Publications by authors named "Anne I Sperling"

76 Publications

IL-33-mediated Eosinophilia Protects Against Acute Lung Injury.

Am J Respir Cell Mol Biol 2021 Feb 11. Epub 2021 Feb 11.

Kaiser Permanente Hawaii, 50679, Hawaii Permanente Medical Group , Honolulu, Hawaii, United States;

Pneumonia-induced lung injury and acute respiratory distress syndrome can develop due to an inappropriate inflammatory response to acute infections, leading to a compromised alveolar barrier. Recent work suggests that hospitalized patients with allergies/asthma are less likely to die from pulmonary infections, and that there is a correlation between survival from acute respiratory distress syndrome and higher eosinophil counts; thus, we hypothesized that eosinophils associated with a type 2 immune response may protect against pneumonia-induced acute lung injury. To test this hypothesis, mice were treated with the type 2-initiating cytokine IL-33 intratracheally 3 days prior to induction of pneumonia with airway administration of a lethal dose of S. aureus. Interestingly, IL-33 pretreatment promoted survival by inhibiting acute lung injury: levels of bronchoalveolar lavage proinflammatory cytokines and pulmonary edema were both reduced, with an associated increase in oxygen saturation. Pulmonary neutrophilia was also reduced, while eosinophilia was strongly increased. This eosinophilia was key to protection; eosinophil reduction eliminated both IL-33-mediated protection against mortality as well as inhibition of neutrophilia and pulmonary edema. Together, these data reveal a novel role for eosinophils in protection against lung injury and suggest that modulation of pulmonary type 2 immunity may represent a novel therapeutic strategy.
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http://dx.doi.org/10.1165/rcmb.2020-0166OCDOI Listing
February 2021

Modeling human adaptive immune responses with tonsil organoids.

Nat Med 2021 01 11;27(1):125-135. Epub 2021 Jan 11.

Department of Microbiology and Immunology, Stanford University, Stanford, CA, USA.

Most of what we know about adaptive immunity has come from inbred mouse studies, using methods that are often difficult or impossible to confirm in humans. In addition, vaccine responses in mice are often poorly predictive of responses to those same vaccines in humans. Here we use human tonsils, readily available lymphoid organs, to develop a functional organotypic system that recapitulates key germinal center features in vitro, including the production of antigen-specific antibodies, somatic hypermutation and affinity maturation, plasmablast differentiation and class-switch recombination. We use this system to define the essential cellular components necessary to produce an influenza vaccine response. We also show that it can be used to evaluate humoral immune responses to two priming antigens, rabies vaccine and an adenovirus-based severe acute respiratory syndrome coronavirus 2 vaccine, and to assess the effects of different adjuvants. This system should prove useful for studying critical mechanisms underlying adaptive immunity in much greater depth than previously possible and to rapidly test vaccine candidates and adjuvants in an entirely human system.
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http://dx.doi.org/10.1038/s41591-020-01145-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7891554PMC
January 2021

Temperature Trajectory Subphenotypes Correlate With Immune Responses in Patients With Sepsis.

Crit Care Med 2020 Nov;48(11):1645-1653

Department of Medicine, University of Chicago Medical Center, Chicago, IL.

Objectives: We recently found that distinct body temperature trajectories of infected patients correlated with survival. Understanding the relationship between the temperature trajectories and the host immune response to infection could allow us to immunophenotype patients at the bedside using temperature. The objective was to identify whether temperature trajectories have consistent associations with specific cytokine responses in two distinct cohorts of infected patients.

Design: Prospective observational study.

Setting: Large academic medical center between 2013 and 2019.

Subjects: Two cohorts of infected patients: 1) patients in the ICU with septic shock and 2) hospitalized patients with Staphylococcus aureus bacteremia.

Interventions: Clinical data (including body temperature) and plasma cytokine concentrations were measured. Patients were classified into four temperature trajectory subphenotypes using their temperature measurements in the first 72 hours from the onset of infection. Log-transformed cytokine levels were standardized to the mean and compared with the subphenotypes in both cohorts.

Measurements And Main Results: The cohorts consisted of 120 patients with septic shock (cohort 1) and 88 patients with S. aureus bacteremia (cohort 2). Patients from both cohorts were classified into one of four previously validated temperature subphenotypes: "hyperthermic, slow resolvers" (n = 19 cohort 1; n = 13 cohort 2), "hyperthermic, fast resolvers" (n = 18 C1; n = 24 C2), "normothermic" (n = 54 C1; n = 31 C2), and "hypothermic" (n = 29 C1; n = 20 C2). Both "hyperthermic, slow resolvers" and "hyperthermic, fast resolvers" had high levels of G-CSF, CCL2, and interleukin-10 compared with the "hypothermic" group when controlling for cohort and timing of cytokine measurement (p < 0.05). In contrast to the "hyperthermic, slow resolvers," the "hyperthermic, fast resolvers" showed significant decreases in the levels of several cytokines over a 24-hour period, including interleukin-1RA, interleukin-6, interleukin-8, G-CSF, and M-CSF (p < 0.001).

Conclusions: Temperature trajectory subphenotypes are associated with consistent cytokine profiles in two distinct cohorts of infected patients. These subphenotypes could play a role in the bedside identification of cytokine profiles in patients with sepsis.
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http://dx.doi.org/10.1097/CCM.0000000000004610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7837282PMC
November 2020

Fibroblast-enriched endoplasmic reticulum protein TXNDC5 promotes pulmonary fibrosis by augmenting TGFβ signaling through TGFBR1 stabilization.

Nat Commun 2020 08 26;11(1):4254. Epub 2020 Aug 26.

Department and Graduate Institute of Pharmacology, National Taiwan University College of Medicine, Taipei, Taiwan.

