Nutrition influences host physiological processes, yet how diets reshape host physiology, microbial functions, or host-microbe interactions to promote regeneration remains poorly explored. Here, we show that a ketogenic diet (KD), enriched in fats and low in carbohydrates, reprograms both skin microbial and immune functions to promote tissue repair. KD enhances IL-17A activity in γδ T cells and mucosal-associated invariant T (MAIT) cells, accelerating tissue repair, while KD-induced skin lipidomic alterations enhance both the abundance and metabolic output of Staphylococcus epidermidis. Metatranscriptomic and lipidomic analyses revealed increased riboflavin biosynthesis and sphingomyelinase (Sph)-dependent ceramide production in S. epidermidis under KD conditions. Genetic depletion of microbial ribD, a key enzyme for riboflavin biosynthesis, or of sph compromised the ability of the bacteria to promote tissue repair. Thus, host nutritional status drives tissue regeneration by synergistically rewiring host and microbial functions, providing new insights into how diet can be harnessed to regulate host physiology.

Intestinal immunity defends against enteric pathogens, mediates symbiotic relationships with the resident microbiota, and provides tolerance to food antigens, safeguarding critical nutrient absorption and barrier functions of this mucosal tissue. Despite the abundance of tissue resident activated T cells, their contributions to these various roles remain poorly understood. Here, we identify a dominant population of IL-10 producing, T-bet-expressing Tr1 T cells, residing in the small intestinal lamina propria at homeostasis. Remarkably, these intestinal Tr1 cells emerge at the time of weaning and accumulate independently of the microbiota displaying similar abundance, function, and TCR repertoire under germ-free conditions. Instead, the small intestinal T-bet+ Tr1 program is driven and shaped by dietary antigens, and accumulates in a cDC1-IL-27-dependent manner. Upon activation, these cells robustly express IL-10 and multiple inhibitory receptors, establishing a distinct suppressive profile. Altogether, this work uncovers a previously unappreciated dominant player in homeostatic small intestinal immunity with the potential to play critical suppressive roles in this tissue, raising important implications for the understanding of immune regulation in the intestine.

The interaction between the immune system and the somatosensory system plays a fundamental role in the regulation of diverse biological processes. Chronic itch is a common yet hard-to-treat symptom of many inflammatory skin conditions. One hallmark of chronic itch is the hyperinnervation of the skin by sensory fibers, yet what drives this aberrant nerve growth or how it contributes to disease progression remains unclear. Here, we identify IL-17A and immunity to skin microbiota as key triggers of sensory neuron plasticity and pruritus. In a murine model of psoriatic itch, we show that exposure to Staphylococcus aureus prior to experimental psoriasis results in heightened skin inflammation, increased itch, and marked hyperinnervation of CGRPα+ sensory neurons. Accordingly, single-nuclei RNA sequencing of dorsal root ganglia reveals that microbiota-driven inflammation induces a regenerative transcriptional program in sensory neurons, including upregulation of axonal growth, injury response, and IL-17RA signaling pathways. Mechanistically, we show that IL-17A/IL-17RA signaling within TRPV1+ sensory neurons drives hyperinnervation and pruritus, establishing a causal link between IL-17A and microbiota-driven immune responses in sensory circuit remodeling. Further, we identify sensory hyperinnervation as a key driver of chronic itch and inflammation. Collectively, we reveal that aberrant IL-17A signaling in sensory neurons, triggered by dysregulated microbiota immunity, promotes neuronal remodeling that amplifies itch and inflammation. These findings provide a framework for targeting microbiota-neuroimmune interactions as a therapeutic strategy for pruritus.

Breastfeeding is an obligatory requirement of mammalian survival. This fundamental process is associated with the adaptation of maternal physiology, including the transformation of the mammary gland into a milk-secreting organ. How maternal immunity contributes to mammary gland remodeling and function remains largely unknown. Here, we show that maternal adaptive immunity plays a critical role in shaping lactogenesis. Specifically, physiological adaptation during pregnancy is associated with thymic involution and a paradoxical enrichment in intraepithelial lymphocyte (IEL) precursors that no longer migrate to the gut but instead preferentially accumulate within the mammary gland. IEL precursors differentiate into T-bet-expressing unconventional CD8αα lymphocytes in an IL-15-dependent manner. Mammary IELs control milk production by favoring the differentiation and maturation of contractile and milk-secreting cells, thereby promoting offspring fitness. Altogether, this work uncovers a contribution of the maternal adaptive immune system in organismal remodeling during pregnancy that is associated with mammary gland development and function.

