Rhinovirus-induced neutrophil extracellular traps (NETs) contribute to acute asthma exacerbations; however, the molecular factors that trigger NETosis in this context remain ill-defined. Here, we sought to implicate a role for IL-33, an epithelial cell-derived alarmin rapidly released in response to infection. In mice with chronic experimental asthma (CEA), but not naïve controls, rhinovirus inoculation induced an early (1day post infection; dpi) inflammatory response dominated by neutrophils, neutrophil-associated cytokines (IL-1α, IL-1β, CXCL1), and NETosis, followed by a later, type-2 inflammatory phase (3-7 dpi), characterised by eosinophils, elevated IL-4 levels, and goblet cell hyperplasia. Notably, both phases were ablated by HpARI (Heligmosomoides polygyrus Alarmin Release Inhibitor), which blocks IL-33 release and signalling. Instillation of exogenous IL-33 recapitulated the rhinovirus-induced early phase, including the increased presence of NETs in the airway mucosa, in a PAD4-dependent manner. Ex vivo IL-33-stimulated neutrophils from mice with CEA, but not naïve mice, underwent NETosis and produced greater amounts of IL-1α/β, IL-4, and IL-5. In nasal samples from rhinovirus-infected people with asthma, but not healthy controls, IL-33 levels correlated with neutrophil elastase and dsDNA. Our findings suggest that IL-33 blockade ameliorates the severity of an asthma exacerbation by attenuating neutrophil recruitment and the downstream generation of NETs.

Mast cells (MCs) accumulate in the epithelium of patients with eosinophilic esophagitis (EoE), an inflammatory disorder characterized by extensive esophageal eosinophilic infiltration. Esophageal barrier dysfunction plays an important role in the pathophysiology of EoE. We hypothesized that MCs contribute to the observed impaired esophageal epithelial barrier. Herein, we demonstrate that coculture of differentiated esophageal epithelial cells with immunoglobulin E-activated MCs significanly decreased epithelial resistance by 30% and increased permeability by 22% compared with non-activated MCs. These changes were associated with decreased messenger RNA expression of barrier proteins filaggrin, desmoglein-1 and involucrin, and antiprotease serine peptidase inhibitor kazal type 7. Using targeted proteomics, we detected various cytokines in coculture supernatants, most notably granulocyte-macrophage colony-stimulating factor and oncostatin M (OSM). OSM expression was increased by 12-fold in active EoE and associated with MC marker genes. Furthermore, OSM receptor-expressing esophageal epithelial cells were found in the esophageal tissue of patients with EoE, suggesting that the epithelial cells may respond to OSM. Stimulation of esophageal epithelial cells with OSM resulted in a dose-dependent decrease in barrier function and expression of filaggrin and desmoglein-1 and an increase in protease calpain-14. Taken together, these data suggest a role for MCs in decreasing esophageal epithelial barrier function in EoE, which may in part be mediated by OSM.

Mucosa-associated invariant T (MAIT) cells are the largest population of unconventional T cells in humans. These antimicrobial T cells are poised with rapid effector responses following recognition of the cognate riboflavin (vitamin B2)-like metabolite antigens derived from microbial riboflavin biosynthetic pathway. Presentation of this unique class of small molecule metabolite antigens is mediated by the highly evolutionarily conserved major histocompatibility complex class I-related protein. In humans, MAIT cells are widely found along the upper and lower gastrointestinal tracts owing to their high expression of chemokine receptors and homing molecules directing them to these tissue sites. In this review, we discuss recent findings regarding the roles MAIT cells play in various gastrointestinal bacterial infections, and how their roles appear to differ depending on the etiological agents and the anatomical location. We further discuss the potential mechanisms by which MAIT cells contribute to pathogen control, orchestrate adaptive immunity, as well as their potential contribution to inflammation and tissue damage during gastrointestinal bacterial infections, and the ensuing tissue repair following resolution. Finally, we propose and discuss the use of the emerging three-dimensional organoid technology to test different hypotheses regarding the role of MAIT cells in gastrointestinal bacterial infections, inflammation, and immunity.