Pulmonary fibrosis (PF) is a major public health problem with limited therapeutic options. There is a clear need to identify novel mediators of PF to develop effective therapeutics. Here we show that an ER protein disulfide isomerase, thioredoxin domain containing 5 (TXNDC5), is highly upregulated in the lung tissues from both patients with idiopathic pulmonary fibrosis and a mouse model of bleomycin (BLM)-induced PF. Global deletion of Txndc5 markedly reduces the extent of PF and preserves lung function in mice following BLM treatment. Mechanistic investigations demonstrate that TXNDC5 promotes fibrogenesis by enhancing TGFβ1 signaling through direct binding with and stabilization of TGFBR1 in lung fibroblasts. Moreover, TGFβ1 stimulation is shown to upregulate TXNDC5 via ER stress/ATF6-dependent transcriptional control in lung fibroblasts. Inducing fibroblast-specific deletion of Txndc5 mitigates the progression of BLM-induced PF and lung function deterioration. Targeting TXNDC5, therefore, could be a novel therapeutic approach against PF.
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http://dx.doi.org/10.1038/s41467-020-18047-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7449970PMC
August 2020

Adjuvant-free nanofiber vaccine induces in situ lung dendritic cell activation and T17 responses.

Sci Adv 2020 Aug 7;6(32):eaba0995. Epub 2020 Aug 7.

Department of Surgery, The University of Chicago, Chicago, IL 60637, USA.

The current paradigm that subunit vaccines require adjuvants to optimally activate innate immunity implies that increased vaccine reactogenicity will invariably be linked to improved immunogenicity. Countering this paradigm, nanoparticulate vaccines have been reported to act as delivery systems for vaccine antigens and induce immunity without the need for exogenous adjuvants or local inflammation; however, the mechanisms underlying the immunogenicity of nanoparticle vaccines are incompletely identified. Here, we show that antigens displayed on self-assembling nanofiber scaffolds and delivered intranasally are presented by CD103 and CD11b lung dendritic cells that up-regulate CD80 and migrate into the draining lymph node (LN). This was accompanied by a nearly exclusive priming and accumulation of antigen-specific T17 cells occurring independently in both LN and lung. Thus, self-assembling peptide nanofiber vaccines may represent a novel, needle- and adjuvant-free means of eliciting protective immunity against fungal and bacterial infections at skin and mucosal barrier surfaces.
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http://dx.doi.org/10.1126/sciadv.aba0995DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7413739PMC
August 2020

Pro-lymphangiogenic VEGFR-3 signaling modulates memory T cell responses in allergic airway inflammation.

Mucosal Immunol 2021 01 9;14(1):144-151. Epub 2020 Jun 9.

Pritzker School of Molecular Engineering, University of Chicago, Chicago, IL, USA.

In allergic airway inflammation, VEGFR-3-mediated lymphangiogenesis occurs in humans and mouse models, yet its immunological roles, particularly in adaptive immunity, are poorly understood. Here, we explored how pro-lymphangiogenic signaling affects the allergic response to house dust mite (HDM). In the acute inflammatory phase, the lungs of mice treated with blocking antibodies against VEGFR-3 (mF4-31C1) displayed less inflammation overall, with dramatically reduced innate and T-cell numbers and reduced inflammatory chemokine levels. However, when inflammation was allowed to resolve and memory recall was induced 2 months later, mice treated with mF4-31C1 as well as VEGF-C/-D knockout models showed exacerbated type 2 memory response to HDM, with increased Th2 cells, eosinophils, type 2 chemokines, and pathological inflammation scores. This was associated with lower CCL21 and decreased T in the lymph nodes. Together, our data imply that VEGFR-3 activation in allergic airways helps to both initiate the acute inflammatory response and regulate the adaptive (memory) response, possibly in part by shifting the T/Th2 balance. This introduces new immunomodulatory roles for pro-lymphangiogenic VEGFR-3 signaling in allergic airway inflammation and suggests that airway lymphatics may be a novel target for treating allergic responses.
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http://dx.doi.org/10.1038/s41385-020-0308-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7725864PMC
January 2021

Transcriptional programming and T cell receptor repertoires distinguish human lung and lymph node memory T cells.

Commun Biol 2019 13;2:411. Epub 2019 Nov 13.

2Department of Human Genetics, The University of Chicago, Chicago, USA.

Antigen-specific memory T cells persist for years after exposure to a pathogen and provide effective recall responses. Many memory T cell subsets have been identified and differ in abundance throughout tissues. This study focused on CD4 and CD8 memory T cells from paired human lung and lung draining lymph node (LDLN) samples and identified substantial differences in the transcriptional landscape of these subsets, including higher expression of an array of innate immune receptors in lung T cells which were further validated by flow cytometry. Using T cell receptor analysis, we determined the clonal overlap between memory T cell subsets within the lung and within the LDLN, and this was greater than the clonal overlap observed between memory T cell subsets compared across tissues. Our results suggest that lung and LDLN memory T cells originate from different precursor pools, recognize distinct antigens and likely have separate roles in immune responses.
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http://dx.doi.org/10.1038/s42003-019-0657-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6853923PMC
May 2020

T-cell phenotypes are associated with serum IgE levels in Amish and Hutterite children.

J Allergy Clin Immunol 2019 11 8;144(5):1391-1401.e10. Epub 2019 Aug 8.

Department of Medicine, Section of Pulmonary and Critical Care Medicine, University of Chicago, Chicago, Ill; Committee on Immunology, University of Chicago, Chicago, Ill. Electronic address:

Objectives: Amish children raised on traditional farms have lower atopy and asthma risk than Hutterite children raised on modern farms. In our previous study we established that the Amish environment affects the innate immune response to decrease asthma and atopy risk. Here we investigated T-cell phenotypes in the same Amish and Hutterite children as in our earlier study to elucidate how this altered innate immunity affects adaptive T cells.

