Publications by authors named "Rafeul Alam"

99 Publications

Management Strategies to Reduce Exacerbations in non-T2 Asthma.

J Allergy Clin Immunol Pract 2021 07;9(7):2588-2597

Center for Lung Biology, Department of Medicine, University of Washington, Seattle, Wash; Division of Allergy and Immunology, University of Washington, Seattle, Wash.

There have been considerable advances in our understanding of asthmatic airway inflammation, resulting in a paradigm shift of classifying individuals on the basis of either the presence or the absence of type 2 (T2) inflammatory markers. Several novel monoclonal antibody therapies targeting T2 cytokines have demonstrated significant clinical effects including reductions in acute exacerbations and improvements in asthma-related quality of life and lung function for individuals with T2-high asthma. However, there have been fewer advancements in developing therapies for those without evidence of T2 airway inflammation (so-called non-T2 asthma). Here, we review the heterogeneity of molecular mechanisms responsible for initiation and regulation of non-T2 inflammation and discuss both current and potential future therapeutic options for individuals with non-T2 asthma.
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http://dx.doi.org/10.1016/j.jaip.2021.04.033DOI Listing
July 2021

The molecular and epigenetic mechanisms of innate lymphoid cell (ILC) memory and its relevance for asthma.

J Exp Med 2021 Jul 2;218(7). Epub 2021 Jun 2.

Division of Allergy & Immunology, Department of Medicine, National Jewish Health, Denver, CO.

Repetitive exposure of Rag1-/- mice to the Alternaria allergen extract generated a form of memory that elicited an asthma-like response upon a subthreshold recall challenge 3-15 wk later. This memory was associated with lung ICOS+ST2+ ILC2s. Genetic, pharmacologic, and antibody-mediated inhibition and adoptive transfer established an essential role for ILC2s in memory-driven asthma. ATAC-seq demonstrated a distinct epigenetic landscape of memory ILC2s and identified Bach2 and AP1 (JunD and Fosl2) motifs as major drivers of altered gene accessibility. scRNA-seq, gene knockout, and signaling studies suggest that repetitive allergenic stress induces a gene repression program involving Nr4a2, Zeb1, Bach2, and JunD and a preparedness program involving Fhl2, FosB, Stat6, Srebf2, and MPP7 in memory ILC2s. A mutually regulated balance between these two programs establishes and maintains memory. The preparedness program (e.g., Fhl2) can be activated with a subthreshold cognate stimulation, which down-regulates repressors and activates effector pathways to elicit the memory-driven phenotype.
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http://dx.doi.org/10.1084/jem.20201354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8176441PMC
July 2021

Sprouty2 positively regulates T cell function and airway inflammation through regulation of CSK and LCK kinases.

PLoS Biol 2021 03 8;19(3):e3001063. Epub 2021 Mar 8.

Division of Allergy and Immunology, Department of Medicine, National Jewish Health, Denver, Colorado, United States of America.

The function of Sprouty2 (Spry2) in T cells is unknown. Using 2 different (inducible and T cell-targeted) knockout mouse strains, we found that Spry2 positively regulated extracellular signal-regulated kinase 1/2 (ERK1/2) signaling by modulating the activity of LCK. Spry2-/- CD4+ T cells were unable to activate LCK, proliferate, differentiate into T helper cells, or produce cytokines. Spry2 deficiency abrogated type 2 inflammation and airway hyperreactivity in a murine model of asthma. Spry2 expression was higher in blood and airway CD4+ T cells from patients with asthma, and Spry2 knockdown impaired human T cell proliferation and cytokine production. Spry2 deficiency up-regulated the lipid raft protein caveolin-1, enhanced its interaction with CSK, and increased CSK interaction with LCK, culminating in augmented inhibitory phosphorylation of LCK. Knockdown of CSK or dislodgment of caveolin-1-bound CSK restored ERK1/2 activation in Spry2-/- T cells, suggesting an essential role for Spry2 in LCK activation and T cell function.
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http://dx.doi.org/10.1371/journal.pbio.3001063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7971865PMC
March 2021

IL-33/ST2 signaling modulates Afghanistan particulate matter induced airway hyperresponsiveness in mice.

Toxicol Appl Pharmacol 2020 10 8;404:115186. Epub 2020 Aug 8.

Department of Medicine, Medicine Office of Research, National Jewish Health, Denver, CO, United States of America. Electronic address:

Increased symptoms of asthma-like respiratory illnesses have been reported in soldiers returning from tours of duty in Afghanistan. Inhalation of desert particulate matter (PM) may contribute to this deployment-related lung disease (DRLD), but little is known about disease mechanisms. The IL-33 signaling pathway, including its receptor ST2, has been implicated in the pathogenesis of lung diseases including asthma, but its role in PM-mediated airway dysfunction has not been studied. The goal of this study was to investigate whether IL-33/ST2 signaling contributes to airway dysfunction in preclinical models of lung exposure to Afghanistan PM (APM). Wild-type (WT) and ST2 knockout (KO) mice on the BALB/C background were oropharyngeally instilled with a single dose of saline or 50 μg of APM in saline. Airway hyperresponsiveness (AHR) and inflammation were assessed after 24 h. In WT mice, a single APM exposure induced AHR and neutrophilic inflammation. Unlike the WT mice, ST2 KO mice that lack the receptor for IL-33 did not demonstrate AHR although airway neutrophilic inflammation was comparable to the WT mice. Oropharyngeal delivery of a soluble ST2 decoy receptor in APM-exposed WT mice significantly blocked AHR. Additional data in mouse tracheal epithelial cell and lung macrophage cultures demonstrated a role of APM-induced IL-33/ST2 signaling in suppression of regulator of G protein signaling 2 (RGS2), a gene known to protect against bronchoconstriction. We present for the first time that APM may increase AHR, one of the features of asthma, in part through the IL-33/ST2/RGS2 pathway.
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http://dx.doi.org/10.1016/j.taap.2020.115186DOI Listing
October 2020

Maternal diesel particle exposure promotes offspring asthma through NK cell-derived granzyme B.

