on the patient's management remains limited. Moreover, BAL can be poorly tolerated and should be avoided in the case of tracheobronchial malacia or current asthma symptoms.

SNAP, CRACKLE, AND POP: A CASE OF PEMETREXED-INDUCED INTERSTITIAL PNEUMONITIS

TOPIC: Diffuse Lung Disease TYPE: Medical Student/Resident Case Reports INTRODUCTION: Pemetrexed is a folic acid inhibitor which is used for the treatment of non-small cell lung cancer (NSCLC) (1). Pemetrexed is generally well tolerated, but interstitial pneumonitis is an exceedingly rare, potentially fatal adverse effect (2). The diagnosis of drug induced interstitial pneumonitis is one of exclusion and based on nonspecific clinical and radiographic findings. Onset of symptoms must correlate clinically with timing of administration of the offending agent, and the symptoms improve with removal of the medication and initiation of corticosteroids (3). It is important for clinicians to have a high index of suspicion to initiate early treatment, reducing morbidity and mortality. Below is a case of a patient with lung adenocarcinoma who developed pemetrexed induced interstitial pneumonitis. CASE PRESENTATION: Our patient is an 85 year old man who was diagnosed with stage IVA lung adenocarcinoma. He received two cycles of Carboplatin and Pemetrexed with good radiologic response but experienced intolerable side effects. The decision was made to transition to Pemetrexed monotherapy. Two weeks after his second cycle, he developed dyspnea, low grade fevers and fatigue, requiring supplemental oxygen (Vapotherm 60% FiO2 at 40 L/min) and hospital admission. Physical exam was notable for diffuse crackles bilaterally. CTA Chest demonstrated bilateral ground glass opacities and was negative for pulmonary embolism. He was empirically started on broad spectrum antibiotics for multifocal pneumonia. COVID-19, respiratory viral test, blood and sputum cultures were all negative. Bronchoscopy with bronchoalveolar lavage (BAL) demonstrated normal airways. There was no evidence of alveolar hemorrhage. BAL cell count and differential was notable for a lymphocytic predominance. BAL cytology demonstrated pulmonary macrophages and mixed inflammatory cells but no atypical cells. High dose corticosteroids were started on hospital day five. Oxygen requirement steadily decreased, and he was discharged a few days later on 2 L NC and a prolonged corticosteroid taper. DISCUSSION: Pemetrexed is a widely used first-line treatment for NSCLC. Pemetrexed-induced interstitial pneumonitis has a range of clinical presentations, making it challenging to diagnose. Presentations vary from asymptomatic to life-threatening requiring mechanical ventilation. Moreover, it is a very rare side effect of Pemetrexed. This case highlights the importance of having strong clinical suspicion so that interstitial pneumonitis can be identified and treated early, mitigating morbidity and mortality. A thorough workup must be completed to rule out other causes, especially infection. CONCLUSIONS: Through early diagnosis, pemetrexed can be discontinued and corticosteroids can be initiated sooner, reducing risk to the patient and improving clinical outcomes (3). The patient should not be rechallenged with Pemetrexed, as this could be fatal. REFERENCE #1: Dhakal, B., Singh, V., Shrestha, A., Rao, A., & Choong, N. (2011). Pemetrexed induced pneumonitis. Clinics and Practice 1(4), 232-6. DOI: 10.4081/cp.2011.e106 REFERENCE #2: Kim, KH. et al. (2013). Interstitial pneumonitis after treatment with pemetrexed for non-small cell lung cancer. Cancer Research and Treatment 45(1), 74-7. DOI: 10.4143/crt.2013.45.1.74 REFERENCE #3: Skeoch, S. et al. (2018). Drug-induced interstitial lung disease: A systematic review. Journal of Clinical Medicine 7(10), 1-30. DOI: 10.3390/jcm7100356 DISCLOSURES: No relevant relationships by Brent Bagley, source=Web Response No relevant relationships by Ethan Tope, source=Web Response

