Innate lymphoid cells (ILC) distribution and compartmentalization in human lymphoid tissues are incompletely described. Through combined multiplex immunofluorescence, multispectral imaging, and advanced computer vision methods, we provide a map of ILCs at the whole-slide single-cell resolution level, and study their proximity to T helper (Th) cells. The results show that ILC2 predominates in thymic medulla; by contrast, immature Th cells prevail in the cortex. Unexpectedly, we find that Th2-like and Th17-like phenotypes appear before complete T cell receptor gene rearrangements in these immature thymocytes. In the periphery, ILC2 are more abundant in lymph nodes and tonsils, penetrating lymphoid follicles. NK cells are uncommon in lymphoid tissues but abundant in the spleen, whereas ILC1 and ILC3 predominate in the ileum and appendix. Under pathogenic conditions, a deep perturbation of both ILC and Th populations is seen in follicular lymphoma compared with non-neoplastic conditions. Lastly, all ILCs are preferentially in close proximity to their Th counterparts. In summary, our histopathology tool help present a spatial mapping of human ILCs and Th cells, in normal and neoplastic lymphoid tissues.

Type 3 Innate lymphoid cells (ILC3s) play a crucial role in intestinal immune function by serving as an innate effector that contributes to early-life defense against pathogens and helps protect the intestines from bacterial infections. ILC3s exert their immune function through cytokine secretion, patrolling actions and the generation of memory ILC3s that aid in repairing epithelial tissue and preserving mucosal barrier integrity. Moreover, dysregulation of ILC3s function has been implicated in the pathogenesis and progression of autoimmune diseases. This comprehensive review aims to explore the interactions between gut microbes, gut microbial metabolites, and diet in relation to ILC3s within the context of the gut microenvironment. Furthermore, the gut microenvironment has the potential to influence distant extra-intestinal sites through immunomodulation, thereby modifying their risk of inflammation. The gut has emerged as a significant focus of autoimmune disease research in recent years. However, the relationship between gut ILC3s and autoimmune diseases remains poorly understood. This paper aims to examine the potential association between ILC3s and autoimmune diseases.

Innate lymphoid cells (ILCs) have emerged as pivotal players in the field of immunology, expanding our understanding of innate immunity beyond conventional paradigms. This comprehensive review delves into the multifaceted world of ILCs, beginning with their serendipitous discovery and traversing their ontogeny and heterogeneity. We explore the distinct subsets of ILCs unraveling their intriguing plasticity, which adds a layer of complexity to their functional repertoire. As we journey through the functional activities of ILCs, we address their role in immune responses against various infections, categorizing their interactions with helminthic parasites, bacterial pathogens, fungal infections, and viral invaders. Notably, this review offers a detailed examination of ILCs in the context of specific infections, such as Mycobacterium tuberculosis, Citrobacter rodentium, Clostridium difficile, Salmonella typhimurium, Helicobacter pylori, Listeria monocytogenes, Staphylococcus aureus, Pseudomonas aeruginosa, Influenza virus, Cytomegalovirus, Herpes simplex virus, and severe acute respiratory syndrome coronavirus 2. This selection aimed for a comprehensive exploration of ILCs in various infectious contexts, opting for microorganisms based on extensive research findings rather than considerations of virulence or emergence. Furthermore, we raise intriguing questions about the potential immune functional resemblances between ILCs and epithelial cells, shedding light on their interconnectedness within the mucosal microenvironment. The review culminates in a critical assessment of the therapeutic prospects of targeting ILCs during infection, emphasizing their promise as novel immunotherapeutic targets. Nevertheless, due to their recent discovery and evolving understanding, effectively manipulating ILCs is challenging. Ensuring specificity and safety while evaluating long-term effects in clinical settings will be crucial.

External signals from the thymic microenvironment and the activities of lineage-specific transcription factors (TFs) instruct T-cell versus innate lymphoid cell (ILC) fates. However, mechanistic insights into how factors such as Notch1-Delta-like-4 (Dll4) signaling and E-protein TFs collaborate to establish T-cell identity remain rudimentary. Using multiple in vivo approaches and single-cell multiome analysis, we identified a feedback amplifier circuit that specifies fetal and adult T-cell fates. In early T progenitors (ETPs) in the fetal thymus, Notch signaling minimally lowered E-protein antagonist Id2 levels, and high Id2 abundance favored the differentiation of ETPs into ILCs. Conversely, in the adult thymus, Notch signaling markedly decreased Id2 abundance in ETPs, substantially elevating E-protein DNA binding and in turn promoting the activation of a T-cell lineage-specific gene expression program linked with V(D)J gene recombination and T-cell receptor signaling. Our findings indicate that, in the fetal versus the adult thymus, a simple feedback amplifier circuit dictated by Notch-mediated signals and Id2 abundance enforces T-cell identity and suppresses ILC development.