J Clin Invest 2020 08;130(8):4133-4151

Division of Allergy and Clinical Immunology, Department of Medicine, National Jewish Health (NJH), Denver, Colorado, USA.

Mothers living near high-traffic roads before or during pregnancy are more likely to have children with asthma. Mechanisms are unknown. Using a mouse model, here we showed that maternal exposure to diesel exhaust particles (DEP) predisposed offspring to allergic airway disease (AAD, murine counterpart of human asthma) through programming of their NK cells; predisposition to AAD did not develop in DEP pups that lacked NK cells and was induced in normal pups receiving NK cells from WT DEP pups. DEP NK cells expressed GATA3 and cosecreted IL-13 and the killer protease granzyme B in response to allergen challenge. Extracellular granzyme B did not kill, but instead stimulated protease-activated receptor 2 (PAR2) to cooperate with IL-13 in the induction of IL-25 in airway epithelial cells. Through loss-of-function and reconstitution experiments in pups, we showed that NK cells and granzyme B were required for IL-25 induction and activation of the type 2 immune response and that IL-25 mediated NK cell effects on type 2 response and AAD. Finally, experiments using human cord blood and airway epithelial cells suggested that DEP might induce an identical pathway in humans. Collectively, we describe an NK cell-dependent endotype of AAD that emerged in early life as a result of maternal exposure to DEP.
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http://dx.doi.org/10.1172/JCI130324DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7410053PMC
August 2020

Volumetric assessment of paranasal sinus opacification on computed tomography can be automated using a convolutional neural network.

Int Forum Allergy Rhinol 2020 11 15;10(11):1218-1225. Epub 2020 Jun 15.

Department of Radiology, National Jewish Health, Denver, CO.

Background: Computed tomography (CT) plays a key role in evaluation of paranasal sinus inflammation, but improved, and standardized, objective assessment is needed. Computerized volumetric analysis has benefits over visual scoring, but typically relies on manual image segmentation, which is difficult and time-consuming, limiting practical applicability. We hypothesized that a convolutional neural network (CNN) algorithm could perform automatic, volumetric segmentation of the paranasal sinuses on CT, enabling efficient, objective measurement of sinus opacification. In this study we performed initial clinical testing of a CNN for fully automatic quantitation of paranasal sinus opacification in the diagnostic workup of patients with chronic upper and lower airway disease.

Methods: Sinus CT scans were collected on 690 patients who underwent imaging as part of multidisciplinary clinical workup at a tertiary care respiratory hospital between April 2016 and November 2017. A CNN was trained to perform automatic segmentation using a subset of CTs (n = 180) that were segmented manually. A nonoverlapping set (n = 510) was used for testing. CNN opacification scores were compared with Lund-MacKay (LM) visual scores, pulmonary function test results, and other clinical variables using Spearman correlation and linear regression.

Results: CNN scores were correlated with LM scores (rho = 0.82, p < 0.001) and with forced expiratory volume in 1 second (FEV ) percent predicted (rho = -0.21, p < 0.001), FEV /forced vital capacity ratio (rho = -0.27, p < 0.001), immunoglobulin E (rho = 0.20, p < 0.001), eosinophil count (rho = 0.28, p < 0.001), and exhaled nitric oxide (rho = 0.40, p < 0.001).

Conclusion: Segmentation of the paranasal sinuses on CT can be automated using a CNN, providing truly objective, volumetric quantitation of sinonasal inflammation.
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http://dx.doi.org/10.1002/alr.22588DOI Listing
November 2020

Optimal identification of human conventional and nonconventional (CRTH2IL7Rα) ILC2s using additional surface markers.

J Allergy Clin Immunol 2020 08 4;146(2):390-405. Epub 2020 Feb 4.

Division of Allergy and Immunology, Department of Medicine, Denver, Colo; School of Medicine, University of Colorado Denver, Denver, Colo. Electronic address:

Background: Human type 2 innate lymphoid cells (ILC2s) are identified by coupled detection of CRTH2 and IL7Rα on lineage negative (Lin) cells. Type 2 cytokine production by CRTH2IL7Rα innate lymphoid cells (ILCs) is unknown.

Objective: We sought to identify CRTH2IL7Rα type 2 cytokine-producing ILCs and their disease relevance.

Methods: We studied human blood and lung ILCs from asthmatic and control subjects by flow cytometry, ELISA, RNA sequencing, quantitative PCR, adoptive transfer to mice, and measurement of airway hyperreactivity by Flexivent.