ACUTE EOSINOPHILIC PNEUMONIA: COMPLICATIONS OF SMOKE EXPOSURE AND VAPING-ASSOCIATED LUNG INJURY

TOPIC: Occupational and Environmental Lung Diseases TYPE: Medical Student/Resident Case Reports INTRODUCTION: E-cigarette or vaping product use-associated lung injury (EVALI) encompass a host of pulmonary complications including diffuse alveolar hemorrhage, lipoid pneumonia, hypersensitivity pneumonitis, and rarely, acute eosinophilic pneumonia (AEP) [1]. AEP presents as an acute febrile illness with hypoxemia, diffuse pulmonary infiltrates, and eosinophilia on bronchoalveolar lavage (BAL) without evidence of prior infection or atopic illness [2]. Presented is a young adult patient with a history of continuous vaping and extended exposure to kitchen smoke who developed acute hypoxemic respiratory failure later diagnosed as AEP. CASE PRESENTATION: An 18-year-old male with no significant past medical history presented to the emergency department with acute onset malaise, fever, non-productive cough, and shortness of breath over 12 hours. Social history revealed 3-week workplace exposure to smoke as a barbeque cook and a 5-year history of daily vaping with reported cessation 3 weeks prior. Vitals were notable for hypoxemia which was corrected with administration of 8L of oxygen via nasal cannula. Quickly after admission, he had worsening respiratory status: tachypnea, increased oxygen requirement, use of accessory muscles, and bilaterally diminished breath sounds. He was transferred to the medical intensive care unit for emergent endotracheal intubation. Laboratory testing revealed a neutrophil predominant leukocytosis of 19.3x109/L. COVID-19, rapid influenza A and B, and urine antigens for streptococcus and legionella were negative as were autoimmune serologies. A computed tomography (CT) scan of the chest revealed diffuse bilateral consolidative opacities [Figure 1] with worsening over the next 24 hours [Figure 2]. Further investigation was undertaken with bronchoscopy. A BAL was obtained, and cell count and cultures were sent. BAL cell count was remarkable for 33% eosinophilia. The diagnosis of AEP was made, and the patient was started on systemic glucocorticoids, with significant improvement within 24 hours. DISCUSSION: The modified Philit criteria is used to make the diagnosis of AEP, which the patient satisfied [3]. Although a 5-year vaping history was present, no previous significant respiratory symptoms were reported. Prior studies have demonstrated a relationship between workplace smoke exposure and AEP [3]. Based on the modified Philit criteria, we suspect that the patient had acute respiratory failure from AEP as a result of chronic EVALI acutely exacerbated by workplace smoke exposure. The treatment of AEP treatment involves high-dose intravenous glucocorticoids followed by a prolonged oral steroid taper. CONCLUSIONS: Obtaining occupation history as well as smoke exposure is as important as obtaining vaping history in an otherwise healthy young patient who presents with acute hypoxemic respiratory failure with bilateral diffuse opacities on imaging. REFERENCE #1: Winnicka L, Shenoy MA. EVALI and the Pulmonary Toxicity of Electronic Cigarettes: A Review. J Gen Intern Med. 2020;35(7):2130-2135. doi:10.1007/s11606-020-05813-2 REFERENCE #2: Allen JN, Pacht ER, Gadek JE, Davis WB. Acute Eosinophilic Pneumonia as a Reversible Cause of Noninfectious Respiratory Failure. N Engl J Med. 1989;321(9):569-574. doi:10.1056/nejm198908313210903 REFERENCE #3: Philit F, Etienne-Mastroïanni B, Parrot A, Guérin C, Robert D, Cordier JF. Idiopathic acute eosinophilic pneumonia: A study of 22 patients. Am J Respir Crit Care Med. 2002;166(9):1235-1239. doi:10.1164/rccm.2112056 DISCLOSURES: No relevant relationships by Nurjahan Khatun, source=Web Response No relevant relationships by Parth Patel, source=Web Response No relevant relationships by Naleen Patel, source=Web Response No relevant relationships by Tarang Patel, source=Web Response No relevant relationships by SACHIN PATIL, source=Web Response No relevant relationships by Shyam Shankar, source=Web Response No relevant relationships by Rodger Wilhite, source=Web Response

Cytotoxic action reduction of crystalline silicon dioxide in a toxicological experiment associated with Lymphomyosot

Introduction. Among the most relevant issues in occupational health is the risk management of the contamination of working air by crystalline silicon dioxide (CSD) in enterprises dealing with its extraction and processing. Cytotoxicity of this silicon type that underlies its fibrogenic action is of particular concern. Thereby, it is an urgent goal to find drugs that increase the body’s resistance to cytotoxic effects of silicon dioxide. Materials and methods. For the study of changes in the cellular composition of the bronchoalveolar lavage (BAL) and some of its biochemical characteristics, a respirable portion of Pervouralsk quartzite (PQ) containing 98% CSD was inserted intratracheally to outbred white female rats in the form of a suspension of 10 mg of PQ in 1 ml of distilled water in the dose of 1 ml. Student’s t-test was applied for statistical analysis of the obtained data. Results. PQ sample induces the alveolar phagocytosis response typical for cytotoxic dust expressed by an increase in the total BAL cell count, the number of both alveolar macrophages (AM) and neutrophilic leukocytes (NL), and particularly the NL/AM ratio (which is a reliable comparative index of cytotoxicity). The administration of «Lymphomyosot» considerably reduced this cytotoxic effect of PQ within one month before the intratracheal injection. It showed a decrease in NL/AM ratio and some biochemical indicators of cytolysis in BAL. Conclusion. There was demonstrated the applicability of «Lymphomyosot» as a precaution against the cytotoxic effect of CSD dust.