Asthma is a common chronic inflammatory disease of the airways. A substantial number of patients present with severe and therapy-resistant asthma, for which the underlying biological mechanisms remain poorly understood. In most asthma patients, airway inflammation is characterized by chronic activation of type 2 immunity. CD4+ T helper 2 (Th2) cells are the canonical producers of the cytokines that fuel type 2 inflammation: interleukin (IL)-4, IL-5, IL-9, and IL-13. However, more recent findings have shown that other lymphocyte subsets, in particular group 2 innate lymphoid cells (ILC2s) and type 2 CD8+ cytotoxic T (Tc2) cells, can also produce large amounts of type 2 cytokines. Importantly, a substantial number of severe therapy-resistant asthma patients present with chronic type 2 inflammation, despite the high sensitivity of Th2 cells for suppression by corticosteroids-the mainstay drugs for asthma. Emerging evidence indicates that ILC2s and Tc2 cells are more abundant in severe asthma patients and can adopt corticosteroid-resistance states. Moreover, many severe asthma patients do not present with overt type 2 airway inflammation, implicating non-type 2 immunity as a driver of disease. In this review, we will discuss asthma pathophysiology and focus on the roles played by ILC2s, Tc2 cells, and non-type 2 lymphocytes, placing special emphasis on severe disease forms.

The helper-like ILC contains various functional subsets, such as ILC1, ILC2, ILC3 and LTi cells, mediating the immune responses against viruses, parasites, and extracellular bacteria, respectively. Among them, LTi cells are also crucial for the formation of peripheral lymphoid tissues, such as lymph nodes. Our research, along with others', indicates a high proportion of LTi cells in the fetal ILC pool, which significantly decreases after birth. Conversely, the proportion of non-LTi ILCs increases postnatally, corresponding to the need for LTi cells to mediate lymphoid tissue formation during fetal stages and other ILC subsets to combat diverse pathogen infections postnatally. However, the regulatory mechanism for this transition remains unclear. In this study, we observed a preference for fetal ILC progenitors to differentiate into LTi cells, while postnatal bone marrow ILC progenitors preferentially differentiate into non-LTi ILCs. Particularly, this differentiation shift occurs within the first week after birth in mice. Further analysis revealed that adult ILC progenitors exhibit stronger activation of the Notch signaling pathway compared to fetal counterparts, accompanied by elevated Gata3 expression and decreased Rorc expression, leading to a transition from fetal LTi cell-dominant states to adult non-LTi ILC-dominant states. This study suggests that the body can regulate ILC development by modulating the activation level of the Notch signaling pathway, thereby acquiring different ILC subsets to accommodate the varying demands within the body at different developmental stages.

Respiratory failure occurs more frequently after thoracic surgery than abdominal surgery. Although the etiology for this complication is frequently attributed to underlying lung disease present in patients undergoing thoracic surgery, this notion is often unfounded because many patients with normal preoperative pulmonary function often require prolonged oxygen supplementation even after minimal resection of lung tissue. Using a murine model of pulmonary resection and peripheral blood samples from patients undergoing resection of the lung or abdominal organs, we demonstrated that lung surgery initiates a proinflammatory loop that results in damage to the remaining lung tissue, noncardiogenic pulmonary edema, hypoxia, and even death. Specifically, we demonstrated that resection of murine lung tissue increased concentrations of the homeostatic cytokine interleukin-7, which led to local and systemic activation of type 2 innate lymphoid cells. This process activated lung-resident eosinophils and facilitated stress-induced eosinophil maturation in the bone marrow in a granulocyte-macrophage colony-stimulating factor-dependent manner, resulting in systemic eosinophilia in both mice and humans. Up-regulation of inducible nitric oxide synthase in lung-resident eosinophils led to tissue nitrosylation, pulmonary edema, hypoxia, and, at times, death. Disrupting this activation cascade at any stage ameliorated deleterious outcomes and improved survival after lung resection in the mouse model. Our data suggest that repurposing US Food and Drug Administration-approved eosinophil-targeting strategies may potentially offer a therapeutic intervention to improve outcomes for patients who require lung resection for benign or malignant etiology.

In squamous cell carcinoma (SCC), macrophages responding to interleukin (IL)-33 create a TGF-β-rich stromal niche that maintains cancer stem cells (CSCs), which evade chemotherapy-induced apoptosis in part via activation of the NRF2 antioxidant program. Here, we examined how IL-33 derived from CSCs facilitates the development of an immunosuppressive microenvironment. CSCs with high NRF2 activity redistributed nuclear IL-33 to the cytoplasm and released IL-33 as cargo of large oncosomes (LOs). Mechanistically, NRF2 increased the expression of the lipid scramblase ATG9B, which exposed an "eat me" signal on the LO surface, leading to annexin A1 (ANXA1) loading. These LOs promoted the differentiation of AXNA1 receptor+ myeloid precursors into immunosuppressive macrophages. Blocking ATG9B's scramblase activity or depleting ANXA1 decreased niche macrophages and hindered tumor progression. Thus, IL-33 is released from live CSCs via LOs to promote the differentiation of alternatively activated macrophage, with potential relevance to other settings of inflammation and tissue repair.

