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The immune system of laboratory mice raised in an ultra-hygienic environment resembles that ofnewborn humans, but can be induced to resemble the immune system of adult humans or 'dirty' mice by co-housing with pet store-bought mice. Do 'dirty' mice make better immunological models? The laboratory mouse is by far the dominant model organism for in vivo immunological research which — particularly in the light of disappointing results obtained with some recent transfers of disease treatments from laboratory to clinic — raises the question of how accurately the model reflects the human immune system. These authors compare the immune status of laboratory mice with that of feral mice and with mice bought commercially as pets. They find that the immune system of the ubiquitous laboratory 'specific pathogen free' mouse approximates that of human neonates, rather than human adults. Co-housing laboratory mice with 'pet store' mice leads to maturation of the immune system, making it more similar to that of the human adult, and resulting in increased resistance in several models of infection. The use of such 'dirty' mice could supplement current models to either increase translational potential to human disease or to better inform the efficacy of preclinical prophylactic and therapeutic modalities. Our current understanding of immunology was largely defined in laboratory mice, partly because they are inbred and genetically homogeneous, can be genetically manipulated, allow kinetic tissue analyses to be carried out from the onset of disease, and permit the use of tractable disease models. Comparably reductionist experiments are neither technically nor ethically possible in humans. However, there is growing concern that laboratory mice do not reflect relevant aspects of the human immune system, which may account for failures to translate disease treatments from bench to bedside 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 . Laboratory mice live in abnormally hygienic specific pathogen free (SPF) barrier facilities. Here we show that standard laboratory mouse husbandry has profound effects on the immune system and that environmental changes produce mice with immune systems closer to those of adult humans. Laboratory mice—like newborn, but not adult, humans—lack effector-differentiated and mucosally distributed memory T cells. These cell populations were present in free-living barn populations of feral mice and pet store mice with diverse microbial experience, and were induced in laboratory mice after co-housing with pet store mice, suggesting that the environment is involved in the induction of these cells. Altering the living conditions of mice profoundly affected the cellular composition of the innate and adaptive immune systems, resulted in global changes in blood cell gene expression to patterns that more closely reflected the immune signatures of adult humans rather than neonates, altered resistance to infection, and influenced T-cell differentiation in response to a de novo viral infection. These data highlight the effects of environment on the basal immune state and response to infection and suggest that restoring physiological microbial exposure in laboratory mice could provide a relevant tool for modelling immunological events in free-living organisms, including humans.

Segmented filamentous bacteria drive the acquisition of the T H 17 phenotype in an antigen-specific manner; these findings begin to elucidate how gut-induced T H 17 cells can contribute to distal organ-specific autoimmune disease. T H 17 cell differentiation linked to intestinal bacteria Colonization of the small intestine by microbes such as segmented filamentous bacteria is known to enhance the induction of T-helper-17 (T H 17) cells, which are important factors in both mucosal defence and in autoimmune disease pathogenesis. Here Dan Littman and colleagues demonstrate that the vast majority of T H 17 cells in mice colonized with segmented filamentous bacteria are directed at antigens encoded by these bacteria, and identify specific bacterial epitopes that are recognized by T H 17 T-cell receptors. This work provides insights into how microbiota communicate with the host immune system, and suggests possible routes for developing novel mucosal vaccines. T-helper-17 (T H 17) cells have critical roles in mucosal defence and in autoimmune disease pathogenesis 1 , 2 , 3 . They are most abundant in the small intestine lamina propria, where their presence requires colonization of mice with microbiota 4 , 5 , 6 , 7 . Segmented filamentous bacteria (SFB) are sufficient to induce T H 17 cells and to promote T H 17-dependent autoimmune disease in animal models 8 , 9 , 10 , 11 , 12 , 13 , 14 . However, the specificity of T H 17 cells, the mechanism of their induction by distinct bacteria, and the means by which they foster tissue-specific inflammation remain unknown. Here we show that the T-cell antigen receptor (TCR) repertoire of intestinal T H 17 cells in SFB-colonized mice has minimal overlap with that of other intestinal CD4 + T cells and that most T H 17 cells, but not other T cells, recognize antigens encoded by SFB. T cells with antigen receptors specific for SFB-encoded peptides differentiated into RORγt-expressing T H 17 cells, even if SFB-colonized mice also harboured a strong T H 1 cell inducer, Listeria monocytogenes , in their intestine. The match of T-cell effector function with antigen specificity is thus determined by the type of bacteria that produce the antigen. These findings have significant implications for understanding how commensal microbiota contribute to organ-specific autoimmunity and for developing novel mucosal vaccines.

