Koji Atarashi

An individual's immune status is an essential factor affecting the efficacy of microbiome‐based therapies. To enable representative studies of microbiome–immune system interactions, we developed protocols to facilitate the coculture of intestinal epithelial cells and human peripheral blood mononuclear cells (PBMCs) in our microphysiological model of the human gut, HuMiX. Using the resulting immunoHuMiX model, we assessed responses to human–microbial cocultures with a probiotic mixture comprised of 17 strictly anaerobic Clostridial strains (17‐mix). We demonstrated the ability of our model to sustain metabolically active microbiota, while maintaining human cell viability, and generated relevant read‐outs for host–microbiome interaction studies. The composition of the strain mixture was not affected by the individual sources of PBMCs, and the abundances of the different immune cell populations were comparable when incubated with or without the 17‐mix after 24 h in the system, and between individuals. We observed increased cytokine production, PBMC viability, and PBMC cell counts upon coculture with the 17‐mix, thereby demonstrating immune sensing of the microbiota by the PBMCs. In summary, our immunoHuMiX model provides an important personalizable tool for screening microbiota‐driven immune responses.

Interactions between diet and the gut microbiota are fundamental to metabolic health, shaping energy balance and disease susceptibility^1-5. However, the underlying mechanisms by which dietary and microbial factors converge to regulate host physiology remain unclear. Here we show that protein availability profoundly modulates the functional landscape of the gut microbiota and promotes remodelling of white adipose tissue (WAT). Specifically, low-protein diets (LPDs) robustly induce signature genes of browning in WAT to a similar extent to that seen in response to classical stimuli, such as cold exposure or β-adrenergic receptor activation^6-8. LPD-mediated browning was markedly diminished in germ-free mice, and this defect was rescued by colonization with defined bacterial consortia made up of strains that were isolated and down-selected from the faeces of either LPD-fed mice or healthy human volunteers with ¹⁸F-fluorodeoxyglucose positron emission tomography (FDG-PET)-confirmed brown- or beige-fat activity^9-12. Microbiota-induced browning was mediated both by bile acids driving the activation of the farnesoid X receptor (FXR) in adipose progenitor cells, and by nrfA-encoding commensal-derived ammonia driving the expression of fibroblast growth factor 21 (FGF21) in hepatocytes. The bile acid-FXR and ammonia-FGF21 axes both have non-redundant, essential roles in promoting WAT browning. These findings highlight a mechanistic link between diet, gut microbial metabolism and adipose tissue remodelling, uncovering microbiota-dependent pathways by which the host responds to dietary cues.

Activation of brown and beige fat biogenesis promotes metabolic health in rodents and humans, but typically requires cold exposure or pharmacological activation of β-adrenergic receptors, which may pose cardiovascular risks. Dietary intervention represents a clinically viable alternative strategy to induce beige cells and thus enhance metabolic health, though the underlying mechanisms remain poorly understood. In this study, we identified specific microbiota members in both mice and humans that promote browning of white adipose tissue (WAT) and ameliorate metabolic disorders in the context of a low-protein diet (LPD). Diets with low protein, regardless of fat and carbohydrate content, induced robust WAT browning to a degree comparable to that achieved by cold exposure or β-adrenergic receptor agonist administration. LPD-mediated browning was markedly diminished in germ-free (GF) mice, highlighting the essential role of the gut microbiota. Microbiota-induced browning occurred independently of the immune system and primarily through mechanisms involving increased circulating deconjugated bile acids, activation of the farnesoid X receptor (FXR) in WAT progenitor cells, and enhanced hepatic production of FGF21. The browning defect in GF mice was rescued by transplanting microbiota from conventional mice or from brown/beige fat-positive healthy human volunteers, as determined by fluorodeoxyglucose positron-emission tomography (FDG-PET). Defined bacterial consortia, comprising strains isolated from mice or FDG-PET-positive human volunteers, were sufficient to elevate plasma bile acids and hepatic FGF21 levels by modulating nitrogen metabolism, ultimately restoring browning in response to a LPD. Our findings highlight the significant impact of diet-microbiota interactions on WAT browning and suggest their therapeutic potential for managing metabolic diseases.