In mice, skin-resident type 2 innate lymphoid cells (ILC2s) exhibit some ILC3-like characteristics. However, the underlying mechanism remains elusive. Here, we observed lower expression of the ILC2 master regulator GATA3 specifically in cutaneous ILC2s (cILC2s) compared with canonical ILC2s, in line with its functionally divergent role in transcriptional control in cILC2s. Decreased levels of GATA3 enabled the expansion of RORγt fate-mapped (RORγtfm+) cILC2s after postnatal days, displaying certain similarities to ILC3s. Single-cell trajectory analysis showed a sequential promotion of the RORγtfm+ cILC2 divergency by RORγt and GATA3. Notably, during hair follicle recycling, these RORγtfm+ cILC2s accumulated around the hair follicle dermal papilla (DP) region to facilitate the process. Mechanistically, we found that GATA3-mediated integrin α3β1 upregulation on RORγtfm+ cILC2s was required for their positioning around the DP. Overall, our study demonstrates a distinct regulatory role of GATA3 in cILC2s, particularly in promoting the divergence of RORγtfm+ cILC2s to facilitate hair follicle recycling.

Group 2 innate lymphoid cells (ILC2s) are a subset of innate lymphocytes that produce type 2 cytokines, including IL-4, IL-5, and IL-13. GATA3 is a critical transcription factor for ILC2 development at multiple stages. However, when and how GATA3 is induced to the levels required for ILC2 development remains unclear. Herein, we identify ILC2-specific GATA3-related tandem super-enhancers (G3SE) that induce high GATA3 in ILC2-committed precursors. G3SE-deficient mice exhibit ILC2 deficiency in the bone marrow, lung, liver, and small intestine with minimal impact on other ILC lineages or Th2 cells. Single-cell RNA-sequencing and subsequent flow cytometry analysis show that GATA3 induction mechanism, which is required for entering the ILC2 stage, is lost in IL-17RB+PD-1- late ILC2-committed precursor stage in G3SE-deficient mice. Cnot6l, part of the CCR4-NOT deadenylase complex, is a possible GATA3 target during ILC2 development. Our findings implicate a stage-specific regulatory mechanism for GATA3 expression during ILC2 development.

Natural killer (NK) cells function by eliminating virus-infected or tumor cells. Here we identified an NK-lineage-biased progenitor population, referred to as early NK progenitors (ENKPs), which developed into NK cells independently of common precursors for innate lymphoid cells (ILCPs). ENKP-derived NK cells (ENKP_NK cells) and ILCP-derived NK cells (ILCP_NK cells) were transcriptionally different. We devised combinations of surface markers that identified highly enriched ENKP_NK and ILCP_NK cell populations in wild-type mice. Furthermore, Ly49H+ NK cells that responded to mouse cytomegalovirus infection primarily developed from ENKPs, whereas ILCP_NK cells were better IFNγ producers after infection with Salmonella and herpes simplex virus. Human CD56dim and CD56bright NK cells were transcriptionally similar to ENKP_NK cells and ILCP_NK cells, respectively. Our findings establish the existence of two pathways of NK cell development that generate functionally distinct NK cell subsets in mice and further suggest these pathways may be conserved in humans.

Innate lymphoid cells (ILCs) and adaptive T lymphocytes promote tissue homeostasis and protective immune responses. Their production depends on the transcription factor GATA3, which is further elevated specifically in ILC2s and T helper 2 cells to drive type-2 immunity during tissue repair, allergic disorders, and anti-helminth immunity. The control of this crucial up-regulation is poorly understood. Using CRISPR screens in ILCs we identified previously unappreciated myocyte-specific enhancer factor 2d (Mef2d)-mediated regulation of GATA3-dependent type-2 lymphocyte differentiation. Mef2d-deletion from ILC2s and/or T cells specifically protected against an allergen lung challenge. Mef2d repressed Regnase-1 endonuclease expression to enhance IL-33 receptor production and IL-33 signaling and acted downstream of calcium-mediated signaling to translocate NFAT1 to the nucleus to promote type-2 cytokine-mediated immunity.