Results: We found that IL-5 and IL-13 were expressed not only by CRTH2 but also by CRTH2IL7Rα and CRTH2IL7Rα (double-negative [DN]) human blood and lung cells. All 3 ILC populations expressed type 2 genes and induced airway hyperreactivity when adoptively transferred to mice. The frequency of type 2 cytokine-positive IL7Rα and DN ILCs were similar to that of CRTH2 ILCs in the blood and lung. Their frequency was higher in asthmatic patients than in disease controls. Transcriptomic analysis of CRTH2, IL7Rα, and DN ILCs confirmed the expression of mRNA for type 2 transcription factors in all 3 populations. Unexpectedly, the mRNA for GATA3 and IL-5 correlated better with mRNA for CD30, TNFR2, ICOS, CCR4, and CD200R1 than for CRTH2. By using a combination of these surface markers, especially CD30/TNFR2, we identified a previously unrecognized ILC2 population.

Conclusions: The commonly used surface markers for human ILC2s leave a majority of type 2 cytokine-producing ILC2s unaccounted for. We identified top GATA3-correlated cell surface-expressed genes in human ILCs by RNA sequencing. These new surface markers, such as CD30 and TNFR2, identified a previously unrecognized human ILC2 population. This ILC2 population is likely to contribute to asthma.
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http://dx.doi.org/10.1016/j.jaci.2020.01.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7398840PMC
August 2020

Interleukin 1 Receptor-Like 1 (IL1RL1) Promotes Airway Bacterial and Viral Infection and Inflammation.

Infect Immun 2019 07 20;87(7). Epub 2019 Jun 20.

National Jewish Health, Denver, Colorado, USA

Interleukin 1 receptor-like 1 (IL1RL1), also known as suppression of tumorigenicity 2 (ST2), is the receptor for interleukin 33 (IL-33) and has been increasingly studied in type 2 inflammation. An increase in airway IL-33/ST2 signaling in asthma has been associated with eosinophilic inflammation, but little is known about the role of ST2 in neutrophilic inflammation. Airway and human rhinovirus (HRV) infections are linked to neutrophilic inflammation during acute exacerbations of asthma. However, whether ST2 contributes to - and HRV-mediated airway inflammation is poorly understood. The current study sought to determine the functions of ST2 during airway or HRV infection. In cultured normal human primary airway epithelial cells, ST2 overexpression (OE) increased the production of neutrophilic chemoattractant IL-8 in the absence or presence of or HRV1B infection. ST2 OE also enhanced HRV1B-induced IP-10, a chemokine involved in asthma exacerbations. In the -infected mouse model, ST2 deficiency, in contrast to sufficiency, significantly reduced the levels of neutrophils following acute (≤24 h) infection, while in the HRV1B-infected mouse model, ST2 deficiency significantly reduced the levels of proinflammatory cytokines KC, IP-10, and IL-33 in bronchoalveolar lavage (BAL) fluid. Overall, ST2 overexpression in human epithelial cells and ST2 sufficiency in mice increased the and HRV loads in cell supernatants and BAL fluid. After pathogen infection, ST2-deficient mice showed a higher level of the host defense protein lactotransferrin in BAL fluid. Our data suggest that ST2 promotes proinflammatory responses (e.g., neutrophils) to airway bacterial and viral infection and that blocking ST2 signaling may broadly attenuate airway infection and inflammation.
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http://dx.doi.org/10.1128/IAI.00340-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6589056PMC
July 2019

Toll-Interacting Protein, Tollip, Inhibits IL-13-Mediated Pulmonary Eosinophilic Inflammation in Mice.

J Innate Immun 2018 27;10(2):106-118. Epub 2018 Jan 27.

Toll-interacting protein (Tollip) is a key negative regulator of innate immunity by preventing excessive proinflammatory responses. Tollip genetic variation has been associated with airflow limitation in asthma subjects and Tollip expression. Whether Tollip regulates lung inflammation in a type 2 cytokine milieu (e.g., IL-13) is unclear. Our goal was to determine the in vivo role of Tollip in IL-13-mediated lung eosinophilic inflammation and the underlying mechanisms. Tollip-knockout (KO) and wild-type (WT) mice were inoculated intranasally with recombinant mouse IL-13 protein to examine lung inflammation. To determine how Tollip regulates inflammation, alveolar macrophages and bone marrow-derived macrophages from Tollip KO and WT mice were cultured with or without IL-13 and/or IL-33. IL-13-treated Tollip KO mice significantly increased lung eosinophilic inflammation and eotaxin-2 (CCL24) levels compared with the WT mice. IL-13- treated Tollip KO (vs. WT) macrophages, in the absence and particularly in the presence of IL-33, increased expression of the IL-33 receptor ST2L and CCL24, which was in part dependent on enhanced activation of interleukin (IL)-1 receptor-associated kinase 1 (IRAK1) and signal transducer and activator of transcription 6 (STAT6). Our results suggest that Tollip downregulates IL-13-mediated pulmonary eosinophilia in part through inhibiting the activity of the ST2L/IL-33/IRAK1 axis and STAT6.
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http://dx.doi.org/10.1159/000485850DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5860932PMC
September 2019

Experimental asthma persists in IL-33 receptor knockout mice because of the emergence of thymic stromal lymphopoietin-driven IL-9 and IL-13 type 2 innate lymphoid cell subpopulations.

J Allergy Clin Immunol 2018 09 10;142(3):793-803.e8. Epub 2017 Nov 10.