Mine-site derived particulate matter exposure exacerbates neurological and pulmonary inflammatory outcomes in an autoimmune mouse model

ABSTRACT The Southwestern United States has a legacy of industrial mining due to the presence of rich mineral ore deposits. The relationship between environmental inhaled particulate matter (PM) exposures and neurological outcomes within an autoimmune context is understudied. The aim of this study was to compare two regionally-relevant dusts from high-priority abandoned mine-sites, Claim 28 PM, from Blue Gap Tachee, AZ and St. Anthony mine PM, from the Pueblo of Laguna, NM and to expose autoimmune-prone mice (NZBWF1/J). Mice were randomly assigned to one of three groups (n = 8/group): DM (dispersion media, control), Claim 28 PM, or St. Anthony PM, subjected to oropharyngeal aspiration of (100 µg/50 µl), once per week for a total of 4 consecutive doses. A battery of immunological and neurological endpoints was assessed at 24 weeks of age including: bronchoalveolar lavage cell counts, lung gene expression, brain immunohistochemistry, behavioral tasks and serum autoimmune biomarkers. Bronchoalveolar lavage results demonstrated a significant increase in number of polymorphonuclear neutrophils following Claim 28 and St. Anthony mine PM aspiration. Lung mRNA expression showed significant upregulation in CCL-2 and IL-1ß following St. Anthony mine PM aspiration. In addition, neuroinflammation was present in both Claim 28 and St. Anthony mine-site derived PM exposure groups. Behavioral tasks resulted in significant deficits as determined by Y-maze new arm frequency following Claim 28 aspiration. Neutrophil elastase was significantly upregulated in the St. Anthony mine exposure group. Interestingly, there were no significant changes in serum autoantigens suggesting systemic inflammatory effects may be mediated through other molecular mechanisms following low-dose PM exposures.

Carboxypeptidase inhibition by NvCI suppresses airway hyperreactivity in a mouse asthma model

Carboxypeptidase inhibition by NvCI suppresses airway hyperreactivity in a mouse asthma model

Concordance of Bronchoalveolar Lavage Cytology with Transbronchial Lung Biopsies in Non-neoplastic Pulmonary Diseases

Introduction: In many pulmonary diseases, despite radiological &amp; clinical investigations, laboratory tests and function studies, the diagnosis becomes difficult. Bronchoalveolar Lavage (BAL) is a minimally invasive method in which cells are collected from bronchial and alveolar spaces for cytology. This is facilitated by using a flexible bronchoscope with which a biopsy is taken following BAL. Bronchoscopy with BAL when used appropriately can offer correct diagnosis which in turn aids in proper management of the patient. Aim: To find the concordance of BAL findings with the histopathological features of Transbronchial Lung Biopsy (TBLB) in non-neoplastic lung diseases. Materials and Methods: It was a retrospective study conducted in Kempegowda Institute of Medical Sciences, Bengaluru, Karnataka. A total of 40 patients presenting with clinico-radiological findings, suggesting a non-neoplastic lung disease in the year 2019, undergoing bronchoscopy with BAL and concurrent TBLB were chosen. The BAL fluid was processed and differential count of cells was done to classify according to the American Thoracic Society Guidelines. Concordance was checked between the diagnoses made on TBLB and BAL analysis. Results: In the present study, a total of 40 cases were included of which 13 (32.5%) cases showed neutrophilic, 16 (40%) cases showed lymphocytic, 5 (12.5%) cases showed eosinophilic and 6 (15%) cases showed normal cellular distribution on BAL cytology. Diagnoses on studying TBLB included nine cases of Nonspecific Interstitial Pneumonia, seven cases of Usual Interstitial Pneumonia, six cases of Bronchiolitis Obliterans Organising Pneumonia, three cases of Bronchiolitis, two cases each of pulmonary tuberculosis and granulomatous inflammation. There was one case each of actinomycosis, sarcoidosis, lung abscess and mucor mycosis. Normal histology was noted in seven cases. The sensitivity of BAL fluid analysis was found to be 84.84% and the concordance was 80%. The Kappa value obtained was 0.71 indicating good agreement/concordance between BAL cytology and TBLB. Conclusion: The data from the current study suggest that differential cell counts in BAL provide diagnostic information of fundamental importance in frequently occurring non-neoplastic lung diseases in the community.