Methods: Blood was collected from 30 Amish and 30 Hutterite age- and sex-matched children; cells were cryopreserved until analysis. Flow cytometry was used to analyze cell subsets. Atopy was determined based on allergen-specific and total IgE levels.

Results: Children exposed to Amish farms had increased activated regulatory CD4 T-cell phenotypes, whereas conventional CD4 T cells expressed lower levels of costimulation molecules and other activation markers. The increase in numbers of circulating activated regulatory CD4 T cells was associated with an increase in inhibitory receptors on monocytes in Amish, but not Hutterite, children. Strikingly, the Amish children had a higher proportion of CD28 CD8 T cells than the Hutterite children (P < .0001, nonparametric t test), a difference that remained even after accounting for the effects of age and sex (conditional log regression exponential β = 1.08, P = .0053). The proportion of these cells correlated with high T-cell IFN-γ production (r = 0.573, P = .005) and low serum IgE levels (r = -0.417, P = .025). Furthermore, CD28 CD8 T-cell numbers were increased in Amish children, with high expression of the innate genes TNF and TNF-α-induced protein 3 (TNFAIP3) in peripheral blood leukocytes.

Conclusion: Amish children's blood leukocytes are not only altered in their innate immune status but also have distinct T-cell phenotypes that are often associated with increased antigen exposure.
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http://dx.doi.org/10.1016/j.jaci.2019.07.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6842432PMC
November 2019

Improving the Quality and Reproducibility of Flow Cytometry in the Lung. An Official American Thoracic Society Workshop Report.

Am J Respir Cell Mol Biol 2019 08;61(2):150-161

Defining responses of the structural and immune cells in biologic systems is critically important to understanding disease states and responses to injury. This requires accurate and sensitive methods to define cell types in organ systems. The principal method to delineate the cell populations involved in these processes is flow cytometry. Although researchers increasingly use flow cytometry, technical challenges can affect its accuracy and reproducibility, thus significantly limiting scientific advancements. This challenge is particularly critical to lung immunology, as the lung is readily accessible and therefore used in preclinical and clinical studies to define potential therapeutics. Given the importance of flow cytometry in pulmonary research, the American Thoracic Society convened a working group to highlight issues and technical challenges to the performance of high-quality pulmonary flow cytometry, with a goal of improving its quality and reproducibility.
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http://dx.doi.org/10.1165/rcmb.2019-0191STDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6670040PMC
August 2019

Associations between fungal and bacterial microbiota of airways and asthma endotypes.

J Allergy Clin Immunol 2019 11 3;144(5):1214-1227.e7. Epub 2019 Jul 3.

Section of Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, Ill. Electronic address:

Background: The relationship between asthma, atopy, and underlying type 2 (T2) airway inflammation is complex. Although the bacterial airway microbiota is known to differ in asthmatic patients, the fungal and bacterial markers that discriminate T2-high (eosinophilic) and T2-low (neutrophilic/mixed-inflammation) asthma and atopy are still incompletely identified.

Objectives: The aim of this study was to demonstrate the fungal microbiota structure of airways in asthmatic patients associated with T2 inflammation, atopy, and key clinical parameters.

Methods: We collected endobronchial brush (EB) and bronchoalveolar lavage (BAL) samples from 39 asthmatic patients and 19 healthy subjects followed by 16S gene and internal transcribed spacer-based microbiota sequencing. The microbial sequences were classified into exact sequence variants. The T2 phenotype was defined by using a blood eosinophil count with a threshold of 300 cells/μL.

Results: Fungal diversity was significantly lower in EB samples from patients with T2-high compared with T2-low inflammation; key fungal genera enriched in patients with T2-high inflammation included Trichoderma species, whereas Penicillium species was enriched in patients with atopy. In BAL fluid samples the dominant genera were Cladosporium, Fusarium, Aspergillus, and Alternaria. Using generalized linear models, we identified significant associations between specific fungal exact sequence variants and FEV, fraction of exhaled nitric oxide values, BAL fluid cell counts, and corticosteroid use. Investigation of interkingdom (bacterial-fungal) co-occurrence patterns revealed different topologies between asthmatic patients and healthy control subjects. Random forest models with fungal classifiers predicted asthma status with 75% accuracy for BAL fluid samples and 80% accuracy for EB samples.

Conclusions: We demonstrate clear differences in bacterial and fungal microbiota in asthma-associated phenotypes. Our study provides additional support for considering microbial signatures in delineating asthma phenotypes.
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http://dx.doi.org/10.1016/j.jaci.2019.06.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6842419PMC
November 2019

Evidence for an IL-6-high asthma phenotype in asthmatic patients of African ancestry.

J Allergy Clin Immunol 2019 07 25;144(1):304-306.e4. Epub 2019 Apr 25.

Department of Human Genetics, University of Chicago, Chicago, Ill.

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http://dx.doi.org/10.1016/j.jaci.2019.04.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612296PMC
July 2019

Protection against Staphylococcus aureus bacteremia-induced mortality depends on ILC2s and eosinophils.

JCI Insight 2019 03 21;4(6). Epub 2019 Mar 21.

Department of Medicine, Section of Pulmonary and Critical Care, University of Chicago, Chicago, Illinois, USA.