Persistent antigen exposure in chronic infections and cancer leads to a progressive state of T cell dysfunction known as exhaustion, which represents a major barrier to effective immune control, but allows antigen-specific T cells to persist. Understanding signaling pathways that mitigate exhaustion and reinvigorate CD8+ T cell effector function is a key goal for immunotherapeutic strategies. Here, we show that an activating mutant of phosphoinositide-3-kinase delta (PI3Kdelta) led to a reduction of FoxO1-dependent TCF-1+ stem-like progenitor CD8+ T cells that are required for sustaining antigen-specific T cells in response to chronic viral infection. Nonetheless, mice expressing activated PI3Kdelta maintained CD8+ T cell responses that were skewed instead towards effector-like cells in a FoxO1-independent manner, associated with an amplified IL-21-STAT3 response axis and improved viral control. Activated PI3Kdelta limited TOX expression, prevented epigenetic changes associated with T cell exhaustion, and promoted effector differentiation and function from both progenitor stem-like cells and cells with an exhausted phenotype. Together, this work uncovers a key role for PI3Kdelta activation in shaping the balance and plasticity between effector function and exhaustion while promoting T cell persistence during chronic infection, providing insight for immunotherapeutic strategies. ### Competing Interest Statement The authors have declared no competing interest. NIAID, Division of Intramural Research, AI001240-07

Males and females exhibit profound differences in immune responses and disease susceptibility. However, the factors responsible for sex differences in tissue immunity remain poorly understood. Here, we uncovered a dominant role for type 2 innate lymphoid cells (ILC2s) in shaping sexual immune dimorphism within the skin. Mechanistically, negative regulation of ILC2s by androgens leads to a reduction in dendritic cell accumulation and activation in males, along with reduced tissue immunity. Collectively, our results reveal a role for the androgen-ILC2-dendritic cell axis in controlling sexual immune dimorphism. Moreover, this work proposes that tissue immune set points are defined by the dual action of sex hormones and the microbiota, with sex hormones controlling the strength of local immunity and microbiota calibrating its tone.

Pregnancy presents a challenging immunological dilemma. The maternal immune system must tolerate fetal antigens, which implies pregnancy may induce specific mechanisms of immunoregulation. Yet within this regulatory milieu, protection against pathogens must be maintained. How does the maternal immune system navigate these opposing demands? Successfully navigating this immunological dilemma is essential for supporting the reproductive fitness of the organism, and generally, the survival of any mammalian species. Of particular importance in this context are barrier tissues, like the skin, which represent the primary portal of entry for pathogens. These barrier tissues are an especially important first line of defense during pregnancy, since adaptive immunity must be regulated to tolerate the fetal antigens which distribute throughout maternal tissues. However, little is known about how immunity in barrier tissues is altered during pregnancy. Our preliminary studies have identified an increase in ceramide levels in the skin during pregnancy. Ceramides have been shown to be critical mediators of skin barrier function, and may prevent pathogens from breaching the skin. Additionally, in the setting of S. aureus infection, pregnant skin demonstrated increased infiltration of innate immune cells, as well as an increase in type I immunity among adaptive cells. Together, these data suggest that the protective mechanisms of the skin are altered during pregnancy. Further insights in this area may elucidate an especially critical role for barrier tissue immunity in defending against infection during pregancy, and may eventually help guide improved clinical strategies to prevent and treat infections in pregnant mothers. Supported by Perelman School of Medicine's Medical Scientist Training Program and intramural research funding from NIAID.