K2P2.1 (TREK-1) potassium channel activation protects against hyperoxia-induced lung injury

No targeted therapies exist to counteract Hyperoxia (HO)-induced Acute Lung Injury (HALI). We previously found that HO downregulates alveolar K2P2.1 (TREK-1) K+ channels, which results in worsening lung injury. This decrease in TREK-1 levels leaves a subset of channels amendable to pharmacological intervention. Therefore, we hypothesized that TREK-1 activation protects against HALI. We treated HO-exposed mice and primary alveolar epithelial cells (AECs) with the novel TREK-1 activators ML335 and BL1249, and quantified physiological, histological, and biochemical lung injury markers. We determined the effects of these drugs on epithelial TREK-1 currents, plasma membrane potential (Em), and intracellular Ca2+ (iCa) concentrations using fluorometric assays, and blocked voltage-gated Ca2+ channels (CaV) as a downstream mechanism of cytokine secretion. Once-daily, intra-tracheal injections of HO-exposed mice with ML335 or BL1249 improved lung compliance, histological lung injury scores, broncho-alveolar lavage protein levels and cell counts, and IL-6 and IP-10 concentrations. TREK-1 activation also decreased IL-6, IP-10, and CCL-2 secretion from primary AECs. Mechanistically, ML335 and BL1249 induced TREK-1 currents in AECs, counteracted HO-induced cell depolarization, and lowered iCa2+ concentrations. In addition, CCL-2 secretion was decreased after L-type CaV inhibition. Therefore, Em stabilization with TREK-1 activators may represent a novel approach to counteract HALI.

Echinacoside attenuates lipopolysaccharide-induced acute lung injury in newborn mice via inactivation of NF- κB/NLRP3 signaling pathway

Purpose: To investigate the effect of echinacoside (ECH) on acute lung injury (ALI) and the underlying mechanism of action.Methods: The ALI model was established through intranasal instillation of lipopolysaccharide (LPS). Lung tissue damage was determined using hematoxylin and eosin (H&amp;E) staining and lung wet-to-dry–weight ratio. Bronchoalveolar lavage fluid (BALF) protein concentration, cell count, and cytokine level were evaluated. Western blotting was used to determine protein expression level.Results: ECH attenuated lung tissue injury and lung wet-to-dry–weight ratio in the ALI model (p &lt; 0.01). The total protein content and number of total cells, neutrophils, and macrophages increased in BALF of mice treated with LPS, but these increases were reversed by ECH treatment (p &lt; 0.01). The levels of TNF-α and IL-1β increased in BALF and lung tissue of LPS-treated mice; however, ECH treatment decreased these changes (p &lt; 0.01). In addition, ECH inhibited the activation of the nuclear factor-κB (NF-κB)/NLR family pyrin domain containing 3 (NLRP3) pathway in LPS-treated mice (p &lt; 0.01).Conclusion: Echinacoside attenuates LPS-induced ALI via inactivation of the NF-κB/NLRP3 pathway, making echinacoside a potential drug for the treatment of ALI.&#x0D; Keywords: Echinacoside, Acute lung injury, Lipopolysaccharide, Nuclear factor-κB, NLR family pyrin domain containing 3

English

Uncaria guianensis (Aubl.) J. F. Gmel. (&ldquo;cat&rsquo;s claw&rdquo;, Rubiaceae) is a plant with potential to treat asthma because of its anti-inflammatory and antioxidant activities. The aim of this study was to evaluate the effects of two extracts of U. guianensis in an animal model of allergic asthma. Balb/c mice were sensitized twice with ovalbumin intraperitoneally one week apart, then challenged with intranasal ovalbumin for three days. Animals were treated with aqueous or hydroethanolic extracts (100 mg/kg) for three days, simultaneously with ovalbumin challenges. Control mice received saline solution on the same days. In vivo bronchial hyper responsiveness, airway and lung inflammation, IgE levels, and total antioxidant capacity were measured. Treatment with the hydroethanolic extract significantly reduced total cell and eosinophil counts in bronchoalveolar lavage, and in vivo bronchial hyper responsiveness. Moreover, U. guianensis hydroethanolic extract significantly reduced interleukin 13 levels in lung homogenate. Total antioxidant capacity and IgE serum levels were not affected with the extract administration. Of note, treatment with the aqueous extract did not elicit significant effects on asthma-like characteristics. Only the hydroethanolic extract of U. guianensis reduced lung inflammation and bronchial hyper responsiveness in asthmatic mice. Key words: Anti-inflammatory agents, asthma, oxindolics, phenols, respiratory hypersensitivity, Uncaria guianensis, Rubiaceae. &nbsp

17β-Estradiol Treatment Protects Lungs Against Brain Death Effects in Female Rat Donor.