While the transcription factor GATA-3 is well-established for its crucial role in T cell development, its specific influence on invariant natural killer T (iNKT) cells remains relatively unexplored. Using flow cytometry and single-cell transcriptomic analysis, we demonstrated that GATA-3 deficiency in mice leads to the absence of iNKT2 and iNKT17 cell subsets, as well as an altered distribution of iNKT1 cells. Thymic iNKT cells lacking GATA-3 exhibited diminished expression of PLZF and T-bet, key transcription factors involved in iNKT cell differentiation, and reduced production of Th2, Th17, and cytotoxic effector molecules. Single-cell transcriptomics revealed a comprehensive absence of iNKT17 cells, a substantial reduction in iNKT2 cells, and an increase in iNKT1 cells in GATA-3-deficient thymi. Differential expression analysis highlighted the regulatory role of GATA-3 in T cell activation signaling and altered expression of genes critical for iNKT cell differentiation, such as Icos, Cd127, Eomes, and Zbtb16. Notably, restoration of Icos, but not Cd127, expression could rescue iNKT cell development in GATA-3-deficient mice. In conclusion, our study demonstrates the pivotal role of GATA-3 in orchestrating iNKT cell effector lineage differentiation through the regulation of T cell activation pathways and Icos expression, providing insights into the molecular mechanisms governing iNKT cell development and function.

Epithelium-derived cytokines or alarmins, such as interleukin-33 (IL-33) and thymic stromal lymphopoietin (TSLP), are major players in type 2 immunity and asthma. Here, we demonstrate that TNF-like ligand 1A (TL1A) is an epithelial alarmin, constitutively expressed in alveolar epithelium at steady state in both mice and humans, which cooperates with IL-33 for early induction of IL-9high ILC2s during the initiation of allergic airway inflammation. Upon synergistic activation by IL-33 and TL1A, lung ILC2s acquire a transient IL-9highGATA3low "ILC9" phenotype and produce prodigious amounts of IL-9. A combination of large-scale proteomic analyses, lung intravital microscopy, and adoptive transfer of ILC9 cells revealed that high IL-9 expression distinguishes a multicytokine-producing state-of-activated ILC2s with an increased capacity to initiate IL-5-dependent allergic airway inflammation. Similar to IL-33 and TSLP, TL1A is expressed in airway basal cells in healthy and asthmatic human lungs. Together, these results indicate that TL1A is an epithelium-derived cytokine and an important cofactor of IL-33 in the airways.

IL-33 is a danger signal that binds to its receptor ST2L to promote tumor progression. This study identifies the IL-33/ST2L positive-feedback loop and the trafficking of ST2L membrane presentation in macrophages that contribute to lung tumor progression. Mechanistically, IL-33 induces ST2L upregulation by activating NF-κB, which binds to the promoter region of the ST2L gene. Moreover, Rab37, a small GTPase involved in membrane trafficking, mediates ST2L trafficking to the plasma membrane of M2 macrophages. This IL-33/NF-κB/ST2L/Rab37 axis promotes positive-feedback loops that enhance ST2L expression and membrane trafficking in M2 macrophages. Notably, neutralizing antibodies against IL-33 or ST2L block NF-κB activity, suppress M2 macrophage polarization, and synergistically inhibit tumor growth when combined with cisplatin treatment in vitro/vivo. Clinically, Rab37+/ST2L+/CD206+ tumor-infiltrating M2 macrophages correlate with advanced-stage lung cancer patients with poor response to chemotherapy. These findings unveil a positive-feedback mechanism and provide a basis for IL-33/ST2L-targeting therapy for cancer.

Group 3 innate lymphoid cells (ILC3) are the major subset of gut-resident ILC with essential roles in infections and tissue repair, but how they adapt to the gut environment to maintain tissue residency is unclear. We report that Tox2 is critical for gut ILC3 maintenance and function. Gut ILC3 highly expressed Tox2, and depletion of Tox2 markedly decreased ILC3 in gut but not at central sites, resulting in defective control of Citrobacter rodentium infection. Single-cell transcriptional profiling revealed decreased expression of Hexokinase-2 in Tox2-deficient gut ILC3. Consistent with the requirement for hexokinases in glycolysis, Tox2-/- ILC3 displayed decreased ability to utilize glycolysis for protein translation. Ectopic expression of Hexokinase-2 rescued Tox2-/- gut ILC3 defects. Hypoxia and interleukin (IL)-17A each induced Tox2 expression in ILC3, suggesting a mechanism by which ILC3 adjusts to fluctuating environments by programming glycolytic metabolism. Our results reveal the requirement for Tox2 to support the metabolic adaptation of ILC3 within the gastrointestinal tract.

Evolution has created complex mechanisms to sense environmental danger and protect tissues, with the nervous and immune systems playing pivotal roles. These systems work together, coordinating local and systemic reflexes to restore homeostasis in response to tissue injury and infection. By sharing receptors and ligands, they influence the pathogenesis of various diseases. Recently, a less-explored aspect of neuroimmune communication has emerged: the release of neuropeptides from immune cells and cytokines/chemokines from sensory neurons. This article reviews evidence of this unique neuroimmune interplay and its impact on the development of allergy, inflammation, itch, and pain. We highlight the effects of this neuroimmune signaling on vital processes such as host defense, tissue repair, and inflammation resolution, providing avenues for exploration of the underlying mechanisms and therapeutic potential of this signaling.

