Single cell RNA sequencing reveals distinct clusters of Irf8-expressing pulmonary conventional dendritic cells.

A single population of interferon-regulatory factor 8 (Irf8)-dependent conventional dendritic cell (cDC type1) is considered to be responsible for both immunogenic and tolerogenic responses depending on the surrounding cytokine . Here, we challenge this concept of an omnipotent single Irf8-dependent cDC1 cluster through analysis of pulmonary cDCs at single cell resolution. We report existence of a pulmonary cDC1 cluster lacking Xcr1 with an immunogenic signature that clearly differs from the Xcr1 positive cDC1 cluster.

Competitive survival of clonal serial isolates from cystic fibrosis airways in human neutrophils.

Chronic airway infections with are the major co-morbidity in most people with cystic fibrosis (CF) sustained by neutrophils as the major drivers of lung inflammation, damage, and remodeling. Phagocytosis assays were performed with clonal consortia of longitudinal airway isolates collected from people with CF since the onset of lung colonization until patient's death or replacement by another clone. The extra- and intracellular abundance of individual strains was assessed by deep amplicon sequencing of strain-specific single nucleotide variants in the bacterial genome.

neutrophil migration is associated with inhaled corticosteroid treatment and serum cytokines in pediatric asthma.

Different asthma phenotypes are driven by molecular endotypes. A Th1-high phenotype is linked to severe, therapy-refractory asthma, subclinical infections and neutrophil inflammation. Previously, we found neutrophil granulocytes (NGs) from asthmatics exhibit decreased chemotaxis towards leukotriene B4 (LTB), a chemoattractant involved in inflammation response.

Interleukin-17A and interleukin-22 production by conventional and non-conventional lymphocytes in three different end-stage lung diseases.

Objectives: The contribution of adaptive innate lymphocytes to IL-17A and IL-22 secretion at the end stage of chronic lung diseases remains largely unexplored. In order to uncover tissue- and disease-specific secretion patterns, we compared production patterns of IL-17A and IL-22 in three different human end-stage lung disease entities.

Methods: Production of IL-17A, IL-22 and associated cytokines was assessed in supernatants of re-stimulated lymphocytes by multiplex assays and multicolour flow cytometry of conventional T cells, iNKT cells, γδ T cells and innate lymphoid cells in bronchial lymph node and lung tissue from patients with emphysema ( = 19), idiopathic pulmonary fibrosis ( = 14) and cystic fibrosis ( = 23), as well as lung donors ( = 17).

IgA memory B-cells are significantly increased in patients with asthma and small airway dysfunction.

Background: Comprehensive studies investigated the role of T-cells in asthma which led to personalised treatment options targeting severe eosinophilic asthma. However, little is known about the contribution of B-cells to this chronic inflammatory disease. In this study we investigated the contribution of various B-cell populations to specific clinical features in asthma.

Loss of vascular endothelial notch signaling promotes spontaneous formation of tertiary lymphoid structures.

Tertiary lymphoid structures (TLS) are lymph node-like immune cell clusters that emerge during chronic inflammation in non-lymphoid organs like the kidney, but their origin remains not well understood. Here we show, using conditional deletion strategies of the canonical Notch signaling mediator Rbpj, that loss of endothelial Notch signaling in adult mice induces the spontaneous formation of bona fide TLS in the kidney, liver and lung, based on molecular, cellular and structural criteria. These TLS form in a stereotypical manner around parenchymal arteries, while secondary lymphoid structures remained largely unchanged.

T2-high asthma phenotypes across lifespan.

Rationale: In adults, personalised asthma treatment targets patients with type 2 (T2)-high and eosinophilic asthma phenotypes. It is unclear whether such classification is achievable in children.

Objectives: To define T2-high asthma with easily accessible biomarkers and compare resulting phenotypes across all ages.

Rescue from Pseudomonas aeruginosa Airway Infection via Stem Cell Transplantation.

Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, which lead to impaired ion transport in epithelial cells. Although lung failure due to chronic infection is the major comorbidity in individuals with cystic fibrosis, the role of CFTR in non-epithelial cells has not been definitively resolved. Given the important role of host defense cells, we evaluated the Cftr deficiency in pulmonary immune cells by hematopoietic stem cell transplantation in cystic fibrosis mice.