Brain death (BD) affects the viability of lungs for transplantation. A correlation exists between high-lung inflammation after BD and the decrease in female sex hormones, especially estradiol. Therefore, we investigated the effects of 17β-estradiol (E2) treatment on the lungs of female brain dead rats. Female Wistar rats were divided into 4 groups: BD (submitted to BD for 6 h), sham (false operated), E2-T0 (treated with E2 immediately after BD; 50 μg/mL, 2 mL/h), and E2-T3 (treated with E2 after 3 h of BD; 50 μg/mL, 2 mL/h). Lung edema, hemorrhage, and leukocyte infiltration were analyzed. Adhesion molecules were evaluated, and analysis of NO synthase gene and protein expression was performed using real-time PCR and immunohistochemistry, respectively. Release of chemokines and matrix degradation in the lungs was analyzed. BD increased leukocyte infiltration, as shown by intravital microscopy (P = 0.017), bronchoalveolar lavage cell count (P = 0.016), the release of inflammatory mediators (P = 0.02), and expression of adhesion molecules. BD also increased microvascular permeability and the expression and activity of matrix metalloproteinase-9 in the lungs. E2 treatment reduced leukocyte infiltration, especially in the E2-T3 group, release of inflammatory mediators, adhesion molecules, and matrix metalloproteinase activity in the lungs. E2 treatment was successful in controlling the lung inflammatory response in females submitted to BD. Our results suggest that E2 directly decreases the release of chemokines, restraining cell traffic into the lungs. Thus, E2 has a therapeutic potential, and its role in improving donor lung quality should be explored further.

Comparison of respiratory toxicity of TiO2 and Fe3O4 nanoparticles after intravenous instillation: an experimental study.

Nanomaterials consist of particles smaller than 100 nm - nanoparticles (NPs). Their nano dimensions allow them to penetrate through various membranes and enter into the bloodstream and disseminate into different body organs. Massive expansion of nanotechnologies together with production of new nanoparticles which have not yet been in contact with living organisms may pose a potential health problem. It is therefore necessary to investigate the health impact of NPs after experimental exposure. Comparison of the effect of TiO2 and NPs Fe3O4 in Wistar rats at time intervals 1, 7, 14 and 28 days was performed by studying the cytotoxic effect in the isolated inflammatory cells from bronchoalveolar lavage (BAL). Wistar rats were intravenously (i.v.) given a suspension of NPs TiO2 or Fe3O4 (coated by sodium oleate) via the tail vein. After time intervals of 1, 7, 14 and 28 days, we sacrificed the animals under anaesthesia, performed BAL and isolated the cells. The number of animals in the individual groups was 7-8. We examined the differential count of BAL cells (alveolar macrophages - AM, polymorphonuclear leukocytes - PMN, lymphocytes - Ly); viability and phagocytic activity of AM; the proportion of immature and polynuclear cells and enzymes - cathepsin D - CAT D, lactate dehydrogenase - LDH and acid phosphatase - ACP. We found that TiO2 NPs are relatively inert - without induction of inflammatory and cytotoxic response. Exposure to nanoparticles Fe3O4 induced - under the same experimental conditions - in comparison with the control and TiO2 a more extensive inflammatory and cytotoxic response, albeit only at 1, 7 and 14 days after injection. The results suggest that TiO2 and Fe3O4 nanoparticles used in our study were transferred from the bloodstream to the respiratory tract, but this effect was not observed at 28 days after i.v. injection, probably due to their removal from the respiratory tract.

Antiretroviral therapy does not reduce tuberculosis reactivation in a tuberculosis-HIV coinfection model.

While the advent of combination antiretroviral therapy (ART) has significantly improved survival, tuberculosis (TB) remains the leading cause of death in the HIV-infected population. We used Mycobacterium tuberculosis/simian immunodeficiency virus-coinfected (M. tuberculosis/SIV-coinfected) macaques to model M. tuberculosis/HIV coinfection and study the impact of ART on TB reactivation due to HIV infection. Although ART significantly reduced viral loads and increased CD4+ T cell counts in blood and bronchoalveolar lavage (BAL) samples, it did not reduce the relative risk of SIV-induced TB reactivation in ART-treated macaques in the early phase of treatment. CD4+ T cells were poorly restored specifically in the lung interstitium, despite their significant restoration in the alveolar compartment of the lung as well as in the periphery. IDO1 induction in myeloid cells in the inducible bronchus-associated lymphoid tissue (iBALT) likely contributed to dysregulated T cell homing and impaired lung immunity. Thus, although ART was indispensable for controlling viral replication, restoring CD4+ T cells, and preventing opportunistic infection, it appeared inadequate in reversing the clinical signs of TB reactivation during the relatively short duration of ART administered in this study. This finding warrants the modeling of concurrent treatment of TB and HIV to potentially reduce the risk of reactivation of TB due to HIV to inform treatment strategies in patients with M. tuberculosis/HIV coinfection.

