Naive CD4 + T cells differentiate into functionally diverse T helper (Th) cell subsets. Th2 cells play a pathogenic role in asthma, yet a clear picture of their transcriptional profile is lacking. We performed single-cell RNA sequencing (scRNA-seq) of T helper cells from lymph node, lung, and airways in the house dust mite (HDM) model of allergic airway disease. scRNA-seq resolved transcriptional profiles of naive CD4 + T, Th1, Th2, regulatory T (Treg) cells, and a CD4 + T cell population responsive to type I interferons. Th2 cells in the airways were enriched for transcription of many genes, including Cd200r1, Il6, Plac8, and Igfbp7, and their mRNA profile was supported by analysis of chromatin accessibility and flow cytometry. Pathways associated with lipid metabolism were enriched in Th2 cells, and experiments with inhibitors of key metabolic pathways supported roles for glucose and lipid metabolism. These findings provide insight into the differentiation of pathogenic Th2 cells in the context of allergy.

A hallmark of immunity to worm infections and many allergies is a strong type 2 immune response. This is characterized by the production of cytokines interleukin-5 (IL-5) and IL-13 by adaptive T helper 2 (T2) cells and/or type 2 innate lymphoid cells. Peroxisome proliferator activated receptor-γ (PPAR-γ) is typically regarded as an anti-inflammatory factor. We report that T2 cells express high levels of PPAR-γ in response to the allergen house dust mite and after infection with the parasite Mice lacking PPAR-γ in T cells failed to effectively differentiate into IL-5- and IL-13-secreting cells and, hence, did not develop T2 cell-associated pathologies, including goblet cell metaplasia and eosinophilia, in response to allergen challenge. Furthermore, these mice could not mount protective immune responses to nematode infection. In addition, mice lacking PPAR-γ in T cells had greatly reduced frequencies of T2 cells in visceral adipose tissue. Mechanistically, PPAR-γ appeared to promote the expression of the IL-33 receptor on the surface of T2 cells. These results pinpoint PPAR-γ as a factor that drives type 2 responses in allergy, worm infection, and visceral adipose tissue.

Upon activation, naïve CD4+ T cells differentiate into a number of specialized T helper (Th) cell subsets. Th2 cells are central players in immunity to helminths and are implicated in mediating the inflammatory pathology associated with allergies. The differentiation of Th2 cells is dependent on transcription factors such as GATA3 and STAT6, which prime Th2 cells for the secretion of interleukin- (IL-) 4, IL-5, and IL-13. Several lines of work now suggest that differentiating Th2 cells in the lymph node are potent IL-4 cytokine producers, but do not become competent IL-5- and IL-13-producing cells until after receiving cues from non-lymphoid tissue. It is evident that Th2 cells that enter tissues undergo considerable changes in chromatin architecture and gene expression, and that over this time, the metabolic requirements of these cells change considerably. Herein, we discuss the metabolic requirements of Th2 cells during their early and late differentiation, focusing on the impact of glucose and lipid metabolism, mTOR activation, the nuclear receptor PPAR-γ and several metabolites.

Objective- Dyslipidemia is a component of the metabolic syndrome, an established risk factor for atherosclerotic cardiovascular disease, and is also observed in various autoimmune and chronic inflammatory conditions. However, there are limited opportunities to study the impact of acquired dyslipidemia on cardiovascular and immune pathology. Approach and Results- We designed a model system that allows for the conversion to a state of acute hyperlipidemia in adult life, so that the consequences of such a transition could be observed, through conditionally deleting APOE (apolipoprotein E) in the adult mouse. The transition to hypercholesterolemia was accompanied by adaptive immune responses, including the expansion of T lymphocyte helper cell 1, T follicular helper cell, and T regulatory subsets and the formation of germinal centers. Unlike steady-state Apoe mice, abrupt loss of APOE induced rapid production of antibodies recognizing rheumatoid disease autoantigens. Genetic ablation of the germinal center reduced both autoimmunity and atherosclerosis, indicating that the immune response that follows loss of APOE is independent of atherosclerosis but nevertheless promotes plaque development. Conclusions- Our findings suggest that immune activation in response to hyperlipidemia could contribute to a wide range of inflammatory autoimmune diseases, including atherosclerosis.