The dysregulated, unbalanced immune response of sepsis results in a mortality exceeding 20%, yet recent findings by our group indicate that patients with allergic, type 2-mediated immune diseases are protected from developing sepsis. We evaluated CD4+ Th cell polarization among patients with Staphylococcus aureus bacteremia and confirmed that survivors had a higher percentage of circulating Th2 cells but lower frequencies of Th17 cells and neutrophils early in the course of infection. To establish the mechanism of this protection, we used a mouse model of lethal S. aureus bacteremia and found that intratracheal pretreatment with the type 2-initiating cytokine IL-33 activated pulmonary type 2 innate lymphoid cells (ILC2s) and promoted eosinophilia. In addition, stimulation of type 2 immunity before lethal infection suppressed the pulmonary neutrophilic response to S. aureus. Mice lacking functional ILC2s did not respond to IL-33 and were not protected from lethal bacteremia, but treatment of these mice with the type 2 cytokines IL-5 and IL-13 rescued them from death. Depletion of eosinophils abrogated IL-33-mediated protection, indicating that eosinophilia is also necessary for the survival benefit. Thus, we have identified a potentially novel mechanism by which type 2 immunity can balance dysregulated septic inflammatory responses, thereby clarifying the protective benefit of type 2 immune diseases on sepsis mortality.
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http://dx.doi.org/10.1172/jci.insight.124168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6482999PMC
March 2019

T cell Co-Stimulatory molecules ICOS and CD28 stratify idiopathic pulmonary fibrosis survival.

Respir Med X 2019 1;1. Epub 2019 Feb 1.

Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, IL, USA.

Idiopathic pulmonary fibrosis (IPF) is a devastating disease that kills as many Americans as breast cancer each year. This study investigated whether lung function decline and survival associates with adaptive immunity in patients with IPF, specifically the expression of checkpoint molecules ICOS, CD28 and PD-1 on circulating CD4 T cells. Clinical data, blood samples and pulmonary function tests were collected prospectively and longitudinally from 59 patients with IPF over a study period of 5 years. Patients were followed until death, lung transplantation, or study end, and cell surface expression of CD45RO, CD28, ICOS, and PD-1 was measured on CD4 T cells via flow cytometry. Repeated measures of ICOS and CD28 on CD4 T cells revealed significant associations between declining ICOS and CD28 expression, and declining lung function parameters FVC and DLCO, independent of age, sex, race, smoking history, or immunosuppressant use. Strikingly, patients in the highest quintile of ICOS at study entry had markedly improved survival, while those with low CD28 fared poorly. No change in PD-1 expression was found. Analysis of ICOS and CD28 from the first blood draw identified three populations of IPF patients; those at high risk for early death, those with intermediate risk, and those at low risk. These results highlight the role of T cell mediated immunity in IPF survival, finding the assessment of two T cell stimulatory checkpoint molecules, CD28 and ICOS, was sufficient to discriminate three distinct survival trajectories over 5 years of patient follow up.
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http://dx.doi.org/10.1016/j.yrmex.2019.100002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7243672PMC
February 2019

Single-Cell Transcriptomic Analysis of Human Lung Provides Insights into the Pathobiology of Pulmonary Fibrosis.

Am J Respir Crit Care Med 2019 06;199(12):1517-1536

1 Division of Pulmonary and Critical Care Medicine, Department of Medicine.

The contributions of diverse cell populations in the human lung to pulmonary fibrosis pathogenesis are poorly understood. Single-cell RNA sequencing can reveal changes within individual cell populations during pulmonary fibrosis that are important for disease pathogenesis. To determine whether single-cell RNA sequencing can reveal disease-related heterogeneity within alveolar macrophages, epithelial cells, or other cell types in lung tissue from subjects with pulmonary fibrosis compared with control subjects. We performed single-cell RNA sequencing on lung tissue obtained from eight transplant donors and eight recipients with pulmonary fibrosis and on one bronchoscopic cryobiospy sample from a patient with idiopathic pulmonary fibrosis. We validated these data using RNA hybridization, immunohistochemistry, and bulk RNA-sequencing on flow-sorted cells from 22 additional subjects. We identified a distinct, novel population of profibrotic alveolar macrophages exclusively in patients with fibrosis. Within epithelial cells, the expression of genes involved in Wnt secretion and response was restricted to nonoverlapping cells. We identified rare cell populations including airway stem cells and senescent cells emerging during pulmonary fibrosis. We developed a web-based tool to explore these data. We generated a single-cell atlas of pulmonary fibrosis. Using this atlas, we demonstrated heterogeneity within alveolar macrophages and epithelial cells from subjects with pulmonary fibrosis. These results support the feasibility of discovery-based approaches using next-generation sequencing technologies to identify signaling pathways for targeting in the development of personalized therapies for patients with pulmonary fibrosis.
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http://dx.doi.org/10.1164/rccm.201712-2410OCDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6580683PMC
June 2019

Non-apoptotic Fas (CD95) Signaling on T Cells Regulates the Resolution of Th2-Mediated Inflammation.

Front Immunol 2018 1;9:2521. Epub 2018 Nov 1.

Committee on Molecular Pathology and Molecular Medicine, Chicago, IL, United States.

Fas (CD95/APO-1) and its ligand (FasL/CD95L) promote the resolution of type 2 lung inflammation and eosinophilia. We previously found that Fas-deficiency on T cells, but not eosinophils, delayed resolution of inflammation. However, Fas can signal both cell death and have a positive signaling function that can actually activate cells. In this study, we investigated whether Fas-induced death or Fas-activated signaling pathways promote resolution of allergic lung inflammation. By increasing T cell survival through two Fas-independent pathways, using Bim-deficient T cells or Bcl-x overexpressing T cells, no differences in resolution of Th2-mediated inflammation was observed. Furthermore, Th2 cells were inherently resistant to Fas-mediated apoptosis and preferentially signaled through non-apoptotic pathways following FasL treatment. Utilizing Fas-mutant mice deficient in apoptotic but sufficient for non-apoptotic Fas signaling pathways, we demonstrate that non-apoptotic Fas signaling in T cells drives resolution of Th2-mediated airway inflammation. Our findings reveal a previously unknown role for non-apoptotic Fas signaling on Th2 cells in the induction of resolution of type 2 inflammation.
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http://dx.doi.org/10.3389/fimmu.2018.02521DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6221963PMC
October 2019

Allergen Exposure in Lymphopenic Fas-Deficient Mice Results in Persistent Eosinophilia Due to Defects in Resolution of Inflammation.