Mild or transient dietary restriction (DR) improves many aspects of health and aging. Emerging evidence from us and others has demonstrated that DR also optimizes the development and quality of immune responses. However, the factors and mechanisms involved remain to be elucidated. Here, we propose that DR-induced optimization of immunological memory requires a complex cascade of events involving memory T cells, the intestinal microbiota, and myeloid cells. Our findings suggest that DR enhances the ability of memory T cells to recruit and activate myeloid cells in the context of a secondary infection. Concomitantly, DR promotes the expansion of commensal Bifidobacteria within the large intestine, which produce the short-chain fatty acid acetate. Acetate conditioning of the myeloid compartment during DR enhances the capacity of these cells to kill pathogens. Enhanced host protection during DR is compromised when Bifidobacteria expansion is prevented, indicating that microbiota configuration and function play an important role in determining immune responsiveness to this dietary intervention. Altogether, our study supports the idea that DR induces both memory T cells and the gut microbiota to produce distinct factors that converge on myeloid cells to promote optimal pathogen control. These findings suggest that nutritional cues can promote adaptation and co-operation between multiple immune cells and the gut microbiota, which synergize to optimize immunity and protect the collective metaorganism.

Barrier tissues such as the skin and gut form the first line of defense against pathogens and establish an immune dialogue with the resident microbiota. To that end, they harbor immune populations geared towards barrier function, tolerance, and repair. Of particular interest are the large numbers of T cells. Some are specific towards commensal microbes or recognize previously encountered pathogens. However, the origin, antigen specificity, and function of most barrier tissue-resident T cells remains elusive. We have identified a dominant CD4 T cell population in the small intestine lamina propria characterized by the expression of T-bet. Surprisingly, this population is present in both germ-free and conventionally reared (SPF) mice at homeostasis, and accounts for almost half of T cells at this site. Bulk and single-cell transcriptome analysis revealed that this population has cytotoxic potential. Furthermore, single-cell TCR sequencing showed parallel T cell repertoires in germ-free and SPF mice, suggesting shared antigen specificity. To explore the ontogeny of this population, we have identified clonally expanded TCRs and are generating low frequency retrogenic mice. Additionally, we have generated several T cell hybridomas from small intestine Th1 cells. Interestingly, preliminary results suggest that small intestinal Th1 TCRs exhibit some degree of self-reactivity, which may determine their development and differentiation in a foreign antigen-independent manner. Collectively, these studies will shed light on the function of a dominant intestinal T cell population and reveal novel insights into homeostatic tissue immune networks. This research was supported by the Intramural Research Program of NIAID, NIH.

The skin immune system mediates the skin health status, but the sex differences of the skin immune system are still unclear. This study aims to investigate sex differences in the skin immune system and decipher the underlying regulatory factors. We found that female mice have a higher magnitude of adaptive immune responses during commensal bacteria association and pathogen infection. Correspondingly, females have a higher level of skin dendritic cells (DCs), which play a fundamental role in commensal-induced adaptive immune responses. These sex differences were regulated by male sex hormones, that castration of males normalized the sex differences in the level of skin immune cells as well as the adaptive immune responses to bacteria. However, our scRNA-seq data reveals that androgen receptor is not expressed in DCs, but highly expressed in skin ILC2. Females have a significantly higher level of ILC2 than males and ILC2 from females also have a more activating gene expression signature than ILC2 from males. Females also have a significantly higher level of ILC2-produced IL-13 and GM-CSF than male mice. In addition, our data reveal that IL-13 and GM-CSF play a critical role to maintain the level of skin DCs, that IL-13/GM-CSF deficient mice have a remarkable decrease of skin DCs, and the sex differences of DCs are also largely impaired. Therefore, based on these findings, we suggest that androgen signaling negatively regulates the level of skin ILC2 and DCs, thereby shaping sex-specific skin immunity to commensals, pathogens, and/or other stimuli. Collectively our work proposes a mechanism for the heighten immunity observed in female and have important implication for our understanding of tissue immunity in the context of infection and cancer.

Breastfeeding has long been associated with health benefits for the infant. Breast milk is composed of essential nutrients and bioactive molecules shaping the growth, and the immune development of the offspring. An optimal development of the mammary gland (MG) during pregnancy is critical to maintaining the beneficial effects of lactation. Nevertheless, how the lactation process is regulated by the immune system remains largely unexplored. Our data reveal that the commitment of mammary epithelial cells into milk-secreting cells is associated with the accumulation of T-bet +lymphoid cells, preferentially localized at the proximity and inside the mammary epithelium. Among mammary T-bet +cells, we characterized an unconventional T cell population exhibiting common features with gut CD8αα +intra-epithelial lymphocytes and deriving from the same thymic progenitor. Such process is associated with thymus involution during pregnancy and specific enrichment in thymic precursor that preferentially migrates into the MG during pregnancy. These cells promote milk production and expression of β-casein, one of the main milk protein, resulting in an increase in newborn weight. This work uncovers a previously unappreciated contribution for the adaptive immune system in mammary gland development and lactation. Decoding the immune regulation of the MG will allow to understand the profound transformation of this compartment as a primary nutritional source for the offspring and may allow to understand how dysregulation of this process could affect the long-term development of newborns. This work was supported in part by intramural funds of NIAID, NIH.