Department of Medicine, Division of Allergy & Immunology, National Jewish Health, Denver, Colo; School of Medicine, University of Colorado Denver, Denver, Colo. Electronic address:

Background: IL-33 plays an important role in the development of experimental asthma.

Objective: We sought to study the role of the IL-33 receptor suppressor of tumorigenicity 2 (ST2) in the persistence of asthma in a mouse model.

Methods: We studied allergen-induced experimental asthma in ST2 knockout (KO) and wild-type control mice. We measured airway hyperresponsiveness by using flexiVent; inflammatory indices by using ELISA, histology, and real-time PCR; and type 2 innate lymphoid cells (ILC2s) in lung single-cell preparations by using flow cytometry.

Results: Airway hyperresponsiveness was increased in allergen-treated ST2 KO mice and comparable with that in allergen-treated wild-type control mice. Peribronchial and perivascular inflammation and mucus production were largely similar in both groups. Persistence of experimental asthma in ST2 KO mice was associated with an increase in levels of thymic stromal lymphopoietin (TSLP), IL-9, and IL-13, but not IL-5, in bronchoalveolar lavage fluid. Expectedly, ST2 deletion caused a reduction in IL-13 CD4 T cells, forkhead box P3-positive regulatory T cells, and IL-5 ILC2s. Unexpectedly, ST2 deletion led to an overall increase in innate lymphoid cells (CD45linCD25 cells) and IL-13 ILC2s, emergence of a TSLP receptor-positive IL-9 ILC2 population, and an increase in intraepithelial mast cell numbers in the lung. An anti-TSLP antibody abrogated airway hyperresponsiveness, inflammation, and mucus production in allergen-treated ST2 KO mice. It also caused a reduction in innate lymphoid cell, ILC2, and IL-9 and IL-13 ILC2 numbers in the lung.

Conclusions: Genetic deletion of the IL-33 receptor paradoxically increases TSLP production, which stimulates the emergence of IL-9 and IL-13 ILC2s and mast cells and leads to development of chronic experimental asthma. An anti-TSLP antibody abrogates all pathologic features of asthma in this model.
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http://dx.doi.org/10.1016/j.jaci.2017.10.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945345PMC
September 2018

Pre-pregnancy exposure to diesel exhaust predisposes offspring to asthma through IL-1β and IL-17A.

J Allergy Clin Immunol 2018 03 21;141(3):1118-1122.e3. Epub 2017 Sep 21.

Department of Medicine, Division of Allergy and Clinical Immunology, National Jewish Health, Denver, Colo; Department of Medicine, Division of Allergy and Clinical Immunology, University of Colorado Denver, Aurora, Colo. Electronic address:

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http://dx.doi.org/10.1016/j.jaci.2017.09.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5844783PMC
March 2018

The R213G polymorphism in SOD3 protects against allergic airway inflammation.

JCI Insight 2017 09 7;2(17). Epub 2017 Sep 7.

Department of Medicine, National Jewish Health, Denver, Colorado, USA.

Oxidative stress is important in the pathogenesis of allergic asthma. Extracellular superoxide dismutase (EC-SOD; SOD3) is the major antioxidant in lungs, but its role in allergic asthma is unknown. Here we report that asthmatics have increased SOD3 transcript levels in sputum and that a single nucleotide polymorphism (SNP) in SOD3 (R213G; rs1799895) changes lung distribution of EC-SOD, and decreases likelihood of asthma-related symptoms. Knockin mice analogous to the human R213G SNP had lower airway hyperresponsiveness, inflammation, and mucus hypersecretion with decreased interleukin-33 (IL-33) in bronchoalveolar lavage fluid and reduced type II innate lymphoid cells (ILC2s) in lungs. SOD mimetic (Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin) attenuated Alternaria-induced expression of IL-33 and IL-8 release in BEAS-2B cells. These results suggest that R213G SNP potentially benefits its carriers by resulting in high EC-SOD in airway-lining fluid, which ameliorates allergic airway inflammation by dampening the innate immune response, including IL-33/ST2-mediated changes in ILC2s.
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http://dx.doi.org/10.1172/jci.insight.95072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5621928PMC
September 2017

Steroid resistance of airway type 2 innate lymphoid cells from patients with severe asthma: The role of thymic stromal lymphopoietin.

J Allergy Clin Immunol 2018 01 20;141(1):257-268.e6. Epub 2017 Apr 20.

Department of Medicine, Division of Allergy & Immunology, and the Division of Pulmonary and Critical Care Medicine, National Jewish Health, Denver, Colo; Department of Medicine, University of Colorado Denver, School of Medicine, Denver, Colo. Electronic address:

Background: Type 2 innate lymphoid cells (ILC2s) represent an important type 2 immune cell. Glucocorticoid regulation of human ILC2s is largely unknown.

Objective: We sought to assess steroid resistance of human blood and airway ILC2s from asthmatic patients and to examine its mechanism of induction.

Methods: We studied human blood and lung ILC2s from asthmatic patients and control subjects using flow cytometry and ELISA.