Front Immunol 2018 30;9:2395. Epub 2018 Oct 30.

Section of Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, IL, United States.

Asthma is characterized by chronic airway type-2 inflammation and eosinophilia, yet the mechanisms involved in chronic, non-resolving inflammation remain poorly defined. Previously, our group has found that when Rag-deficient mice were reconstituted with Fas-deficient B6 LPR T cells and sensitized and challenged, the mice developed a prolonged type-2-mediated airway inflammation that continued for more than 6 weeks after the last antigen exposure. Surprisingly, no defect in resolution was found when intact B6 LPR mice or T cell specific Fas-conditional knockout mice were sensitized and challenged. We hypothesize that the homeostatic proliferation induced by adoptive transfer of T cells into Rag-deficient mice may be an important mechanism involved in the lack of resolution. To investigate the role of homeostatic proliferation, we induced lymphopenia in the T cell-specific Fas-conditional knockout mice by non-lethal irradiation and sensitized them when T cells began to repopulate. Interestingly, we found that defective Fas signaling on T cells plus antigen exposure during homeostatic proliferation was sufficient to induce prolonged eosinophilic airway inflammation. In conclusion, our data show that the combination of transient lymphopenia, abnormal Fas-signaling, and antigen exposure leads to the development of a prolonged airway eosinophilic inflammatory phase in our mouse model of experimental asthma.
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http://dx.doi.org/10.3389/fimmu.2018.02395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6219400PMC
October 2019

Prognosticating Outcomes in Interstitial Lung Disease by Mediastinal Lymph Node Assessment. An Observational Cohort Study with Independent Validation.

Am J Respir Crit Care Med 2019 03;199(6):747-759

3 Department of Radiology.

Rationale: Mediastinal lymph node (MLN) enlargement on chest computed tomography (CT) is prevalent in patients with interstitial lung disease (ILD) and may reflect immunologic activation and subsequent cytokine-mediated immune cell trafficking.

Objectives: We aimed to determine whether MLN enlargement on chest CT predicts clinical outcomes and circulating cytokine levels in ILD.

Methods: MLN measurements were obtained from chest CT scans of patients with ILD at baseline evaluation over a 10-year period. Patients with sarcoidosis and drug toxicity-related ILD were excluded. MLN diameter and location were assessed. Plasma cytokine levels were analyzed in a subset of patients. The primary outcome was transplant-free survival (TFS). Secondary outcomes included all-cause and respiratory hospitalizations, lung function, and plasma cytokine concentrations. Cox regression was used to assess mortality risk. Outcomes were assessed in three independent ILD cohorts.

Measurements And Main Results: Chest CT scans were assessed in 1,094 patients (mean age, 64 yr; 52% male). MLN enlargement (≥10 mm) was present in 66% (n = 726) and strongly predicted TFS (hazard ratio [HR], 1.53; 95% confidence interval [CI], 1.12-2.10; P = 0.008) and risk of all-cause and respiratory hospitalizations (internal rate of return [IRR], 1.52; 95% CI, 1.17-1.98; P = 0.002; and IRR, 1.71; 95% CI, 1.15-2.53; P = 0.008, respectively) when compared with subjects with MLN <10 mm. Patients with MLN enlargement had lower lung function and decreased plasma concentrations of soluble CD40L (376 pg/ml vs. 505 pg/ml, P = 0.001) compared with those without MLN enlargement. Plasma IL-10 concentration >45 pg/ml predicted mortality (HR, 4.21; 95% CI, 1.21-14.68; P = 0.024). Independent analysis of external datasets confirmed these findings.

Conclusions: MLN enlargement predicts TFS and hospitalization risk in ILD and is associated with decreased levels of a key circulating cytokine, soluble CD40L. Incorporating MLN and cytokine findings into current prediction models might improve ILD prognostication.
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http://dx.doi.org/10.1164/rccm.201804-0761OCDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6423102PMC
March 2019

Effects of an FcγRIIA polymorphism on leukocyte gene expression and cytokine responses to anti-CD3 and anti-CD28 antibodies.

Genes Immun 2019 07 6;20(6):462-472. Epub 2018 Jul 6.

Department of Human Genetics, University of Chicago, Chicago, IL, 60637, USA.

The low affinity Fcγ receptor, FcγRIIA, harbors a common missense mutation, rs1801274 (G>A, Arg131His) that modifies binding affinity to human IgG2 and mouse IgG1 antibodies and is associated with increased risk of autoimmune disease. Despite the important role of the Arg131His variant, little is understood about heterozygous genotype effects on global gene expression and cytokine production during an FcγR-dependent response. To address this gap in knowledge, we treated human whole-blood samples from 130 individuals with mouse IgG1 anti-CD3 and anti-CD28 antibodies and characterized the genome-wide gene expression profiles and cytokine production among individuals stratified by rs1801274 genotype. Our analysis revealed that the levels of four cytokines (IFNγ, IL-12, IL-2, TNFα) and global gene expression patterns differed between all three genotype classes. Surprisingly, the heterozygotes showed suboptimal T cell activation compared to cells from individuals homozygous for the higher-affinity FcγRIIA allele (GG; Arg/Arg). The results of this study demonstrate that IgG response varies among all rs1801274 genotype classes and results in profound differences in both cytokine responses and gene expression patterns in blood leukocytes. Because even heterozygotes showed dampened global responses, our data may provide insight into the heterogeneity of outcomes in cytokine release assays and immunotherapy efficacy.
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http://dx.doi.org/10.1038/s41435-018-0038-8DOI Listing
July 2019

Distinct T-helper cell responses to Staphylococcus aureus bacteremia reflect immunologic comorbidities and correlate with mortality.

Crit Care 2018 Apr 25;22(1):107. Epub 2018 Apr 25.