Imaging Biomarkers and Pathobiological Profiling in a Rat Model of Drug-Induced Interstitial Lung Disease Induced by Bleomycin.

A large number of systemically administered drugs have the potential to cause drug-induced interstitial lung disease (DIILD). We aim to characterize a model of DIILD in the rat and develop imaging biomarkers (IBs) for detection and quantification of DIILD. In this study, Sprague–Dawley rats received one single dose of intratracheal (i.t.) bleomycin and were longitudinally imaged at day 0, 3, 7, 14, 21, and 28 post dosing, applying the imaging techniques magnetic resonance imaging (MRI) and positron emission tomography (PET)/computed tomography (CT). Bronchoalveolar lavage fluid (BALF) was analyzed for total protein and inflammatory cells. Lungs were saved for further evaluation by gene analysis using quantitative-PCR and by histology. Lung sections were stained with Masson’s-Trichrome staining and evaluated by modified Ashcroft score. Gene expression profiling of inflammatory and fibrotic markers was performed on lung tissue homogenates. Bleomycin induced significant increase in total protein concentration and total cell count in bronchoalveolar lavage (BAL), peaking at day 3 (p > 0.001) and day 7 (p > 0.001) compared to control, respectively. Lesions measured by MRI and PET signal in the lungs of bleomycin challenged rats were significantly increased during days 3–14, peaking at day 7. Two subgroups of animals were identified as low- and high-responders by their different change in total lung volume. Both groups showed signs of inflammation initially, while at later time points, the low-responder group recovered toward control, and the high-responder group showed sustained lung volume increase, and significant increase of lesion volume (p < 0.001) compared to control. Lastly, important inflammatory and pro-fibrotic markers were assessed from lung tissue, linking observed imaging pathological changes to gene expression patterns. In conclusion, bleomycin-induced lung injury is an adequate animal model for DIILD studies and for translational lung injury assessment by MRI and PET imaging. The scenario comprised disease responses, with different fractions of inflammation and fibrosis. Thereby, this study improved the understanding of imaging and biological biomarkers in DIILD and lung injury.

Analysis of particles from hamster lungs following pulmonary talc exposures: implications for pathogenicity

BackgroundTalc, a hydrous magnesium silicate, often used for genital hygiene purposes, is associated with ovarian carcinoma in case-control studies. Its potential to cause inflammation, injury, and functional changes in cells has been described. A complication of such studies is that talc preparations may be contaminated with other materials. A previous study by (Beck et al. Toxicol Appl Pharmacol 87:222-34, 1987) used a hamster model to study talc and granite dust exposure effects on various biochemical and cellular inflammatory markers. Our current study accessed key materials used in that 1987 study; we re-analyzed the original talc dust with contemporary scanning electron microscopy and energy dispersive x-ray analysis (SEM/EDX) for contaminants. We also examined the original bronchoalveolar lavage (BAL) cells with polarized light microscopy to quantify cell-associated birefringent particles to gain insight into the talc used.ResultsSEM/EDX analyses showed that asbestos fibers, quartz, and toxic metal particulates were below the limits of detection in the original talc powder. However, fibers with aspect ratios ≥3:1 accounted for 22% of instilled material, mostly as fibrous talc. Talc (based on Mg/Si atomic weight % ratio) was the most abundant chemical signature, and magnesium silicates with various other elements made up the remainder. BAL cell counts confirmed the presence of acute inflammation, which followed intratracheal instillation. Measurements of cell associated birefringent particles phagocytosis revealed significant differences among talc, granite, and control exposures with high initial uptake of talc compared to granite, but over the 14-day experiment, talc phagocytosis by lavaged cells was significantly less than that of granite. Phagocytosis of talc fibers by macrophages was observed, and birefringent particles were found in macrophages, neutrophils, and multinucleate giant cells in lavaged cells from talc-exposed animals.ConclusionOur data support the contention that talc, even without asbestos and other known toxic contaminants, may elicit inflammation and contribute to lung disease. Our findings support the conclusions of (Beck et al. Toxicol Appl Pharmacol 87:222-34, 1987) study. By analyzing particulate exposures with polarized light microscopy and SEM/EDX, fibrous talc was identified and a distinctive pattern of impaired particulate ingestion was demonstrated.