Breast milk has been shown to play a key role in the transfer of immunity from mother to offspring. Antibodies and lymphoid cells can be passed through breast milk and impact offspring immunity. However, the role of these lymphoid cells in lactation and in the control of offspring immunity remains unclear. We observed that the lymphoid cells present in the breast milk are distinct from those found in the blood. Compared to the blood, breast milk T cells are enriched in a T-bet +unconventional population. Imaging analysis revealed that these T-bet +cells gradually accumulate in the mammary gland during pregnancy and are specifically localized around and inside the mammary epithelium. We will utilize confocal and intravital imaging of the mammary gland in virgin, pregnant, and lactating mice to characterize the dynamics of this mammary-associated T-bet +population throughout pregnancy and lactation. Understanding the dynamics of T-bet +cells and their anatomical localization within the mammary gland will help us decode how these cells impact the remodeling of the mammary epithelium during pregnancy and lactation. We propose that the remodeling of the mammary gland imparted by the immune system can also influence the transfer of immunity from mother to offspring, thereby affecting the development of offspring immunity. This work was supported in part by intramural funds of NIAID, NIH.

Tissue-resident innate lymphoid cells (ILCs) regulate tissue homeostasis, protect against pathogens at mucosal surfaces, and are key players at the interface of innate and adaptive immunity. How ILCs adapt their phenotype and function to environmental cues within tissues remains to be fully understood. Here, we show that Mycobacterium tuberculosis (Mtb) infection alters the phenotype and function of lung IL-18Rα+ ILC toward a protective interferon-γ-producing ILC1-like population. This differentiation is controlled by type 1 cytokines and is associated with a glycolytic program. Moreover, a BCG-driven type I milieu enhances the early generation of ILC1-like cells during secondary challenge with Mtb. Collectively, our data reveal how tissue-resident ILCs adapt to type 1 inflammation toward a pathogen-tailored immune response.

Tissue-resident innate lymphoid cells (ILCs) regulate tissue homeostasis and protect against pathogens at mucosal surfaces and are key players at the interface of innate and adaptive immunity. How ILCs adapt their phenotype and function to environmental cues in their tissue of residence remains to be fully understood. Here we show that Mycobacterium tuberculosis infection alters the biology of lung ILCs and, in particular, induces the emergence of a non-classical, protective, interferon-γ-producing ILC1-like population. Adoptive transfer, fate-mapping and in vitro differentiation experiments revealed that ILC1-like cells originate from immature ILC2 rather than from mature ILC2. This plasticity is controlled by type 1 cytokines and a glycolytic program involving the transcription factor HIF1α. Collectively, our data reveal how tissue-resident ILCs adapt to their inflammatory and metabolic environment to undergo phenotypic and functional changes toward a pathogen-adapted immune response.

Tissue-resident innate lymphoid cells (ILCs) regulate tissue homeostasis, protect against pathogens at mucosal surfaces and are key players at the interface of innate and adaptive immunity. How ILCs adapt their phenotype and function to environmental cues within tissues remains to be fully understood. Here, we show that Mycobacterium tuberculosis infection alters the phenotype and function of immature lung ILC2 toward a protective interferon-γ-producing ILC1-like population. This differentiation is controlled by type 1 cytokines and is associated with a glycolytic program involving the transcription factor HIF1α. Collectively, our data reveal how tissue-resident ILCs adapt to type 1 inflammation toward a pathogen tailored immune response.