Section of Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, IL, USA.

Background: The dysregulated host immune response that defines sepsis varies as a function of both the immune status of the host and the distinct nature of the pathogen. The degree to which immunocompromising comorbidities or immunosuppressive medications affect the immune response to infection is poorly understood because these patients are often excluded from studies about septic immunity. The objectives of this study were to determine the immune response to a single pathogen (Staphylococcus aureus) among a diverse case mix of patients and to determine whether comorbidities affect immune and clinical outcomes.

Methods: Blood samples were drawn from 95 adult inpatients at multiple time points after the first positive S. aureus blood culture. Cox proportional hazards modeling was used to determine the associations between admission neutrophil counts, admission lymphocyte counts, cytokine levels, and 90-day mortality. A nested case-control flow cytometric analysis was conducted to determine T-helper type 1 (Th1), Th2, Th17, and regulatory T-cell (Treg) subsets among a subgroup of 28 patients. In a secondary analysis, we categorized patients as either having immunocompromising disorders (human immunodeficiency virus and hematologic malignancies), receiving immunosuppressive medications, or being not immunocompromised.

Results: Higher neutrophil-to-lymphocyte count ratios and higher Th17 cytokine responses relative to Th1 cytokine responses early after infection were independently associated with mortality and did not depend on the immune state of the patient (HR 1.93, 95% CI 1.17-3.17, p = 0.01; and HR 1.13, 95% CI 1.01-1.27, p = 0.03, respectively). On the basis of flow cytometric analysis of CD4 T-helper subsets, an increasing Th17/Treg response over the course of the infection was most strongly associated with increased mortality (HR 4.41, 95% CI 1.69-11.5, p < 0.01). This type of immune response was most common among patients who were not immunocompromised. In contrast, among immunocompromised patients who died, a decreasing Th1/Treg response was most common.

Conclusions: The association of both increased Th17 responses and increased neutrophil counts relative to lymphocyte counts with mortality suggests that an overwhelming inflammatory response is detrimental. However, the differential responses of patients according to immune state suggest that immune status is an important clinical indicator that should be accounted for in the management of septic patients, as well as in the development of novel immunomodulatory therapies.
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http://dx.doi.org/10.1186/s13054-018-2025-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5916828PMC
April 2018

IL-33 Drives Monocyte Recruitment to Lung Interstitium through Chemokine Upregulation.

Immunohorizons 2017 Aug 15;1(6):101-108. Epub 2017 Aug 15.

Committee on Immunology, University of Chicago, Chicago, IL 60637.

Tissue infiltration by circulating monocytes is a critical step in the initiation and augmentation of type 2 inflammatory responses in the lungs. Our studies demonstrate that IL-33 mice have a defect in monocyte extravasation from the vasculature to the lung interstitium during induction of type 2 inflammatory responses. This result suggests that monocyte migration to the lungs is IL-33 dependent, and we found that administration of exogenous recombinant IL-33 is sufficient to restore monocyte localization to the lung interstitium. Further investigation of the effect of early administration of recombinant IL-33 on the lungs identified upregulation of multiple chemokines including the monocyte chemoattractants CCL2, CCL7, and CCL22. Importantly, blockade of G-protein coupled receptor-dependent signaling, and thereby chemokine receptor activity, inhibited IL-33-driven monocyte recruitment. CCR2 deficiency prevented recruitment of monocytes to the lung extravascular space during allergic sensitization, and resulted in reduced eosinophilia after allergen challenge. Thus, IL-33 plays a critical role in the initiation of type 2 inflammatory responses by inducing upregulation of chemokines that promote monocyte recruitment to the lung interstitium.
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http://dx.doi.org/10.4049/immunohorizons.1700024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5889047PMC
August 2017

Immune development and environment: lessons from Amish and Hutterite children.

Curr Opin Immunol 2017 Oct 29;48:51-60. Epub 2017 Aug 29.

Department of Cellular and Molecular Medicine, Asthma and Airway Disease Research Center, and Bio5 Institute, The University of Arizona, Tucson, AZ 85724, USA.

Children who grow up in traditional farm environments are protected from developing asthma and allergy. This 'farm effect' can be largely explained by the child's early life contact with farm animals, in particular cows, and their microbes. Our studies in Amish and Hutterite school children living on farms in the U.S. have further demonstrated that this protection is mediated through innate immune pathways. Although very similar with respect to ancestry and many lifestyle factors that are associated with asthma risk, Amish and Hutterites follow farming practices that are associated with profound differences in the levels of house dust endotoxin, in the prevalence of asthma and atopy among school children, and in the proportions, phenotypes, and functions of immune cells from these children. In this review, we will consider our studies in Amish and Hutterites children in the context of the many previous studies in European farm children and discuss how these studies have advanced our understanding of the asthma-protective 'farm effect'.
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http://dx.doi.org/10.1016/j.coi.2017.08.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5682224PMC
October 2017

DNA methylation in lung cells is associated with asthma endotypes and genetic risk.

JCI Insight 2016 12 8;1(20):e90151. Epub 2016 Dec 8.

Department of Human Genetics.

The epigenome provides a substrate through which environmental exposures can exert their effects on gene expression and disease risk, but the relative importance of epigenetic variation on human disease onset and progression is poorly characterized. Asthma is a heterogeneous disease of the airways, for which both onset and clinical course result from interactions between host genotype and environmental exposures, yet little is known about the molecular mechanisms for these interactions. We assessed genome-wide DNA methylation using the Infinium Human Methylation 450K Bead Chip and characterized the transcriptome by RNA sequencing in primary airway epithelial cells from 74 asthmatic and 41 nonasthmatic adults. Asthma status was based on doctor's diagnosis and current medication use. Genotyping was performed using various Illumina platforms. Our study revealed a regulatory locus on chromosome 17q12-21 associated with asthma risk and epigenetic signatures of specific asthma endotypes and molecular networks. Overall, these data support a central role for DNA methylation in lung cells, which promotes distinct molecular pathways of asthma pathogenesis and modulates the effects of genetic variation on disease risk and clinical heterogeneity.
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http://dx.doi.org/10.1172/jci.insight.90151DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5139904PMC
December 2016

Skewed Lung CCR4 to CCR6 CD4 T Cell Ratio in Idiopathic Pulmonary Fibrosis Is Associated with Pulmonary Function.