Results: Dexamethasone inhibited (P = .04) chemoattractant receptor-homologous molecule expressed on T2 lymphocytes and type 2 cytokine expression by blood ILC2s stimulated with IL-25 and IL-33. However, it did not do so when ILC2s were stimulated with IL-7 and thymic stromal lymphopoietin (TSLP), 2 ligands of IL-7 receptor α. Unlike blood ILC2s, bronchoalveolar lavage (BAL) fluid ILC2s from asthmatic patients were resistant to dexamethasone. BAL fluid from asthmatic patients had increased TSLP but not IL-7 levels. BAL fluid TSLP levels correlated (r = 0.74) with steroid resistance of ILC2s. TSLP was synergistically induced in epithelial cells by IL-13 and human rhinovirus. Mechanistically, dexamethasone upregulated ILC2 expression of IL-7 receptor α, which augmented and sustained signal transducer and activator of transcription (STAT) 5 signaling by TSLP. TSLP induced mitogen-activated protein kinase kinase (MEK), c-Fos, inhibitor of DNA binding 3, phosphorylated signal transducer and activator of transcription (pSTAT) 3, and pSTAT5, molecules linked to steroid resistance. Dexamethasone inhibited c-Fos, inhibitor of DNA binding 3, and pSTAT3 but not pSTAT5 and MEK. The MEK inhibitor trametinib, the Janus kinase-STAT inhibitor tofacitinib, and the STAT5 inhibitor pimozide reversed steroid resistance of BAL ILC2s.

Conclusions: Dexamethasone inhibited type 2 cytokine production by blood ILC2s. IL-7 and TSLP abrogated this inhibition and induced steroid resistance of ILC2s in a MEK- and STAT5-dependent manner. BAL fluid ILC2s from asthmatic patients with increased TSLP levels were steroid resistant, which was reversed by clinically available inhibitors of MEK and STAT5.
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http://dx.doi.org/10.1016/j.jaci.2017.03.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5650571PMC
January 2018

Airway and serum biochemical correlates of refractory neutrophilic asthma.

J Allergy Clin Immunol 2017 Oct 3;140(4):1004-1014.e13. Epub 2017 Feb 3.

Department of Medicine, National Jewish Health, Denver, Colo.

Background: Despite progress in the diagnosis and management of asthma, many patients have poorly controlled or refractory asthma (RA). The mechanism of this RA is not well understood.

Objective: We sought to explore the relationship between neutrophils and other biomarkers of RA.

Method: Sixty patients with RA, 30 patients with nonrefractory asthma (NRA), and 20 healthy subjects were enrolled. We performed a comprehensive characterization of these study subjects, which included laboratory and pulmonary function studies, chest computed tomography, and bronchoscopy with bronchoalveolar lavage (BAL). We analyzed BAL fluid and serum for a total of 244 biomolecules using a multiplex assay and correlated them with clinical and other laboratory parameters.

Results: RA was significantly different from NRA with regard to pulmonary function indices, bronchial basement membrane thickness, and BAL fluid neutrophil and lymphocyte counts but not eosinophil counts. BAL fluid neutrophil counts negatively and positively correlated with forced vital capacity and age, respectively. Of the 244 biomolecules studied, 52 and 14 biomolecules from BAL fluid and serum, respectively, were significantly different among the study groups. Thirteen of these 52 molecules correlated with BAL fluid neutrophil counts. BAL fluid from 40% of patients with RA was positive for a pathogenic microbe. Infection-negative neutrophilic RA was associated with an increase in levels of select biomarkers of inflammation in the serum, suggesting the presence of systemic inflammation.

Conclusions: RA was associated with increased numbers of neutrophils and proneutrophilic biomolecules in the airways. Subclinical infection was present in 40% of patients with RA, which likely contributed to neutrophilic inflammation. A subgroup of patients with noninfected neutrophilic RA was associated with systemic inflammation.
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http://dx.doi.org/10.1016/j.jaci.2016.12.963DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5540819PMC
October 2017

Mechanism of T2/T17-predominant and neutrophilic T2/T17-low subtypes of asthma.

J Allergy Clin Immunol 2017 May 1;139(5):1548-1558.e4. Epub 2016 Oct 1.

Department of Medicine, Division of Allergy & Immunology, and Division of Pulmonary and Critical Care Medicine, National Jewish Health, Denver, Colo; School of Medicine, University of Colorado Denver, Denver, Colo. Electronic address:

Background: The mechanism of T2/T17-predominant and T2/T17-low asthma is unknown.

Objective: We sought to study the immune mechanism of T2/T17-predominant and T2/T17-low asthma.

Methods: In a previously reported cohort of 60 asthmatic patients, 16 patients were immunophenotyped with T2/T17-predominant asthma and 22 patients with T2/T17-low asthma. We examined bronchoalveolar lavage (BAL) fluid leukocytes, cytokines, mediators, and epithelial cell function for these asthma subgroups.

Results: Patients with T2/T17-predominant asthma had increased IL-1β, IL-6, IL-23, C3a, and serum amyloid A levels in BAL fluid, and these correlated with IL-1β and C3a levels. T2/T17 cells expressed higher levels of the IL-1 receptor and phospho-p38 mitogen-activated protein kinase. Anakinra, an IL-1 receptor antagonist protein, inhibited BAL T2/T17 cell counts. T2/T17-low asthma had 2 distinct subgroups: neutrophilic asthma (45%) and pauci-inflammatory asthma (55%). This contrasted with patients with T2/T17-predominant and T2-predominant asthma, which included neutrophilic asthma in 6% and 0% of patients, respectively. BAL fluid neutrophils strongly correlated with BAL fluid myeloperoxidase, IL-8, IL-1α, IL-6, granulocyte colony-stimulating factor, and GM-CSF levels. Sixty percent of the patients with neutrophilic asthma had a pathogenic microorganism in BAL culture, which suggested a subclinical infection.