Diffuse large B-cell lymphoma (DLBCL) exhibits remarkable heterogeneity but still remains undiagnosed in identifying the subpopulation of DLBCL to predict the prognosis and guide clinical treatment.

Molecular subgroups were identified in gene expression data from GSE10846 by a consensus clustering algorithm. And gene set enrichment analysis, immune infiltration, and the proposed cell cycle algorithm were applied to explore the biological functions of different subtypes. Meanwhile, univariate and multivariate Cox regression analyses were used to evaluate independent prognostic factors of DLBCL. Finally, the prognostic model, including some key genes screened by Lasso regression, Random Forest algorithm, and point-biserial correlation, was constructed by an optimal classifier from seven machine learning algorithms and validated by another three external datasets (GSE34171, GSE87371, GSE31312).

Comprehensive genomic analysis of 1,143 DLBCL samples identify 2 molecularly, prognostically relevant subtypes: immune-enriched (IME) and cell-cycle-enriched (CCE). Then a new predictive model including seven key genes (SERPING1, TIMP2, NME1, DCTPP1, RFC4, POLE2, and SNRPD1) was developed with high prediction accuracy (88.6%) and strong predictive power (AUC = 0.973) based on the Support Vector Machine (SVM) algorithm in 414 patients from GSE10846. The predictive power was similar in another three testing sets (HR > 1.400, p < 0.05).

This model could evaluate survival independently with strong predictive power compared with other clinical risk factors. Our study constructed a reliable model to predict two new subtypes of DLBCL patients, which could guide the implementation of individualized treatment.

Dibutyl phthalate (DBP), a commonly used plasticizer, has been found to be strongly linked to a consistently high prevalence of allergic diseases, particularly allergic asthma. Previous animal experiments have demonstrated that exposure to DBP can worsen asthma by triggering the production of calcitonin gene-related peptide (CGRP), a neuropeptide in the lung tissue. However, the precise neuroimmune mechanism and pathophysiology of DBP-exacerbated allergic asthma with the assistance of CGRP remain unclear.

The present study was to investigate the potential pathophysiological mechanism in DBP-exacerbated asthma from the perspective of neural-immune interactions.

C57BL/6 mice were orally exposed to different concentrations (0.4, 4, 40 mg/kg) of DBP for 28 days. They were then sensitized with OVA and nebulized with OVA for 7 consecutive excitations. To investigate whether DBP exacerbates allergic asthma in OVA induced mice, we analyzed airway hyperresponsiveness and lung histopathology. To investigate the activation of JNC and TRPV1 neurons and the release of CGRP by JNC cells, we measured the levels of TRPV1 channels, calcium inward flow, and downstream neuropeptide CGRP. Results showed that TRPV1 expression, inward calcium flux, and CGRP levels were significantly elevated in the lung tissues of the 40DBP + OVA group, suggesting the release of CGRP by JNC cells. To counteract the detrimental effects of DBP mediated by CGRP, we employed olcegepant (also known as BIBN-4096), a CGRP receptor specific antagonist. Results revealed that 40DBP + OVA + olcegepant led to notable decreases in TRPV1, calcium inward flow, and CGRP expression in lung tissues compare with 40DBP + OVA, further supporting the efficacy of olcegepant. Additionally, we also conducted ILC2 flow sorting and observed that neuropeptide CGRP-activated ILC2 cells have a crucial role as key effector cells in DBP-induced neuroimmune positive feedback regulation. Finally, we examined the protein expression of CGRP, GATA3 and P-GATA3, and found that significant upregulations of CGRP and P-GATA3 in the 40DBP + OVA group, suggest that GATA3 acted as a key regulator of CGRP-activated ILC2.

The aforementioned studies indicate that exposure to DBP can exacerbate allergic asthma, leading to airway inflammation. This exacerbation occurs through the activation of TRPV1 in JNC, resulting in the release of CGRP. The excessive release of CGRP further promotes the release of Th2 cytokines by inducing the activation of ILC2 through GATA phosphorylation. Consequently, this process contributes to the development of airway inflammation and allergic asthma. The increased production of Th2 cytokines also triggers the production of IgE, which interacts with FcεRI on JNC neurons, thereby mediating neuro-immune positive feedback regulation.

Innate lymphoid cells (ILCs) play an important role in maintaining tissue homeostasis and various inflammatory responses. ILCs are typically classified into three subsets, as is the case for T-cells. Recent studies have reported that IL-10-producing type 2 ILCs (ILC210s) have an immunoregulatory function dependent on IL-10. However, the surface markers of ILC210s and the role of ILC210s in contact hypersensitivity (CHS) are largely unknown. Our study revealed that splenic ILC210s are extensively included in PD-L1highSca-1+ ILCs and that IL-27 amplifies the development of PD-L1highSca-1+ ILCs and ILC210s. Adoptive transfer of PD-L1highSca-1+ ILCs suppressed oxazolone-induced CHS in an IL-10-dependent manner Taken together, our results demonstrate that ILC210s are critical for the control of CHS and suggest that ILC210s can be used as target cells for the treatment of CHS.