When exposed to antigens, naïve B cells differentiate into different types of effector cells: antibody-producing plasma cells, germinal center cells, or memory cells. Whether an individual naïve B cell can produce all of these different cell fates remains unclear. Using a limiting dilution approach, we found that many individual naïve B cells produced only one type of effector cell subset, whereas others produced all subsets. The capacity to differentiate into multiple subsets was a characteristic of clonal populations that divided many times and resisted apoptosis, but was independent of isotype switching. Antigen receptor affinity also influenced effector cell differentiation. These findings suggest that diverse effector cell types arise in the primary immune response as a result of heterogeneity in responses by individual naïve B cells.

Although lung myeloid cells provide an intracellular niche for Mycobacterium tuberculosis ( Mtb ), CD4 + T cells limit Mtb growth in these cells to protect the host. The CD4 + T cell activities including interferon-γ (IFN-γ) production that account for this protection are poorly understood. Using intravenous antibody labeling and lineage-tracing reporter mice, we show that monocyte-derived macrophages (MDMs), rather than phenotypically similar monocytes or dendritic cells, are preferentially infected with Mtb in murine lungs. MDMs were recruited to the lungs by Mtb- specific CD4 + T cells via IFN-γ, which promoted the extravasation of monocyte precursors from the blood. It was possible that CD4 + T cells recruited infectable MDMs because these cells are uniquely poised to receive cognate MHCII-mediated help to control intracellular bacteria. Mice with MHCII deficiency in monocyte-derived cells had normal Mtb -specific CD4 + T cell activation, expansion and differentiation but the CD4 + T cells were unable to attenuate Mtb growth. Using single cell RNA sequencing, we showed that MDMs receiving cognate MHCII-mediated help from CD4 + T cells upregulated glycolytic genes associated with Mtb control. Overall, the results indicate that CD4 + T cells recruit infectable MDMs to the lungs and then trigger glycolysis-dependent bacterial control in the MDMs by engaging their MHCII-bound Mtb peptides. Moreover, the results suggest that cognate MHCII-mediated help to promote MDM glycolysis is an essential, IFN-γ-independent effector function of Mtb -specific CD4 + T cells.

The canine rabies vaccine consists of the whole killed rabies virus and an alum adjuvant. While it is known to provide immunological protection in dogs, its effects on cell-mediated responses remain largely uncharacterized. Here, we analyzed blood and spleen samples from vaccinated dogs to understand adaptive immune responses ex vivo following restimulation with rabies vaccine antigens. Our results showed that recombinant rabies virus glycoprotein (RABV-G) elicited higher antibody titers and IFNγ production compared to recombinant rabies virus nucleoprotein (RABV-N). CD4 + and CD4 + CD8 + double-positive (DP) T cells proliferate robustly after five days of RABV-G stimulation, which was inhibited by an anti-canine MHC class II blocking antibody. Both RABV-G-specific CD4 + and DP T cells demonstrated a polarized T H 1 phenotype, with minor subsets showing T H 1/T H 17 hybrid and pathogenic T H 1/T H 17 hybrid cell features. CD4 + T cells were primarily effector memory T cells (T EM ), while DP T cells exhibited a terminally differentiated effector memory phenotype that re-expressed CD45RA (T EMRA ). Both RABV-G-specific CD4 + and DP T cells were detectable up to 1,024 days post-vaccination in spleen samples and their proliferative capacities were unaffected by age. Our results provide the first characterization of canine RABV-G-specific T cell phenotypes in the spleen and blood following rabies vaccination.

Pulmonary mycoses are often associated with type-2 helper T (Th2) cell responses. However, mechanisms of Th2 cell accumulation are multifactorial and incompletely known. To investigate Th2 cell responses to pulmonary fungal infection, we developed a peptide-MHCII tetramer to track antigen-specific CD4+ T cells produced in response to infection with the fungal pathogen Cryptococcus neoformans. We noted massive accruement of pathologic cryptococcal antigen-specific Th2 cells in the lungs following infection that was coordinated by lung-resident CD11b+ IRF4-dependent conventional dendritic cells. Other researchers have demonstrated that this dendritic cell subset is also capable of priming protective Th17 cell responses to another pulmonary fungal infection, Aspergillus fumigatus. Thus, higher order detection of specific features of fungal infection by these dendritic cells must direct Th2 cell lineage commitment. Since chitin-containing parasites commonly elicit Th2 responses, we hypothesized that recognition of fungal chitin is an important determinant of Th2 cell-mediated mycosis. Using C. neoformans mutants or purified chitin, we found that chitin abundance impacted Th2 cell accumulation and disease. Importantly, we determined Th2 cell induction depended on cleavage of chitin via the mammalian chitinase, chitotriosidase, an enzyme that was also prevalent in humans experiencing overt cryptococcosis. The data presented herein offers a new perspective on fungal disease susceptibility, whereby chitin recognition via chitotriosidase leads to the initiation of harmful Th2 cell differentiation by CD11b+ conventional dendritic cells in response to pulmonary fungal infection.