Introduction Innate lymphoid cells (ILCs) can provide early cytokine help against a variety of pathogens in the lungs and gastrointestinal tract. Type 2 ILC (ILC2) are comparable to T helper 2 cells found in the adaptive immune system, which secrete cytokines such as interleukin 5 (IL‐5) and IL‐13 and have been found to play roles in host defense against helminth infections and in allergic responses. Recent studies have identified that programmed cell death protein 1 (PD‐1) and peroxisome proliferator activated receptor‐γ (PPAR‐γ) are highly expressed by ILC2. We examined whether PD‐1 plays a role in ILC2 function and whether there was any connection between PD‐1 and PPAR‐γ Methods To ensure that only innate immune cells were present, ILC2 cells were examined from RAG1−/− and PD‐1−/−xRAG1−/− mice under steady‐state or following inoculation with IL‐33. We also tested ILC2 generated from bone marrow of RAG1−/− and PD‐1−/−xRAG1−/− mice for their production of cytokines. These in vitro‐derived ILC2 were also exposed to agonist and antagonist of PPAR‐γ. Results We found that ILC2 from PD‐1−/−xRAG1−/− mice had reduced frequencies of IL‐5 and IL‐13 producing cells both in vitro upon IL‐33 stimulation and in vivo following intraperitoneal administration of IL‐33 when compared with ILC2 from RAG1−/− mice. However, by adding IL‐2, IL‐25, and thymic stromal lymphopoietin to the in vitro cultures, the frequency of IL‐5 and IL‐13 expressing ILC2 from PD‐1−/−xRAG1−/− mice became similar to the frequency observed for ILC2 from RAG1−/− mice. In addition, PPAR‐γ agonists and antagonists were found to increase and decrease PD‐1 expression on ILC2 respectively. Conclusions These findings illustrate that chronic loss of PD‐1 plays a role in ILC2 function and PD‐1 expression can be modulated by PPAR‐γ.

Impaired classical NF‐κB pathway signaling causes reduced antibody responses to T‐independent (TI) antigens. We investigated the potential reasons for defective TI responses in mice lacking the atypical inhibitory kappa B (IκB) protein of the NF‐κB pathway, IκBNS. Analyses of the plasma cell compartment in vitro and in vivo after challenge with lipopolysaccharide (LPS) showed significant decreases in the frequencies of plasma cells in the absence of IκBNS. In vitro activation of B cells via the B cell receptor or via Toll‐like receptor 4 revealed that early activation events were unaffected in IκBNS‐deficient B cells, while proliferation was reduced compared to in similarly stimulated wildtype (wt) B cells. IκBNS‐deficient B cells also displayed impaired upregulation of the transmembrane activator and calcium modulator cyclophilin ligand interactor (TACI), which is essential for TI responses, and decreased sensitivity to TACI ligands upon stimulation. Furthermore, IκBNS‐deficient B cells, in contrast to wt B cells, displayed altered expression of IRF4, Blimp‐1 and Pax5 upon LPS‐induced differentiation, indicating impaired transcriptional regulation of plasma cell generation.

Mice deficient in central components of classical NF-κB signaling have low levels of circulating natural IgM antibodies and fail to respond to immunization with T-independent type 2 (TI-2) antigens. A plausible explanation for these defects is the severely reduced numbers of B-1 and marginal zone B (MZB) cells in such mice. By using an ethyl-N-nitrosourea mutagenesis screen, we identified a role for the atypical IκB protein IκBNS in humoral immunity. IκBNS-deficient mice lack B-1 cells and have severely reduced numbers of MZB cells, and thus resemble several other strains with defects in classical NF-κB signaling. We analyzed mice heterozygous for the identified IκBNS mutation and demonstrate that these mice have an intermediary phenotype in terms of levels of circulating IgM antibodies and responses to TI-2 antigens. However, in contrast to mice that are homozygous for the IκBNS mutation, the heterozygous mice had normal frequencies of B-1 and MZB cells. These results suggest that there is a requirement for IκBNS expression from two functional alleles for maintaining normal levels of circulating natural IgM antibodies and responses to TI-2 antigens.

Marginal zone (MZ) B cells reside in the splenic MZ and play important roles in T cell-independent humoral immune responses against blood-borne pathogens. IκBNS-deficient mice exhibit a severe reduction in the MZ B compartment but regain an MZ B population with age and, thus, represent a valuable model to examine the biology of MZ B cells. In this article, we characterized the MZ B cell defect in further detail and investigated the nature of the B cells that appear in the MZ of aged mice. Flow cytometry analysis of the splenic transitional B cell subsets demonstrated that MZ B cell development was blocked at the transitional-1 to transitional-2-MZ precursor stage in the absence of functional IκBNS. Immunohistochemical analysis of spleen sections from wild-type and mice revealed no alteration in the cellular MZ microenvironment, and analysis of bone marrow chimeras indicated that the MZ B cell development defect in mice was B cell intrinsic. Further, we demonstrate that the B cells that repopulate the MZ in aged mice were distinct from age-matched wild-type MZ B cells. Specifically, the expression of surface markers characteristic for MZ B cells was altered and the L chain Igλ repertoire was reduced in mice. Finally, plasma cell differentiation of sorted LPS-stimulated MZ B cells was impaired, and aged mice were unable to respond to NP-Ficoll immunization. These results demonstrate that IκBNS is required for an intact MZ B cell compartment in C57BL/6 mice.