Front Immunol 2016 23;7:516. Epub 2016 Nov 23.

Section of Pulmonary and Critical Care, Department of Medicine, University of Chicago, Chicago, IL, USA; Committee on Immunology, University of Chicago, Chicago, IL, USA.

Rationale: Idiopathic pulmonary fibrosis (IPF) is a progressive, fatal lung disease. While it has been suggested that T cells may contribute to IPF pathogenesis, these studies have focused primarily on T cells outside of the pulmonary interstitium. Thus, the role of T cells in the diseased lung tissue remains unclear.

Objective: To identify whether specific CD4 T cell subsets are differentially represented in lung tissue from patients with IPF.

Methods: CD4 T cell subsets were measured in lung tissue obtained from patients with IPF at the time of lung transplantation, and from age- and gender-matched organ donors with no known lung disease. Subsets were identified by their surface expression of CCR4, CCR6, and CXCR3 chemokine receptors. CD4 T cell subsets were correlated with measurements of lung function obtained prior to transplantation.

Results: Compared to controls, IPF patients had a higher proportion of lung CD4 T cells, a higher proportion of CCR4 CD4 T cells, and a lower proportion of CCR6 CD4 T cells. The increase in CCR4 CD4 T cells in IPF lung tissue was not due to increased Tregs. Intriguingly, the increase in the ratio of CCR4 cells to CCR6 cells correlated significantly with better lung function.

Conclusion: Our findings suggest a new paradigm that not all T cell infiltrates in IPF lungs are detrimental, but instead, specialized subsets may actually be protective. Thus, augmentation of the chemokines that recruit protective T cells, while blocking chemokines that recruit detrimental T cells, may constitute a novel approach to IPF therapy.
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http://dx.doi.org/10.3389/fimmu.2016.00516DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5120085PMC
November 2016

Innate Immunity and Asthma Risk in Amish and Hutterite Farm Children.

N Engl J Med 2016 Aug;375(5):411-421

Department of Human Genetics (M.M. Stein, C.I., R.L.A., C.O.), the Department of Medicine, Section of Pulmonary and Critical Care Medicine, and the Committee on Immunology (C.L.H., A.I.S.), the Department of Ecology and Evolution (J.A.G.), and the Department of Surgery (J.A.G.), University of Chicago, Chicago, and the Institute for Genomic and Systems Biology, Argonne National Laboratory, Argonne (J.A.G.) - all in Illinois; the NIEHS Training Program in Environmental Toxicology and Graduate Program in Cellular and Molecular Medicine (J.G.), and the Departments of Cellular and Molecular Medicine (V.P., D.V.), Medicine (J.G.L.), Chemical and Environmental Engineering (M. Marques dos Santos), and Soil, Water, and Environmental Science (J.W.N., R.M.M.), University of Arizona, and the Arizona Respiratory Center and Bio5 Institute (J.G., V.P., S.E.M., J.G.L., F.D.M., D.V.) - all in Tucson; the Department of Occupational and Environmental Health, University of Iowa, Iowa City (N.M., P.S.T.); Allergy and Asthma Consultants, Indianapolis (M.H.); and Dr. von Hauner Children's Hospital, Ludwig Maximilians University Munich, Munich, Germany (E.M.).

Background: The Amish and Hutterites are U.S. agricultural populations whose lifestyles are remarkably similar in many respects but whose farming practices, in particular, are distinct; the former follow traditional farming practices whereas the latter use industrialized farming practices. The populations also show striking disparities in the prevalence of asthma, and little is known about the immune responses underlying these disparities.

Methods: We studied environmental exposures, genetic ancestry, and immune profiles among 60 Amish and Hutterite children, measuring levels of allergens and endotoxins and assessing the microbiome composition of indoor dust samples. Whole blood was collected to measure serum IgE levels, cytokine responses, and gene expression, and peripheral-blood leukocytes were phenotyped with flow cytometry. The effects of dust extracts obtained from Amish and Hutterite homes on immune and airway responses were assessed in a murine model of experimental allergic asthma.

Results: Despite the similar genetic ancestries and lifestyles of Amish and Hutterite children, the prevalence of asthma and allergic sensitization was 4 and 6 times as low in the Amish, whereas median endotoxin levels in Amish house dust was 6.8 times as high. Differences in microbial composition were also observed in dust samples from Amish and Hutterite homes. Profound differences in the proportions, phenotypes, and functions of innate immune cells were also found between the two groups of children. In a mouse model of experimental allergic asthma, the intranasal instillation of dust extracts from Amish but not Hutterite homes significantly inhibited airway hyperreactivity and eosinophilia. These protective effects were abrogated in mice that were deficient in MyD88 and Trif, molecules that are critical in innate immune signaling.

Conclusions: The results of our studies in humans and mice indicate that the Amish environment provides protection against asthma by engaging and shaping the innate immune response. (Funded by the National Institutes of Health and others.).
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http://dx.doi.org/10.1056/NEJMoa1508749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5137793PMC
August 2016

Mapping Variation in Cellular and Transcriptional Response to 1,25-Dihydroxyvitamin D3 in Peripheral Blood Mononuclear Cells.

PLoS One 2016 25;11(7):e0159779. Epub 2016 Jul 25.

Department of Human Genetics, University of Chicago, Chicago, Illinois, United States of America.