Conclusion: We uncovered a critical role for the IL-1β pathway in patients with T2/T17-predminant asthma. A subgroup of patients with T2/T17-low asthma had neutrophilic asthma and increased BAL fluid IL-1α, IL-6, IL-8, granulocyte colony-stimulating factor, and GM-CSF levels. IL-1α was directly involved in IL-8 production and likely contributed to neutrophilic asthma. Sixty percent of neutrophilic patients had a subclinical infection.
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http://dx.doi.org/10.1016/j.jaci.2016.08.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5376378PMC
May 2017

Nonsteroidal anti-inflammatory-induced inhibition of signal transducer and activator of transcription 6 (STAT-6) phosphorylation in aspirin-exacerbated respiratory disease.

J Allergy Clin Immunol 2016 08 23;138(2):579-85. Epub 2016 Feb 23.

National Jewish Health, Denver, Colo.

Background: Aspirin desensitization provides long-term clinical benefits. The exact mechanisms of aspirin desensitization are not clearly understood.

Objective: We sought to evaluate the effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on T-cell activation of the IL-4 pathway in aspirin-sensitive patients with asthma and control subjects.

Methods: A total of 11 aspirin-sensitive patients with asthma, 10 aspirin-tolerant patients with asthma, and 10 controls without asthma were studied. PBMCs were stimulated with an anti-CD3 antibody and IL-4 or IL-12, with and without the presence of NSAIDs. The expression of phosphorylated signal transducers and activators of transcription 6 (pSTAT6), phosphorylated signal transducers and activators of transcription 4, and IL-4 was detected in CD4 T cells by flow cytometry.

Results: Stimulation with a combination of anti-CD3 and IL-4 induced pSTAT6 in CD4 T cells from all subjects. The induction of pSTAT6 was significantly higher in aspirin-sensitive patients with asthma than in controls subjects. The increase in pSTAT6 was inhibited in a dose-dependent manner by aspirin and indomethacin and minimally by sodium salicylate. This inhibition was strongest in aspirin-sensitive patients. Two-group comparisons showed significant differences in pSTAT6 inhibition by all concentrations of indomethacin and aspirin: between aspirin-sensitive and aspirin-tolerant groups and between aspirin-sensitive and control groups. No differences were found between aspirin-tolerant and control groups at all 3 concentrations. The inhibition of pSTAT6 was associated with reduced IL-4 expression.

Conclusions: NSAIDs inhibited signal transducers and activators of transcription 6 signaling in CD4 T cells. This inhibition was significantly higher in aspirin-sensitive patients than in aspirin-tolerant subjects and was associated with reduced expression of IL-4. These findings have implications for clinical benefits of aspirin desensitization in aspirin-sensitive patients with asthma.
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http://dx.doi.org/10.1016/j.jaci.2015.11.038DOI Listing
August 2016

Association of B-cell activating factor receptor deficiency with the P21R polymorphism and common variable immunodeficiency.

Ann Allergy Asthma Immunol 2015 Jul 23;115(1):82-3. Epub 2015 May 23.

Division of Allergy & Immunology, National Jewish Health, Department of Medicine, Denver, Colorado; University of Colorado Denver, School of Medicine, Denver, Colorado. Electronic address:

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http://dx.doi.org/10.1016/j.anai.2015.04.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4760690PMC
July 2015

The other side of asthma: Steroid-refractory disease in the absence of TH2-mediated inflammation.

J Allergy Clin Immunol 2015 May 13;135(5):1196-8. Epub 2015 Mar 13.

Department of Medicine, National Jewish Health, Denver, Colo.

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http://dx.doi.org/10.1016/j.jaci.2015.01.032DOI Listing
May 2015

Persistence of asthma requires multiple feedback circuits involving type 2 innate lymphoid cells and IL-33.

J Allergy Clin Immunol 2015 Jul 21;136(1):59-68.e14. Epub 2015 Jan 21.

Division of Allergy and Immunology, National Jewish Health, Denver, Colo; University of Colorado Denver, Denver, Colo. Electronic address:

Background: Asthma in a mouse model spontaneously resolves after cessation of allergen exposure. We developed a mouse model in which asthma features persisted for 6 months after cessation of allergen exposure.

Objective: We sought to elucidate factors contributing to the persistence of asthma.

Methods: We used a combination of immunologic, genetic, microarray, and pharmacologic approaches to dissect the mechanism of asthma persistence.

Results: Elimination of T cells though antibody-mediated depletion or lethal irradiation and transplantation of recombination-activating gene (Rag1)(-/-) bone marrow in mice with chronic asthma resulted in resolution of airway inflammation but not airway hyperreactivity or remodeling. Elimination of T cells and type 2 innate lymphoid cells (ILC2s) through lethal irradiation and transplantation of Rag2(-/-)γc(-/-) bone marrow or blockade of IL-33 resulted in resolution of airway inflammation and hyperreactivity. Persistence of asthma required multiple interconnected feedback and feed-forward circuits between ILC2s and epithelial cells. Epithelial IL-33 induced ILC2s, a rich source of IL-13. The latter directly induced epithelial IL-33, establishing a positive feedback circuit. IL-33 autoinduced, generating another feedback circuit. IL-13 upregulated IL-33 receptors and facilitated IL-33 autoinduction, thus establishing a feed-forward circuit. Elimination of any component of these circuits resulted in resolution of chronic asthma. In agreement with the foregoing, IL-33 and ILC2 levels were increased in the airways of asthmatic patients. IL-33 levels correlated with disease severity.