IL-17 regulates DC migration to the peribronchial LNs and allergen presentation in experimental allergic asthma.

IL-17 is associated with different phenotypes of asthma, however, it is not fully elucidated how it influences induction and maintenance of asthma and allergy. In order to determine the role of IL-17 in development of allergic asthma, we used IL-17A/F double KO (IL-17A/F KO) and WT mice with or without neutralization of IL-17 in an experimental allergic asthma model and analyzed airway hyperresponsiveness, lung inflammation, T helper cell polarization, and DCs influx and activation. We report that the absence of IL-17 reduced influx of DCs into lungs and lung draining LNs.

A comparison of curated gene sets versus transcriptomics-derived gene signatures for detecting pathway activation in immune cells.

Background: Despite the significant contribution of transcriptomics to the fields of biological and biomedical research, interpreting long lists of significantly differentially expressed genes remains a challenging step in the analysis process. Gene set enrichment analysis is a standard approach for summarizing differentially expressed genes into pathways or other gene groupings. Here, we explore an alternative approach to utilizing gene sets from curated databases.

Absence of Regulatory T Cells Causes Phenotypic and Functional Switch in Murine Peritoneal Macrophages.

Tissue macrophages are important components of tissue homeostasis and inflammatory pathologies. In the peritoneal cavity, resident macrophages interact with a variety of immune cells and can exhibit broad range of phenotypes and functions. Forkhead-box-P3 (FOXP3) regulatory T cells (Tregs) play an indispensable role in maintaining immunological tolerance, yet whether, and how the pathological condition that results from the lack of functional Tregs affects peritoneal macrophages (PM) is largely unknown.

Improved protocol for simultaneous analysis of leukocyte subsets and epithelial cells from murine and human lung.

Purpose: To study and isolate lung cells by flow cytometry, enzymatic digestion and generation of single cell suspensions is required. This significantly influences expression of cellular epitopes and protocols need to be adapted for the best isolation and subsequent analysis of specific cellular subsets.

Materials And Methods: We optimized protocols for the simultaneous isolation and characterization of specific human and murine lung cell types.

Suppression of Th17-polarized airway inflammation by rapamycin.

Because Th17-polarized airway inflammation correlates with poor control in bronchial asthma and is a feature of numerous other difficult-to-treat inflammatory lung diseases, new therapeutic approaches for this type of airway inflammation are necessary. We assessed different licensed anti-inflammatory agents with known or expected efficacy against Th17-polarization in mouse models of Th17-dependent airway inflammation. Upon intravenous transfer of in vitro derived Th17 cells and intranasal challenge with the corresponding antigen, we established acute and chronic murine models of Th17-polarised airway inflammation.

B cells control maternofetal priming of allergy and tolerance in a murine model of allergic airway inflammation.

Background: Allergic asthma is a chronic lung disease resulting from inappropriate immune responses to environmental antigens. Early tolerance induction is an attractive approach for primary prevention of asthma.

Objective: We analyzed the mechanisms of perinatal tolerance induction to allergens, with particular focus on the role of B cells in preconception and early intrauterine immune priming.

Gain-of-function STAT1 mutations are associated with intracranial aneurysms.

Chronic mucocutaneous candidiasis, characterized by persistent or recurrent fungal infections, represents the clinical hallmark in gain-of-function (GOF) signal transducer and activator of transcription 1 (STAT1) mutation carriers. Several cases of intracranial aneurysms have been reported in patients with GOF STAT1 mutation but the paucity of reported cases likely suggested this association still as serendipity. In order to endorse this association, we link the development of intracranial aneurysms with STAT1 GOF mutation by presenting the two different cases of a patient and her mother, and demonstrate upregulated phosphorylated STAT4 and IL-12 receptor β1 upon stimulation in patient's blood cells.

B cell subsets are modulated during allergic airway inflammation but are not required for the development of respiratory tolerance in a murine model.