Eosinophilic esophagitis (EoE) is a Th2-mediated inflammatory disorder characterized endoscopically by eosinophilic infiltration leading to fibrosis of the esophagus. Stem cell factor (SCF), a multifunctional cytokine, is upregulated in several allergic diseases, including in patients with EoE. Mast cells and eosinophils express c-kit, the cell surface receptor for SCF, and have been found to play an important role in EoE. Therefore, we investigated whether blocking SCF represents a potential therapeutic approach for EoE. Esophageal inflammation was induced in mice using peanut allergen. In mice with experimental EoE, we found that SCF was upregulated in the esophageal tissue. In EoE mice injected with a polyclonal antibody specific for SCF, we observed a decrease in both mast cells and eosinophils by histological and flow cytometric analysis. Furthermore, Th2 cytokines in the esophagus were decreased in anti-SCF treated mice, as were levels of Th2 cytokines from lung-draining and esophageal lymph nodes. Serum levels of peanut-specific IgE were reduced following treatment with anti-SCF. In Kitlf/f-Col1-Cre-ERT mice, which have SCF deleted primarily in myofibroblasts that develop in EoE, we observed similar results as the anti-SCF treated animals for inflammatory cell accumulation, cytokines and histopathology. These results indicate that therapeutic treatments targeting SCF can reduce allergic inflammation in EoE.

T follicular helper (Tfh) cells are an important component of germinal center (GC)-mediated humoral immunity. Yet, how a chronic type 1 versus protective type 2 helminth infection modulates Tfh-GC responses remains poorly understood. Here, we employ the helminth Trichuris muris model and demonstrate that Tfh cell phenotypes and GC are differentially regulated in acute versus chronic infection. The latter failed to induce Tfh-GC B cell responses, with Tfh cells expressing Τ-bet and IFN-γ. In contrast, IL-4-producing Tfh cells dominate responses to an acute, resolving infection. Heightened expression and increased chromatin accessibility of Th1- and Th2 cell-associated genes is observed in chronic and acute induced Tfh cells, respectively. Blockade of the Th1 cell response by T cell-intrinsic T-bet deletion promoted Tfh cell expansion during chronic infection, pointing to a correlation between a robust Tfh cell response and protective immunity to parasites. Finally, blockade of Tfh-GC interactions impaired type 2 immunity, revealing the critical protective role of GC-dependent Th2-like Tfh cell responses during acute infection. Collectively, these results provide new insights into the protective roles of Tfh-GC responses and identify distinct transcriptional and epigenetic features of Tfh cells that emerge during resolving or chronic Trichuris muris infection.

Group 3 innate lymphoid cells (ILC3) are potent effector cells with critical roles in enforcing immunity, barrier integrity and tissue homeostasis along the gastrointestinal tract. ILC3 are considered primarily tissue-resident cells, seeding the gastrointestinal tract during embryonic stages and early life. However, the mechanisms through which mature ILC3 are maintained within adult tissues are poorly understood. Here, we report that lymphoid tissue-inducer-like (LTi-like) ILC3 exhibit minimal turnover in the healthy adult intestinal tract, persist for extended periods of time, and display a quiescent phenotype. Strikingly, during enteric bacterial infection LTi-like ILC3 also exhibit negligible hematopoietic replenishment and remain non-proliferative, despite robustly producing cytokines. Survival of LTi-like ILC3 was found to be dependent upon the balance between the metabolic activity required to drive effector function and anti-apoptotic programs. Notably, the pro-survival protein B-cell lymphoma-2 (Bcl-2) was required for the survival of LTi-like ILC3 ex vivo but was rendered partially dispensable if mitochondrial respiration was inhibited. Together we demonstrate LTi-like ILC3 are a tissue-resident, quiescent population that persist independently of hematopoietic replenishment to survive within the intestinal microenvironment.

This study investigates sex-associated systemic innate immune differences by examining bone marrow-derived dendritic cells (BMDCs). BMDC grown from 7-day-old mice show enhanced type-I interferon (IFN) signaling in female compared to male BMDC. Upon respiratory syncytial virus (RSV) infection of 7-day-old mice, a significantly altered phenotype of BMDC at 4 weeks post-infection is observed in a sex-dependent manner. The alterations include heightened Ifnb/ interleukin (Il12a) and enhanced IFNAR1+ expression in BMDC from early-life RSV-infected female mice that leads to increased IFN-γ production by T cells. Phenotypic differences were verified upon pulmonary sensitization whereby EL-RSV male-derived BMDC promoted enhanced T helper 2/17 responses and exacerbated disease upon RSV infection while EL-RSV/F BMDC sensitization was relatively protective. Assay for transposase-accessible chromatin using sequencing analysis (ATAC-seq) demonstrated that EL-RSV/F BMDC had enhanced chromatin accessibility near type-I immune genes with JUN, STAT1/2, and IRF1/8 transcription factors predicted to have binding sites in accessible regions. Importantly, ATAC-seq of human cord blood-derived monocytes displayed a similar sex-associated chromatin landscape with female-derived monocytes having more accessibility in type-I immune genes. These studies enhance our understanding of sex-associated differences in innate immunity by epigenetically controlled transcriptional programs amplified by early-life infection in females via type-I immunity.