Conclusions: We present a critical network of feedback and feed-forward interactions between epithelial cells and ILC2s involved in maintaining chronic asthma. Although T cells contributed to the severity of chronic asthma, they were redundant in maintaining airway hyperreactivity and remodeling.
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http://dx.doi.org/10.1016/j.jaci.2014.11.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4494983PMC
July 2015

Reply: To PMID 25042748.

Authors:
Rafeul Alam

J Allergy Clin Immunol 2015 Jan 12;135(1):291. Epub 2014 Nov 12.

Division of Allergy & Immunology, Department of Medicine, National Jewish Health & University of Colorado Denver, Denver, Colo. Electronic address:

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http://dx.doi.org/10.1016/j.jaci.2014.09.038DOI Listing
January 2015

Obesity and asthma--is there a causal association?

Authors:
Rafeul Alam

Immunol Allergy Clin North Am 2014 Nov;34(4):xi-xii

Division of Allergy and Immunology, National Jewish Health, University of Colorado Denver School of Medicine, 1400 Jackson Street, Denver, CO 80206, USA. Electronic address:

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http://dx.doi.org/10.1016/j.iac.2014.09.001DOI Listing
November 2014

Association between specific timothy grass antigens and changes in TH1- and TH2-cell responses following specific immunotherapy.

J Allergy Clin Immunol 2014 Nov 16;134(5):1076-83. Epub 2014 Jul 16.

Division of Vaccine Discovery, La Jolla Institute for Allergy & Immunology, La Jolla, Calif. Electronic address:

Background: Different populations of T cells are involved in the pathogenesis of allergic diseases.

Objective: We investigated changes in TH-cell populations in patients with allergies after specific immunotherapy (SIT).

Methods: PBMCs were isolated from patients with allergies who received SIT and those who did not (controls). We tested the ability of peptides from 93 timothy grass (TG) proteins to induce T-cell responses (cytokine production). We used ELISPOT and staining assays for intracellular cytokines to measure the production of IL-4, IL-5, IL-13, IFN-γ, and IL-10.

Results: Compared with PBMCs from controls, PBMCs from patients who received SIT produced lower levels of TH2 cytokines on incubation with several different TG peptides. These data were used to select 20 peptides to be tested in an independent cohort of 20 patients with allergies who received SIT and 20 controls. We again observed a significant decrease in the production of TH2 cytokines, and an increase in the production of the TH1 cytokine IFN-γ, in PBMCs from the validation groups. These changes correlated with improved symptoms after SIT. Immunization with this selected pool of peptides (or their associated antigens) could protect a substantial proportion of the population from TG allergy.

Conclusions: We observed a significant decrease in the production of TH2 cytokines by PBMCs from patients who received SIT for TG allergy compared to those who did not. These changes might be used to monitor response to therapy. The decrease occurred in response to antigens that elicit little (if any) IgE responses; these antigens might be developed for use in immunotherapy.
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http://dx.doi.org/10.1016/j.jaci.2014.05.033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4253970PMC
November 2014

Increased frequency of dual-positive TH2/TH17 cells in bronchoalveolar lavage fluid characterizes a population of patients with severe asthma.

J Allergy Clin Immunol 2014 Nov 18;134(5):1175-1186.e7. Epub 2014 Jul 18.

Department of Medicine, Division of Allergy & Immunology, and Division of Pulmonary and Critical Care Medicine, National Jewish Health, Denver, Colo; School of Medicine, University of Colorado at Denver, Denver, Colo. Electronic address:

Background: TH2 cells can further differentiate into dual-positive TH2/TH17 cells. The presence of dual-positive TH2/TH17 cells in the airways and their effect on asthma severity are unknown.

Objective: We sought to study dual-positive TH2/TH17 cells in bronchoalveolar lavage (BAL) fluid from asthmatic patients, examine their response to glucocorticoids, and define their relevance for disease severity.

Methods: Bronchoscopy and lavage were performed in 52 asthmatic patients and 25 disease control subjects. TH2 and TH2/TH17 cells were analyzed by using multicolor flow cytometry and confocal immunofluorescence microscopy. Cytokines were assayed by means of ELISA.

Results: Dual-positive TH2/TH17 cells were present at a higher frequency in BAL fluid from asthmatic patients compared with numbers seen in disease control subjects. High-level IL-4 production was typically accompanied by high-level IL-17 production and coexpression of GATA3 and retinoic acid receptor-related orphan receptor γt. Increased presence of TH2/TH17 cells was associated with increased IL-17 production in lavage fluid. TH2/TH17 cell counts and IL-17 production correlated with PC20 for methacholine, eosinophil counts, and FEV1. TH2/TH17 cells, unlike TH2 cells, were resistant to dexamethasone-induced cell death. They expressed higher levels of mitogen-activated protein-extracellular signal-regulated kinase kinase 1, a molecule that induces glucocorticoid resistance. On the basis of the dominance of BAL fluid TH2 or TH2/TH17 cells, we identified 3 subgroups of asthma: TH2(predominant), TH2/TH17(predominant), and TH2/TH17(low). The TH2/TH17(predominant) subgroup manifested the most severe form of asthma, whereas the TH2/TH17(low) subgroup had the mildest asthma.