It is assumed that cells corresponding to innate lymphoid cells (ILCs) in humans, in addition to lymphoid tissue inducer cells (LTi), are also found in teleosts. In this systematic group of organisms, however, they are a poorly understood cell population. In contrast to the data on ILCs in humans, which also remain incomplete despite advanced research, in teleosts, these cells require much more attention. ILCs in teleosts have been presented as cells that may be evolutionary precursors of NK cells or ILCs identified in mammals, including humans. It is a highly heterogeneous group of cells in both humans and fish and their properties, as revealed by studies in humans, are most likely to remain strictly dependent on the location of these cells and the physiological state of the individual from which they originate. They form a bridge between innate and adaptive immunity. The premise of this paper is to review the current knowledge of ILCs in teleosts, taking into account data on similar cells in humans. A review of the knowledge concerning these particular cells, elements of innate immunity mechanisms as equivalent to, or perhaps dominant over, adaptive immunity mechanisms in teleosts, as presented, may inspire the need for further research.

Intestinal tuft cells (TCs) are defined as chemosensory cells that can "taste" danger and induce immune responses. They play a critical role in gastrointestinal parasite invasion, inflammatory bowel diseases and high-fat diet-induced obesity. Intestinal IL-25, the unique product of TCs, is a key activator of type 2 immunity, especially to promote group 2 innate lymphoid cells (ILC2s) to secret IL-13. Then the IL-13 mainly promotes intestinal stem cell (ISCs) proliferation into TCs and goblet cells. This pathway formulates the circuit in the intestine. This paper focuses on the potential role of the intestinal TC, ILC2 and their circuit in obesity-induced intestinal damage, and discussion on further study and the potential therapeutic target in obesity.

Innate lymphoid cells (ILCs), as the innate counterpart of CD4+ T helper (Th) cells, play crucial roles in maintaining tissue homeostasis. While the ILC subsets and their corresponding Th subsets demonstrate significant similarities in core programming related to effector function and regulatory mechanisms, their principal distinctions, given their innate and adaptive lymphocyte nature, remain largely unknown. In this study, we have employed an integrative analysis of 294 bulk RNA-sequencing results across all ILC and Th subsets, using scRNA-seq algorithms. Consequently, we identify two genesets that predominantly differentiate ILCs from Th cells, as well as three genesets that distinguish various immune responses. Furthermore, through chromatin accessibility analysis, we find that the ILC geneset tends to rely on specific transcriptional regulation at promoter regions compared with the Th geneset. Additionally, we observe that ILCs and Th cells are under differential transcriptional regulation. For example, ILCs are under stronger regulation by multiple transcription factors, including RORα, GATA3, and NF-κB. Otherwise, Th cells are under stronger regulation by AP-1. Thus, our findings suggest that, despite the acknowledged similarities in effector functions between ILC subsets and corresponding Th subsets, the underlying regulatory machineries still exhibit substantial distinctions. These insights provide a comprehensive understanding of the unique roles played by each cell type during immune responses.

Group 2 innate lymphoid cells (ILC2s) are critical sources of type 2 cytokines and represent one of the major tissue-resident lymphoid cells in the mouse lung. However, the molecular mechanisms underlying ILC2 activation under challenges are not fully understood.

Here, using single-cell transcriptomics, genetic reporters, and gene knockouts, we identify four ILC2 subsets, including two non-activation subsets and two activation subsets, in the mouse acute inflammatory lung. Of note, a distinct activation subset, marked by the transcription factor Nr4a1, paradoxically expresses both tissue-resident memory T cell (Trm), and effector/central memory T cell (Tem/Tcm) signature genes, as well as higher scores of proliferation, activation, and wound healing, all driven by its particular regulons. Furthermore, we demonstrate that the Nr4a1+ILC2s are restrained from activating by the programmed cell death protein-1 (PD-1), which negatively modulates their activation-related regulons. PD-1 deficiency places the non-activation ILC2s in a state that is prone to activation, resulting in Nr4a1+ILC2 differentiation through different activation trajectories. Loss of PD-1 also leads to the expansion of Nr4a1+ILC2s by the increase of their proliferation ability.

The findings show that activated ILC2s are a heterogenous population encompassing distinct subsets that have different propensities, and therefore provide an opportunity to explore PD-1's role in modulating the activity of ILC2s for disease prevention and therapy.