Allergic asthma is a widespread chronic inflammatory disease of the airways. The role of different B cell subsets in developing asthma and respiratory tolerance is not well known. Especially regulatory B (Breg) cells are proposed to be important in asthma regulation.

IL-27 Is Essential for Suppression of Experimental Allergic Asthma by the TLR7/8 Agonist R848 (Resiquimod).

Different models of experimental allergic asthma have shown that the TLR7/8 agonist resiquimod (R848) is a potential inhibitor of type 2 helper cell-driven inflammatory responses. However, the mechanisms mediating its therapeutic effects are not fully understood. Using a model of experimental allergic asthma, we show that induction of IL-27 by R848 is critical for the observed ameliorative effects.

Pulmonary transplantation of macrophage progenitors as effective and long-lasting therapy for hereditary pulmonary alveolar proteinosis.

Hereditary pulmonary alveolar proteinosis (herPAP) is a rare lung disease caused by mutations in the granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor genes, resulting in disturbed alveolar macrophage differentiation, massive alveolar proteinosis, and life-threatening respiratory insufficiency. So far, the only effective treatment for herPAP is repetitive whole-lung lavage, a merely symptomatic and highly invasive procedure. We introduce pulmonary transplantation of macrophage progenitors as effective and long-lasting therapy for herPAP.

Polysialic acid on neuropilin-2 is exclusively synthesized by the polysialyltransferase ST8SiaIV and attached to mucin-type o-glycans located between the b2 and c domain.

Neuropilin-2 (NRP2) is well known as a co-receptor for class 3 semaphorins and vascular endothelial growth factors, involved in axon guidance and angiogenesis. Moreover, NRP2 was shown to promote chemotactic migration of human monocyte-derived dendritic cells (DCs) toward the chemokine CCL21, a function that relies on the presence of polysialic acid (polySia). In vertebrates, this posttranslational modification is predominantly found on the neural cell adhesion molecule (NCAM), where it is synthesized on N-glycans by either of the two polysialyltransferases, ST8SiaII or ST8SiaIV.

Priming of CD8+ T cells against cytomegalovirus-encoded antigens is dominated by cross-presentation.

CMV can infect dendritic cells (DCs), and direct Ag presentation could, therefore, lead to the priming of CMV-specific CD8(+) T cells. However, CMV-encoded immune evasins severely impair Ag presentation in the MHC class I pathway; thus, it is widely assumed that cross-presentation drives the priming of antiviral T cells. We assessed the contribution of direct versus cross priming in mouse CMV (MCMV) infection using recombinant viruses.

Toll-like receptor-2 agonist-allergen coupling efficiently redirects Th2 cell responses and inhibits allergic airway eosinophilia.

Toll-like receptor (TLR) agonists beneficially modulate allergic airway inflammation. However, the efficiency of TLR agonists varies considerably, and their exact cellular mechanisms (especially of TLR 2/6 agonists) are incompletely understood. We investigated at a cellular level whether the administration of the pharmacologically improved TLR2/6 agonist S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol (BPP) conjugated to antigenic peptide (BPP-OVA) could divert an existing Th2 response and influence airway eosinophilia.

The synthetic TLR2 agonist BPPcysMPEG leads to efficient cross-priming against co-administered and linked antigens.

The property of DC to generate effective CTL responses is influenced by TLR signaling. TLR ligands contain molecular signatures associated with pathogens, have an impact on the antigen processing and presentation by DC, and are being exploited as potential adjuvants. We hypothesized that the TLR2/6 heterodimer agonist S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxyl polyethylene glycol (BPP), a synthetic derivative of the Mycoplasma macrophage activating lipopeptide-2, is a potent adjuvant for cross-priming against cellular antigens.

Contribution of direct and cross-presentation to CTL immunity against herpes simplex virus 1.

Dendritic cells (DC), which can be subdivided into different phenotypic and functional subsets, play a pivotal role in the generation of cytotoxic T cell immunity against viral infections. Understanding the modes of Ag acquisition, processing and presentation by DC is essential for the design of effective antiviral vaccines. We aimed to assess the contribution of direct vs cross-presentation for the induction of HSV1-specific CD8(+) T lymphocyte responses in mice.