CORRELATION OF ENVIRONMENTAL MICROBIAL DIVERSITY WITH AIRWAY DISEASE IN REFRACTORY ASTHMA

TYPE: Abstract Publication TOPIC: Allergy and Airway PURPOSE: The influence of subacute bacterial infection in refractory asthma remains elusive. Bacterial dysbiosis and potentially environmental microbe exposures contribute to disease in asthma. We hypothesize that biodiversity within home environments influences airway microbiome and control of disease in refractory asthma. METHODS: We examined the microbiome of bronchoalveolar lavage (BAL) samples and home dust specimens of 5 refractory asthmatics, at time points 12 months apart. A custom, aggressive genomic DNA extraction was used to obtain a more diverse composition of microbial DNA. Bacterial 16S rDNA (V4-V5) was amplified, packaged into libraries, and sequenced in a multiplex manner using 2X300bp Illumina chemistry. Sequences were processed and profiled using QIIME2. Clinical parameters collected include BAL cell count differential, cultures, Asthma Control Test (ACT) and FEV1. RESULTS: Two subjects had positive BAL cultures, one with rare Pseudomonas and another with Mycobacterium. Follow-up bronchoscopy showed culture resolution and reduced airway neutrophilia after directed treatment. Subjects with clinical improvement (3 of 5) trended toward a significant increase in BAL biodiversity by genus (p=0.065, two-tailed T-test), and reduced environmental microbial DNA at month 12. Subjects without clinical improvement of asthma (2 of 5) demonstrated predominant pathogenic bacterial DNA and negative BAL cultures, therefore untreated clinically. CONCLUSIONS: Preliminary analysis of environmental and airway microbiome suggests asthma control is dependent on treatment of clinical and sub-clinical microbes. Higher airway and lower environmental biodiversity may correlate with improved asthma control. CLINICAL IMPLICATIONS: Our research shows clinical cultures are sub-optimal in identifying bacterial species in the lower airways. DISCLOSURE: No significant relationships. KEYWORDS: airway microbiome, refractory asthma

Modeling Immune Effector Responses in Alveolar Epithelial Cell Driven Lung Injury and Fibrosis

Abstract Mutations in the Surfactant Protein-C gene [SP-C] have been linked to pulmonary fibrosis (PF), with the missense isoleucine to threonine substitution at position 73 [I73T] as the most common. Clinically, PF is characterized by unrelenting tissue scarring and respiratory failure, intermitted by episodic exacerbations, bouts of inflammatory cell influx of unknown cause, which drastically reduce patient prognosis. Current murine models of PF rely on exposure to toxic agents or overexpression of pro-fibrotic signaling molecules in order to generate fibrotic endpoints. This reduces the overall relevance to human disease. To address these shortcomings, we developed a murine model of fibrosis designed to expressed mutant SP-CI73T. The resulting phenotype is accompanied by elevations in bronchoalveolar lavage cell counts, diffuse alveolar damage, and mortality. RNA and protein analysis demonstrate SP-CI73T epithelial cells initiate immune cell recruitment via CCL-2, IL-5, CCL-11, and CCL-17. Flow cytometric analysis highlighted temporally definite waves of inflammatory cells: an initial Ly6C+ inflammatory monocytes influx (3d), transient neutrophilic influx, and subsequent alveolar eosinophilia (2wk). RNA-seq analysis of sorted Ly6C+ monocytes revealed both inflammatory (inos, Il-6) and pro-fibrotic (col1a1, col1a2) phenotype. Pharmacologic and genetic monocyte ablation reduced inflammatory burden and improved survival of SP-CI73T mice. Taken together, we show that our mutant SP-CI73T model recapitulates the clinical features of PF. These findings support the concept that epithelial dysfunction is a key component of fibrogenesis and that inflammatory monocytes is responsible for disease exacerbation.

P45 Interstitial pneumonitis induced by secukinumab therapy in a patient with psoriatic arthritis