Intranasal (i.n.) infections preferentially generate Th17 cells. We explored the basis for this anatomic preference by tracking polyclonal CD4⁺ T cells specific for an MHC class II-bound peptide from the mucosal pathogenStreptococcus pyogenes. S. pyogenesMHC class II-bound peptide-specific CD4⁺ T cells were first activated in the cervical lymph nodes following i.n. inoculation and then differentiated into Th17 cells.S. pyogenes-induced Th17 formation depended on TGF-β1 from dendritic cells and IL-6 from a CD301b⁺ dendritic cell subset located in the cervical lymph nodes but not the spleen. Thus, the tendency of i.n. infection to induce Th17 cells is related to cytokine production by specialized dendritic cells that drain this site.

The immune response elicited after Mycobacterium tuberculosis (Mtb) infection is critically dependent on CD4 T cells during both acute and chronic infection. How CD4 T-cell responses are maintained throughout infection is not well understood, and evidence from other infection models has suggested that, under conditions of chronic antigen stimulation, T cells can undergo replicative exhaustion. These findings led us to determine whether subpopulations of CD4 T cells existed that displayed markers of terminal differentiation or exhaustion during murine Mtb infection. Analysis of antigen-specific effector CD4 T cells revealed that programmed death-1 (PD-1) and the killer cell lectin-like receptor G1 (KLRG1) delineated subpopulations of T cells. PD-1–expressing CD4 T cells were highly proliferative, whereas KLRG1 cells exhibited a short lifespan and secreted the cytokines IFNγ and TNFα. Adoptive transfer studies demonstrated that proliferating PD-1–positive CD4 T cells differentiated into cytokine-secreting KLRG1-positive T cells, but not vice versa. Thus, proliferating PD-1–positive cells are not exhausted, but appear to be central to maintaining antigen-specific effector T cells during chronic Mtb infection. Our findings suggest that antigen-specific T-cell responses are maintained during chronic mycobacterial infection through the continual production of terminal effector cells from a proliferating precursor population.

The autoimmune regulator (Aire) plays a critical role in central tolerance by promoting the display of tissue-specific antigens in the thymus. To study the influence of Aire on thymic selection in a physiological setting, we used tetramer reagents to detect autoreactive T cells specific for the Aire-dependent tissue-specific antigen interphotoreceptor retinoid-binding protein (IRBP), in the polyclonal repertoire. Two class II tetramer reagents were designed to identify T cells specific for two different peptide epitopes of IRBP. Analyses of the polyclonal T-cell repertoire showed a high frequency of activated T cells specific for both IRBP tetramers in Aire–/– mice, but not in Aire+/+ mice. Surprisingly, although one tetramer-binding T-cell population was efficiently deleted in the thymus in an Aire-dependent manner, the second tetramer-binding population was not deleted and could be detected in both the Aire–/– and Aire+/+ T-cell repertoires. We found that Aire-dependent thymic deletion of IRBP-specific T cells relies on intercellular transfer of IRBP between thymic stroma and bone marrow-derived antigen-presenting cells. Furthermore, our data suggest that Aire-mediated deletion relies not only on thymic expression of IRBP, but also on proper antigen processing and presentation of IRBP by thymic antigen-presenting cells.

Group A streptococcus (GAS, Streptococcus pyogenes) is the cause of a variety of clinical conditions, ranging from pharyngitis to autoimmune disease. Peptide-major histocompatibility complex class II (pMHCII) tetramers have recently emerged as a highly sensitive means to quantify pMHCII-specific CD4+ helper T cells and evaluate their contribution to both protective immunity and autoimmune complications induced by specific bacterial pathogens. In lieu of identifying an immunodominant peptide expressed by GAS, a surrogate peptide (2W) was fused to the highly expressed M1 protein on the surface of GAS to allow in-depth analysis of the CD4+ helper T cell response in C57BL/6 mice that express the I-A(b) MHCII molecule. Following intranasal inoculation with GAS-2W, antigen-experienced 2W:I-A(b)-specific CD4+ T cells were identified in the nasal-associated lymphoid tissue (NALT) that produced IL-17A or IL-17A and IFN-γ if infection was recurrent. The dominant Th17 response was also dependent on the intranasal route of inoculation; intravenous or subcutaneous inoculations produced primarily IFN-γ+ 2W:I-A(b+) CD4+ T cells. The acquisition of IL-17A production by 2W:I-A(b)-specific T cells and the capacity of mice to survive infection depended on the innate cytokine IL-6. IL-6-deficient mice that survived infection became long-term carriers despite the presence of abundant IFN-γ-producing 2W:I-A(b)-specific CD4+ T cells. Our results suggest that an imbalance between IL-17- and IFN-γ-producing CD4+ T cells could contribute to GAS carriage in humans.