Asthma is a chronic airway inflammatory disease whose global incidence increases annually. The role of innate lymphoid cells (ILCs) is a crucial aspect of asthma research with respect to different endotypes of asthma. Based on its pathological and inflammatory features, asthma is divided into type 2 high and type 2 low endotypes. Type-2 high asthma is distinguished by the activation of type 2 immune cells, including T helper 2 (Th2) cells and ILC2s; the production of cytokines interleukin (IL)-4, IL-5 and IL-13; eosinophilic aggregation; and bronchial hyper-responsiveness. Type-2 low asthma represents a variety of endotypes other than type 2 high endotype such as the IL-1β/ILC3/neutrophil endotype and a paucigranulocytic asthma, which may be insensitive to corticosteroid treatment and/or associated with obesity. The complexity of asthma is due to the involvement of multiple cell types, including tissue-resident ILCs and other innate immune cells including bronchial epithelial cells, dendritic cells, macrophages and eosinophils, which provide immediate defence against viruses, pathogens and allergens. On this basis, innate immune cells and adaptive immune cells combine to induce the pathological condition of asthma. In addition, the plasticity of ILCs increases the heterogeneity of asthma. This review focuses on the phenotypes of tissue-resident ILCs and their roles in the different endotypes of asthma, as well as the mechanisms of tissue-resident ILCs and other immune cells. Based on the phenotypes, roles and mechanisms of immune cells, the therapeutic strategies for asthma are reviewed.

Lymphoid tissue inducer (LTi) cells, a subset of innate lymphoid cells (ILCs), play an essential role in the formation of secondary lymphoid tissues. However, the regulation of the development and functions of this ILC subset is still elusive. In this study, we report that the transcription factor T cell factor 1 (TCF-1), just as GATA3, is indispensable for the development of non-LTi ILC subsets. While LTi cells are still present in TCF-1-deficient mice, the organogenesis of Peyer's patches (PPs), but not of lymph nodes, is impaired in these mice. LTi cells from different tissues have distinct gene expression patterns, and TCF-1 regulates the expression of lymphotoxin specifically in PP LTi cells. Mechanistically, TCF-1 may directly and/or indirectly regulate Lta, including through promoting the expression of GATA3. Thus, the TCF-1-GATA3 axis, which plays an important role during T cell development, also critically regulates the development of non-LTi cells and tissue-specific functions of LTi cells.

Vitamin D3 (VitD3) is known to have immunomodulatory functions, and VitD3 deficiency is associated with more severe asthma.

We aimed to assess the immunoregulatory effects of VitD3 food supplementation on asthma manifestation, with particular focus on T cells and type 2 innate lymphoid cells.

Preschool children and adult asthmatic cohorts were analyzed in the context of VitD3 supplementation and serum levels. In a murine model of ovalbumin-induced asthma, effects of diet VitD3 sufficiency and deficiency on T cells and type 2 innate lymphoid cells immune mechanisms were investigated.

We found less severe and better-controlled asthma phenotypes along with reduced need for steroid medication in preschool children and asthmatic adults with VitD3 supplementation. VitD3 serum levels correlated with B lymphocyte-induced maturation protein 1 (Blimp-1) expression in blood peripheral mononuclear cells. VitD3-supplement-fed mice showed decreased asthmatic traits, with a decrease in IgE serum levels, reduced airway mucus, and increased IL-10 production by lung cells. Furthermore, we discovered an upregulation of effector T cells and Blimp-1+ lung tissue-resident memory T cells as well as induction of anti-inflammatory Blimp-1+ lung innate lymphoid cells producing IL-10.

Supplementing VitD3 resulted in amelioration of clinical asthma manifestations in human studies as well as in experimental allergic asthma, indicating that VitD3 shifts proinflammatory immune responses to anti-inflammatory immune responses via upregulating Blimp-1 in lung innate lymphoid cells and tissue-resident memory cells.

ILC2s are capable of generating memory. The mechanism of memory induction and memory-driven effector function (trained immunity) in ILC2s is unknown.

NFκB1 is preferentially expressed at a high level in ILC2s. We examined the role of NFkB1 in memory induction and memory-driven effector function in a mouse model of asthma.

Intranasal administration of Alternaria, flexivent, ELISA, histology, real-time PCR, western blot, flow cytometry and immunofluorescence staining.

NFκB1 was essential for the effector phase of memory-driven asthma. NFκB1 was critical for IL33 production, ILC2 generation, and production of type-2 cytokines, which resulted in eosinophilic inflammation and other features of asthma. NFκB1 induction of type-2 cytokines in ILC2s was independent of GATA3. NFκB1 was important for allergen induction of ILC3s and FoxP3+ Tregs. NFκB1 did not affect Th2 cells or their cytokine production. In contrast to its protagonistic role in the effector phase, NFκB1 had an antagonistic role in the memory phase. NFκB1 inhibited allergen-induced upregulation of memory-associated repressor and preparedness genes in ILC2s. NFκB1 upregulated RUNX1. NFκB1 formed a heterodimer with RUNX1 in ILC2s.

NFκB1 positively regulated the effector phase but inhibited the induction phase of memory. The foregoing pointed to an interdependent antagonism between the memory induction and the memory effector processes. The NFκB1-RUNX1 heterodimer represented a non-canonical transcriptional activator of type-2 cytokines in ILC2s.