Conclusion: Asthma is associated with a higher frequency of dual-positive TH2/TH17 cells in BAL fluid. The TH2/TH17(predominant) subgroup of asthmatic patients manifested glucocorticoid resistance in vitro. They also had the greatest airway obstruction and hyperreactivity compared with the TH2(predominant) and TH2/TH17(low) subgroups.
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http://dx.doi.org/10.1016/j.jaci.2014.05.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254017PMC
November 2014

The complexity of drug hypersensitivity. Foreword.

Authors:
Rafeul Alam

Immunol Allergy Clin North Am 2014 Aug 13;34(3):xiii-xiv. Epub 2014 May 13.

Division of Allergy and Immunology, National Jewish Health andUniversity of Colorado Denver, 1400 Jackson Street, Denver, CO 80206, USA. Electronic address:

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http://dx.doi.org/10.1016/j.iac.2014.04.014DOI Listing
August 2014

The amazing mast cell.

Authors:
Rafeul Alam

Immunol Allergy Clin North Am 2014 May 16;34(2):xv-xvi. Epub 2014 Feb 16.

Division of Allergy and Immunology, National Jewish Health, University of Colorado Denver, 1400 Jackson Street, Denver, CO 80206, USA. Electronic address:

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http://dx.doi.org/10.1016/j.iac.2014.02.002DOI Listing
May 2014

A mouse model links asthma susceptibility to prenatal exposure to diesel exhaust.

J Allergy Clin Immunol 2014 Jul 22;134(1):63-72. Epub 2013 Dec 22.

Department of Medicine, Division of Allergy and Clinical Immunology, National Jewish Health, Denver, Colo; Department of Medicine, Division of Allergy and Clinical Immunology, University of Colorado Denver, Aurora, Colo. Electronic address:

Background: Most asthma begins in the first years of life. This early onset cannot be attributed merely to genetic factors because the prevalence of asthma is increasing. Epidemiologic studies have indicated roles for prenatal and early childhood exposures, including exposure to diesel exhaust. However, little is known about the mechanisms. This is largely due to a paucity of animal models.

Objective: We aimed to develop a mouse model of asthma susceptibility through prenatal exposure to diesel exhaust.

Methods: Pregnant C57BL/6 female mice were given repeated intranasal applications of diesel exhaust particles (DEPs) or PBS. Offspring underwent suboptimal immunization and challenge with ovalbumin (OVA) or received PBS. Pups were examined for features of asthma; lung and liver tissues were analyzed for transcription of DEP-regulated genes.

Results: Offspring of mice exposed to DEPs were hypersensitive to OVA, as indicated by airway inflammation and hyperresponsiveness, increased serum OVA-specific IgE levels, and increased pulmonary and systemic TH2 and TH17 cytokine levels. These cytokines were primarily produced by natural killer (NK) cells. Antibody-mediated depletion of NK cells prevented airway inflammation. Asthma susceptibility was associated with increased transcription of genes known to be specifically regulated by the aryl hydrocarbon receptor and oxidative stress. Features of asthma were either marginal or absent in OVA-treated pups of PBS-exposed mice.

Conclusion: We created a mouse model that linked maternal exposure to DEPs with asthma susceptibility in offspring. Development of asthma was dependent on NK cells and associated with increased transcription from aryl hydrocarbon receptor- and oxidative stress-regulated genes.
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http://dx.doi.org/10.1016/j.jaci.2013.10.047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4065237PMC
July 2014

Rituximab as successful adjunct treatment in a patient with disseminated nontuberculous mycobacterial infection due to acquired anti-interferon-γ autoantibody.

Clin Infect Dis 2014 Mar 11;58(6):e115-8. Epub 2013 Dec 11.

Division of Mycobacterial and Respiratory Infections.

An acquired immune deficiency due to interferon gamma (IFN-γ) autoantibodies was diagnosed in a 78-year-old Japanese man with treatment-refractory disseminated nontuberculous mycobacterial infection. In addition to standard antimycobacterial therapy, he was successfully treated with rituximab to eliminate B cells and thereby the autoantibody. Subsequently, he obtained a sustained remission from infection.
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http://dx.doi.org/10.1093/cid/cit809DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3935498PMC
March 2014

Urticaria: an evolving story.

Authors:
Rafeul Alam

Immunol Allergy Clin North Am 2014 Feb 25;34(1):xiii-xiv. Epub 2013 Oct 25.

Division of Allergy and Immunology, National Jewish Health, University of Colorado Denver School of Medicine, 1400 Jackson Street, Denver, CO 80206, USA. Electronic address:

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http://dx.doi.org/10.1016/j.iac.2013.10.002DOI Listing
February 2014

Angioedema: what we know and what we need to know.

Authors:
Rafeul Alam

Immunol Allergy Clin North Am 2013 Nov 18;33(4):ix-x. Epub 2013 Sep 18.

Division of Allergy and Immunology, National Jewish Health and University of Colorado Denver, 1400 Jackson Street, Denver, CO 80206, USA. Electronic address:

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http://dx.doi.org/10.1016/j.iac.2013.09.003DOI Listing
November 2013
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