The active hormonal form of vitamin D, 1,25-dihydroxyvitamin D (1,25D) is an important modulator of the immune system, inhibiting cellular proliferation and regulating transcription of immune response genes. In order to characterize the genetic basis of variation in the immunomodulatory effects of 1,25D, we mapped quantitative traits of 1,25D response at both the cellular and the transcriptional level. We carried out a genome-wide association scan of percent inhibition of cell proliferation (Imax) induced by 1,25D treatment of peripheral blood mononuclear cells from 88 healthy African-American individuals. Two genome-wide significant variants were identified: rs1893662 in a gene desert on chromosome 18 (p = 2.32 x 10-8) and rs6451692 on chromosome 5 (p = 2.55 x 10-8), which may influence the anti-proliferative activity of 1,25D by regulating the expression of nearby genes such as the chemokine gene, CCL28, and the translation initiation gene, PAIP1. We also identified 8 expression quantitative trait loci at a FDR<0.10 for transcriptional response to 1,25D treatment, which include the transcriptional regulator ets variant 3-like (ETV3L) and EH-domain containing 4 (EHD4). In addition, we identified response eQTLs in vitamin D receptor binding sites near genes differentially expressed in response to 1,25D, such as FERM Domain Containing 6 (FRMD6), which plays a critical role in regulating both cell proliferation and apoptosis. Combining information from the GWAS of Imax and the response eQTL mapping enabled identification of putative Imax-associated candidate genes such as PAIP1 and the transcriptional repressor gene ZNF649. Overall, the variants identified in this study are strong candidates for immune traits and diseases linked to vitamin D, such as multiple sclerosis.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0159779PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4959717PMC
July 2017

Corticosteroid therapy and airflow obstruction influence the bronchial microbiome, which is distinct from that of bronchoalveolar lavage in asthmatic airways.

J Allergy Clin Immunol 2016 05 25;137(5):1398-1405.e3. Epub 2015 Nov 25.

Section of Pulmonary and Critical Care Medicine, Department of Medicine, University of Chicago, Chicago, Ill. Electronic address:

Background: The lung has a diverse microbiome that is modest in biomass. This microbiome differs in asthmatic patients compared with control subjects, but the effects of clinical characteristics on the microbial community composition and structure are not clear.

Objectives: We examined whether the composition and structure of the lower airway microbiome correlated with clinical characteristics of chronic persistent asthma, including airflow obstruction, use of corticosteroid medications, and presence of airway eosinophilia.

Methods: DNA was extracted from endobronchial brushings and bronchoalveolar lavage fluid collected from 39 asthmatic patients and 19 control subjects, along with negative control samples. 16S rRNA V4 amplicon sequencing was used to compare the relative abundance of bacterial genera with clinical characteristics.

Results: Differential feature selection analysis revealed significant differences in microbial diversity between brush and lavage samples from asthmatic patients and control subjects. Lactobacillus, Pseudomonas, and Rickettsia species were significantly enriched in samples from asthmatic patients, whereas Prevotella, Streptococcus, and Veillonella species were enriched in brush samples from control subjects. Generalized linear models on brush samples demonstrated oral corticosteroid use as an important factor affecting the relative abundance of the taxa that were significantly enriched in asthmatic patients. In addition, bacterial α-diversity in brush samples from asthmatic patients was correlated with FEV1 and the proportion of lavage eosinophils.

Conclusion: The diversity and composition of the bronchial airway microbiome of asthmatic patients is distinct from that of nonasthmatic control subjects and influenced by worsening airflow obstruction and corticosteroid use.
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http://dx.doi.org/10.1016/j.jaci.2015.10.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4860110PMC
May 2016

Intrinsic functional defects of type 2 innate lymphoid cells impair innate allergic inflammation in promyelocytic leukemia zinc finger (PLZF)-deficient mice.

J Allergy Clin Immunol 2016 Feb 23;137(2):591-600.e1. Epub 2015 Oct 23.

Committee on Immunology, University of Chicago, Chicago, Ill; Department of Pathology, University of Chicago, Chicago, Ill. Electronic address:

Background: The transcription factor promyelocytic leukemia zinc finger (PLZF) is transiently expressed during development of type 2 innate lymphoid cells (ILC2s) but is not present at the mature stage. We hypothesized that PLZF-deficient ILC2s have functional defects in the innate allergic response and represent a tool for studying innate immunity in a mouse with a functional adaptive immune response.

Objective: We determined the consequences of PLZF deficiency on ILC2 function in response to innate and adaptive immune stimuli by using PLZF(-/-) mice and mixed wild-type:PLZF(-/-) bone marrow chimeras.

Methods: PLZF(-/-) mice, wild-type littermates, or mixed bone marrow chimeras were treated with the protease allergen papain or the cytokines IL-25 and IL-33 or infected with the helminth Nippostrongylus brasiliensis to induce innate type 2 allergic responses. Mice were sensitized with intraperitoneal ovalbumin-alum, followed by intranasal challenge with ovalbumin alone, to induce adaptive TH2 responses. Lungs were analyzed for immune cell subsets, and alveolar lavage fluid was analyzed for ILC2-derived cytokines. In addition, ILC2s were stimulated ex vivo for their capacity to release type 2 cytokines.

Results: PLZF-deficient lung ILC2s exhibit a cell-intrinsic defect in the secretion of IL-5 and IL-13 in response to innate stimuli, resulting in defective recruitment of eosinophils and goblet cell hyperplasia. In contrast, the adaptive allergic inflammatory response to ovalbumin and alum was unimpaired.

Conclusions: PLZF expression at the innate lymphoid cell precursor stage has a long-range effect on the functional properties of mature ILC2s and highlights the importance of these cells for innate allergic responses in otherwise immunocompetent mice.
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http://dx.doi.org/10.1016/j.jaci.2015.07.050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4747811PMC
February 2016