T-bet-expressing Th17 (T-bet+RORγt+) cells are associated with the induction of pathology during experimental autoimmune encephalomyelitis (EAE) and the encephalitic nature of these Th17 cells can be explained by their ability to produce GM-CSF. However, the upstream regulatory mechanisms that control Csf2 (gene encoding GM-CSF) expression are still unclear. In this study, we found that Th17 cells dynamically expressed GATA3, the master transcription factor for Th2 cell differentiation, during their differentiation both in vitro and in vivo. Early deletion of Gata3 in three complimentary conditional knockout models by Cre-ERT2, hCd2 Cre and Tbx21 Cre, respectively, limited the pathogenicity of Th17 cells during EAE, which was correlated with a defect in generating pathogenic T-bet-expressing Th17 cells. These results indicate that early GATA3-dependent gene regulation is critically required to generate a de novo encephalitogenic Th17 response. Furthermore, a late deletion of Gata3 via Cre-ERT2 in the adoptive transfer EAE model resulted in a cell intrinsic failure to induce EAE symptoms which was correlated with a substantial reduction in GM-CSF production without affecting the generation and/or maintenance of T-bet-expressing Th17 cells. RNA-Seq analysis of Gata3-sufficient and Gata3-deficient CNS-infiltrating CD4+ effector T cells from mixed congenic co-transfer recipient mice revealed an important, cell-intrinsic, function of GATA3 in regulating the expression of Egr2, Bhlhe40, and Csf2. Thus, our data highlights a novel role for GATA3 in promoting and maintaining the pathogenicity of T-bet-expressing Th17 cells in EAE, via putative regulation of Egr2, Bhlhe40, and GM-CSF expression.

T-cell differentiation is a tightly regulated developmental program governed by interactions between transcription factors (TFs) and chromatin landscapes and affected by signals received from the thymic stroma. This process is marked by a series of checkpoints: T-lineage commitment, T-cell receptor (TCR)β selection, and positive and negative selection. Dynamically changing combinations of TFs drive differentiation along the T-lineage trajectory, through mechanisms that have been most extensively dissected in adult mouse T-lineage cells. However, fetal T-cell development differs from adult in ways that suggest that these TF mechanisms are not fully deterministic. The first wave of fetal T-cell differentiation occurs during a unique developmental window during thymic morphogenesis, shows more rapid kinetics of differentiation with fewer rounds of cell division, and gives rise to unique populations of innate lymphoid cells (ILCs) and invariant γδT cells that are not generated in the adult thymus. As the characteristic kinetics and progeny biases are cell-intrinsic properties of thymic progenitors, the differences could be based on distinct TF network circuitry within the progenitors themselves. Here, we review recent single-cell transcriptome data that illuminate the TF networks involved in T-cell differentiation in the fetal and adult mouse thymus.

Huge progress has been made in understanding the biology of innate lymphoid cells (ILC) by adopting several well-known concepts in T cell biology. As such, flow cytometry gating strategies and markers, such as CD90, have been applied to indentify ILC. Here, we report that most non-NK intestinal ILC have a high expression of CD90 as expected, but surprisingly a sub-population of cells exhibit low or even no expression of this marker. CD90-negative and CD90-low CD127+ ILC were present amongst all ILC subsets in the gut. The frequency of CD90-negative and CD90-low CD127+ ILC was dependent on stimulatory cues in vitro and enhanced by dysbiosis in vivo. CD90-negative and CD90-low CD127+ ILC were a potential source of IL-13, IFNγ and IL-17A at steady state and upon dysbiosis- and dextran sulphate sodium-elicited colitis. Hence, this study reveals that, contrary to expectations, CD90 is not constitutively expressed by functional ILC in the gut.

Group 2 innate lymphoid cells (ILC2s) are a category of heterogeneous cells that produce the cytokines IL-5 and IL-13, which mediate the type 2 immune response. However, specific drug targets on lung ILC2s have rarely been reported. Previous studies have shown that type 2 cytokines, such as IL-5 and IL-13, are related to depression. Here, we demonstrated the negative correlation between the depression-associated monoamine neurotransmitter serotonin and secretion of the cytokines IL-5 and IL-13 by ILC2s in individuals with depression. Interestingly, serotonin ameliorates papain-induced lung inflammation by suppressing ILC2 activation. Our data showed that the serotonin receptor HTR2A was highly expressed on ILC2s from mouse lungs and human PBMCs. Furthermore, an HTR2A selective agonist (DOI) impaired ILC2 activation and alleviated the type 2 immune response in vivo and in vitro. Mice with ILC2-specific depletion of HTR2A (Il5cre/+·Htr2aflox/flox mice) abolished the DOI-mediated inhibition of ILC2s in a papain-induced mouse model of inflammation. In conclusion, serotonin and DOI could restrict the type 2 lung immune response, indicating a potential treatment strategy for type 2 lung inflammation by targeting HTR2A on ST2+ ILC2s.