Streptococcus pneumoniae is the most common etiology of bacterial pneumonia, one of the leading causes of death in children and the elderly worldwide. During non-lethal infections with S. pneumoniae, lymphocytes accumulate in the lungs and protect against reinfection with serotype-mismatched strains. Cluster of differentiation CD4+ resident memory T (TRM) cells are known to be crucial for this protection, but the diversity of lung CD4+ TRM cells has yet to be fully delineated. We aimed to identify unique subsets and their contributions to lung immunity. After recovery from pneumococcal infections, we identified a distinct subset of CD4+ T cells defined by the phenotype CD11ahiCD69+GL7+ in mouse lungs. Phenotypic analyses for markers of lymphocyte memory and residence demonstrated that GL7+ T cells are a subset of CD4+ TRM cells. Functional studies revealed that unlike GL7- TRM subsets that were mostly (RAR-related Orphan Receptor gamma T) RORγT+, GL7+ TRM cells exhibited higher levels of (T-box expressed in T cells) T-bet and Gata-3, corresponding with increased synthesis of interferon-γ, interleukin-13, and interleukin-5, inherent to both T helper 1 (TH1) and TH2 functions. Thus, we propose that these cells provide novel contributions during pneumococcal pneumonia, serving as important determinants of lung immunity.

Air pollution significantly impacts the aggravation of asthma. Exposure to acrylamide, a volatile organic compound in tobacco smoke, is associated with elevated risks of allergy-related outcomes among active smokers. As group 2 innate lymphoid cells (ILC2s) can act as an environmental sensor and significantly contribute to protease allergen-induced lung inflammation, we aimed to elucidate the causal relationship and how inhaled acrylamide worsens allergic lung inflammation via ILC2s. Intranasal acrylamide exposure at nanomolar levels significantly enhanced allergen-induced or rmIL-33-induced lung inflammation in C57BL/6 mice or Rag1-/- mice, respectively. The cardinal features of lung inflammation included accumulated infiltration of ILC2s and eosinophils. Transcriptomic analysis revealed a gene expression pattern associated with proliferation-related pathways in acrylamide-treated ILC2s. Western blotting revealed significantly higher expression of Ras and p-Erk in acrylamide-treated ILC2s than the control, suggesting Ras-Erk signaling pathway involvement. Ex vivo and in vitro analysis showed that acrylamide treatment mainly increased Ki-67+ ILC2s and the cell number of ILC2s whereas PD98059, a highly selective Erk inhibitor, effectively counteracted the acrylamide effect. Intratracheal administration of acrylamide-treated ILC2s significantly enhanced eosinophil infiltration in Rag1-/- mice. This study suggests that airborne acrylamide may enhance the severity of allergen-induced airway eosinophilic inflammation, partly via altering ILC2 proliferative activity.

In the intestine, mucin 2 (Muc2) forms a network structure and prevents bacterial invasion. Glycans are indispensable for Muc2 barrier function. Among various glycosylation patterns of Muc2, sialylation inhibits bacteria-dependent Muc2 degradation. However, the mechanisms by which Muc2 creates the network structure and sialylation prevents mucin degradation remain unknown. Here, by focusing on two glycosyltransferases, St6 N-acetylgalactosaminide α-2,6-sialyltransferase 6 (St6galnac6) and β-1,3-galactosyltransferase 5 (B3galt5), mediating the generation of desialylated glycans, we show that sialylation forms the network structure of Muc2 by providing negative charge and hydrophilicity. The colonic mucus of mice lacking St6galnac6 and B3galt5 was less sialylated, thinner, and more permeable to microbiota, resulting in high susceptibility to intestinal inflammation. Mice with a B3galt5 mutation associated with inflammatory bowel disease (IBD) also showed the loss of desialylated glycans of mucus and the high susceptibility to intestinal inflammation, suggesting that the reduced sialylation of Muc2 is associated with the pathogenesis of IBD. In mucins of mice with reduced sialylation, negative charge was reduced, the network structure was disturbed, and many bacteria invaded. Thus, sialylation mediates the negative charging of Muc2 and facilitates the formation of the mucin network structure, thereby inhibiting bacterial invasion in the colon to maintain gut homeostasis.