The lungs harbor multiple resident microbial communities, otherwise known as the microbiota. There is an emerging interest in deciphering whether the pulmonary microbiota modulate local immunity, and whether this knowledge could shed light on mechanisms operating in the response to respiratory pathogens. In this study, we investigate the capacity of a pulmonary Lactobacillus strain to modulate the lung T cell compartment and assess its prophylactic potential upon infection with Mycobacterium tuberculosis, the etiological agent of tuberculosis. In naive mice, we report that a Lactobacillus murinus (Lagilactobacillus murinus) strain (CNCM I-5314) increases the presence of lung Th17 cells and of a regulatory T cell (Treg) subset known as RORγt+ Tregs. In particular, intranasal but not intragastric administration of CNCM I-5314 increases the expansion of these lung leukocytes, suggesting a local rather than systemic effect. Resident Th17 and RORγt+ Tregs display an immunosuppressive phenotype that is accentuated by CNCM I-5314. Despite the well-known ability of M. tuberculosis to modulate lung immunity, the immunomodulatory effect by CNCM I-5314 is dominant, as Th17 and RORγt+ Tregs are still highly increased in the lung at 42-d postinfection. Importantly, CNCM I-5314 administration in M. tuberculosis-infected mice results in reduction of pulmonary inflammation, without increasing M. tuberculosis burden. Collectively, our findings provide evidence for an immunomodulatory capacity of CNCM I-5314 at steady state and in a model of chronic inflammation in which it can display a protective role, suggesting that L. murinus strains found in the lung may shape local T cells in mice and, perhaps, in humans.

Mycobacterium tuberculosis (Mtb) regulates the macrophage metabolic state to thrive in the host. Yet, the responsible mechanisms remain elusive. Macrophage activation towards the microbicidal (M1) program depends on the HIF-1α-mediated metabolic shift from oxidative phosphorylation towards glycolysis. Here, we asked whether a tuberculosis (TB) microenvironment changes the M1 macrophage metabolic state. We exposed M1 macrophages to the acellular fraction of tuberculous pleural effusions (TB-PE), and found lower glycolytic activity, accompanied by elevated levels of oxidative phosphorylation and bacillary load, compared to controls. The host-derived lipid fraction of TB-PE drove these metabolic alterations. HIF-1α stabilization reverted the effect of TB-PE by restoring M1 metabolism. As a proof-of-concept, Mtb-infected mice with stabilized HIF-1α displayed lower bacillary loads and a pronounced M1-like metabolic profile in alveolar macrophages. Collectively, we demonstrate that host-derived lipids from a TB-associated microenvironment alter the M1 macrophage metabolic reprogramming by hampering HIF-1α functions, thereby impairing control of Mtb infection.

Tuberculosis (TB), caused by the airborne bacterial pathogen Mycobacterium tuberculosis, remains a major source of morbidity and mortality worldwide. So far, the study of host-pathogen interactions in TB has mostly focused on the physiology and virulence of the pathogen, as well as, on the various innate and adaptive immune compartments of the host. Microbial organisms endogenous to our body, the so-called microbiota, interact not only with invading pathogens, but also with our immune system. Yet, the impact of the microbiota on host defense against M. tuberculosis remains poorly understood. In order to address this question, we adapted a robust and reproducible mouse model of microbial dysbiosis based on a combination of wide-spectrum antibiotics. We found that microbiota dysbiosis resulted in an increased early colonization of the lungs by M. tuberculosis during the first week of infection, correlating with an altered diversity of the gut microbiota during this time period. At the cellular level, no significant difference in the recruitment of conventional myeloid cells, including macrophages, dendritic cells and neutrophils, to the lungs could be detected during the first week of infection between microbiota-competent and -deficient mice. At the molecular level, microbiota depletion did not impact the global production of pro-inflammatory cytokines, such as interferon (IFN)γ, tumor necrosis factor (TNF)α and interleukin (IL)-1β in the lungs. Strikingly, a reduced number of mucosal-associated invariant T (MAIT) cells, a population of innate-like lymphocytes whose development is known to depend on the host microbiota, was observed in the lungs of the antibiotics-treated animals after 1week of infection. These cells produced less IL-17A in antibiotics-treated mice. Notably, dysbiosis correction through the inoculation of a complex microbiota in antibiotics-treated animals reversed these phenotypes and improved the ability of MAIT cells to proliferate. Altogether, our results demonstrate that the host microbiota contributes to early protection of lung colonization by M. tuberculosis, possibly through sustaining the function(s) of MAIT cells. Our study calls for a better understanding of the impact of the microbiota on host-pathogen interactions in TB. Ultimately, this study may help to develop novel therapeutic approaches based on the use of beneficial microbes, or components thereof, to boost anti-mycobacterial immunity.