Abstract Background Inhibition of pro-inflammatory interleukin-17A (IL-17A) with secukinumab (an anti-IL-17A antibody) has been utilised in patients with chronic autoimmune disease such as psoriasis seronegative spondyloarthopathies. In mouse models, IL-17A was involved in the development of pulmonary inflammation and progression of fibrosis, with blockade of IL-17A implicated as a potential therapeutic target in such conditions. Here we report a case of, seemingly paradoxical, interstitial lung disease (ILD) secondary to secukinumab therapy in a 61-year-old patient with psoriatic arthritis. Methods Please refer to the results section. Results Initial therapy was with adalimumab achieving good disease control for 9 years, at which point his arthritis began to progress. Secukinumab was commenced with good clinical response from his arthritis, however he developed a persistent cough with blood-stained secretions, and shortness of breath on exertion. A CT thorax was arranged at 21 weeks after initiation of secukinumab, and showed florid bibasal, lingula and middle lobe ground glass density air space opacification. He went on to have bronchoscopy with bronchoalveolar lavage (BAL) and differential cell count. BAL was negative for atypical infection including P. jirovecii, growing upper respiratory tract flora only. Microscopy of BAL showed lymphocytosis in keeping with the diagnosis of drug induced ILD, with no atypical cells. The patient made a significant improvement on withdrawal of secukinumab therapy within 7-8 weeks of cessation - both clinically and radiologically. Conclusion A review of the current literature at this time indicates this is now a third case of secukinumab induced ILD. Given this potentially idiosyncratic reaction we suggest further work may be necessary to understand the underlying immunogenic potential of secukinumab, and an increased awareness as well as vigilance in reporting, to determine those at-risk populations of this phenomenon. Disclosures C. Saleh None. S. Bilgrami None. T. Gatheral None. L. Ottewell None.

Mammalian Target of Rapamycin Signaling Enhances Ovalbumin-Induced Neutrophilic Airway Inflammation by Promoting Th17 Cell Polarization in Murine Noneosinophilic Asthma Model.

Background: T helper 17 (Th17) is regarded as key immune cell in the pathogenesis of noneosinophilic asthma (NEA) due to the recruitment of neutrophils into the airways. The mammalian target of rapamycin (mTOR) is an important signaling molecule that plays a critical role in immune regulation. This study focused on mTOR signaling pathway in the regulation of Th17-mediated neutrophilic airway inflammation. Methods: Ovalbumin (OVA) T cell receptor transgenic DO11.10 mice (DO11.10 mice) were used to establish NEA model, and few mice received specific mTORC1 inhibitor rapamycin (RAPA) before intranasal administration of OVA. The severity of airway inflammation was determined by differential cell counts in bronchoalveolar lavage (BAL) fluids and histopathologic lung analysis. The levels of various cytokines in BAL fluids and lung tissues were measured. To determine the role of mTORC1 signaling in Th17 differentiation, naive T cells from wild-type (WT) and TSC1 knockout (KO) mice were cultured in Th17 skewing condition with or without RAPA in vitro and the production of IL-17A was compared. Results: Treatment with RAPA markedly attenuated OVA-induced neutrophilic airway inflammation in DO11.10 mice. Also the production of IL-17A was inhibited without affecting the production of interferon-γ (IFN-γ) and IL-4 in lungs. Furthermore, RAPA suppressed differentiation of Th17 cells in vitro, whereas enhanced activity of mTORC1 promoted Th17 cell differentiation and increased the expression of Th17-related transcription factors RORγt and RORα. Conclusion: These results suggested that mTOR promoted Th17 cell polarization and enhanced OVA-induced neutrophilic airway inflammation in experimental NEA.

Aqueous Pyrostegia venusta (Ker Gawl.) Miers extract attenuates allergen-induced asthma in a mouse model via an antioxidant mechanism

Objective: Pyrostegia venusta (Ker-Gawl.) Miers (Bignoniaceae) is a perennial invasive vine, distributed worldwide. In folk medicine, its parts are used for the treatment of inflammatory respiratory diseases. Extracts of P. venusta have antioxidant, antimicrobial, and antinociceptive properties. The aim of this study was to evaluate the effects of two extracts (aqueous and hydroethanolic) of P. venusta in the treatment of asthma in an animal model.Methods: Balb/c mice were sensitized twice with ovalbumin (OVA) intraperitoneally (ip), one week apart, and after one week, challenged with OVA intranasally on four alternate days. Mice were treated ip with 300 mg/kg of aqueous or hydroethanolic extracts for seven consecutive days. Control groups received saline on the same days. Bronchial hyperresponsiveness, production of Th1 and Th2 cytokines, lung and airway inflammation, and antioxidant activity in lung tissue were assessed.Results: Treatment with aqueous extract significantly decreased bronchial hyperresponsiveness, measured by total and tissue resistance and elastance. The administration of hydroethanolic extract did not reduce bronchial hyperresponsiveness. In addition, both extracts significantly reduced total cell and eosinophil counts in bronchoalveolar lavage. Both extracts did not change significantly IL-4, IL-5, IL-9, IL-13, IFN-gamma, and TGF-beta levels. Of note, only the aqueous extract significantly increased the total antioxidant activity and reduced lung inflammation.Conclusion: Aqueous extract of P. venusta reduced bronchial hyperresponsiveness, lung and airway inflammation, probably via an antioxidant mechanism. These results demonstrate that P. venusta may have potential for asthma treatment.