Persistent colonization and outgrowth of potentially pathogenic organisms in the intestine can result from long-term antibiotic use or inflammatory conditions, and may perpetuate dysregulated immunity and tissue damage^1,2. Gram-negative Enterobacteriaceae gut pathobionts are particularly recalcitrant to conventional antibiotic treatment^3,4, although an emerging body of evidence suggests that manipulation of the commensal microbiota may be a practical alternative therapeutic strategy^5–7. Here we isolated and down-selected commensal bacterial consortia from stool samples from healthy humans that could strongly and specifically suppress intestinal Enterobacteriaceae. One of the elaborated consortia, comprising 18 commensal strains, effectively controlled ecological niches by regulating gluconate availability, thereby re-establishing colonization resistance and alleviating Klebsiella- and Escherichia-driven intestinal inflammation in mice. Harnessing these activities in the form of live bacterial therapies may represent a promising solution to combat the growing threat of proinflammatory, antimicrobial-resistant Enterobacteriaceae infection.

The liver is the main gateway from the gut, and the unidirectional sinusoidal flow from portal to central veins constitutes heterogenous zones, including the periportal vein (PV) and the pericentral vein zones^1,2,3,4,5. However, functional differences in the immune system in each zone remain poorly understood. Here intravital imaging revealed that inflammatory responses are suppressed in PV zones. Zone-specific single-cell transcriptomics detected a subset of immunosuppressive macrophages enriched in PV zones that express high levels of interleukin-10 and Marco, a scavenger receptor that sequesters pro-inflammatory pathogen-associated molecular patterns and damage-associated molecular patterns, and consequently suppress immune responses. Induction of Marco⁺ immunosuppressive macrophages depended on gut microbiota. In particular, a specific bacterial family, Odoribacteraceae, was identified to induce this macrophage subset through its postbiotic isoallolithocholic acid. Intestinal barrier leakage resulted in inflammation in PV zones, which was markedly augmented in Marco-deficient conditions. Chronic liver inflammatory diseases such as primary sclerosing cholangitis (PSC) and non-alcoholic steatohepatitis (NASH) showed decreased numbers of Marco⁺ macrophages. Functional ablation of Marco⁺ macrophages led to PSC-like inflammatory phenotypes related to colitis and exacerbated steatosis in NASH in animal experimental models. Collectively, commensal bacteria induce Marco⁺ immunosuppressive macrophages, which consequently limit excessive inflammation at the gateway of the liver. Failure of this self-limiting system promotes hepatic inflammatory disorders such as PSC and NASH.

Persistent colonization and outgrowth of pathogenic organisms in the intestine may occur due to long-term antibiotic usage or inflammatory conditions, which perpetuate dysregulated immunity and tissue damage^1,2. Gram-negative Enterobacteriaceae gut pathobionts are particularly recalcitrant to conventional antibiotic treatment^3,4, though an emerging body of evidence suggests that manipulation of the commensal microbiota may be a practical alternative therapeutic strategy^5-7. In this study, we rationally isolated and down-selected commensal bacterial consortia from healthy human stool samples capable of strongly and specifically suppressing intestinal Enterobacteriaceae . One of the elaborated consortia, consisting of 18 commensal strains, effectively controlled ecological niches by regulating gluconate availability, thereby reestablishing colonization resistance and alleviating antibiotic-resistant Klebsiella -driven intestinal inflammation in mice . Harnessing these microbial activities in the form of live bacterial therapeutics may represent a promising solution to combat the growing threat of proinflammatory, antimicrobial-resistant bacterial infection.

Diabetes is known to increase the risk of nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC). Here we treat male STAM (STelic Animal Model) mice, which develop diabetes, NASH and HCC associated with dysbiosis upon low-dose streptozotocin and high-fat diet (HFD), with insulin or phlorizin. Although both treatments ameliorate hyperglycemia and NASH, insulin treatment alone lead to suppression of HCC accompanied by improvement of dysbiosis and restoration of antimicrobial peptide production. There are some similarities in changes of microflora from insulin-treated patients comorbid with diabetes and NASH. Insulin treatment, however, fails to suppress HCC in the male STAM mice lacking insulin receptor specifically in intestinal epithelial cells (ieIRKO), which show dysbiosis and impaired gut barrier function. Furthermore, male ieIRKO mice are prone to develop HCC merely on HFD. These data suggest that impaired gut insulin signaling increases the risk of HCC, which can be countered by restoration of insulin action in diabetes.

Distinct gut microbiome ecology may be implicated in the prevention of aging-related diseases as it influences systemic immune function and resistance to infections. Yet, the viral component of the microbiome throughout different stages in life remains unexplored. Here we present a characterization of the centenarian gut virome using previously published metagenomes from 195 individuals from Japan and Sardinia. Compared with gut viromes of younger adults (>18 yr) and older individuals (>60 yr), centenarians had a more diverse virome including previously undescribed viral genera, such as viruses associated with Clostridia. A population shift towards higher lytic activity was also observed. Finally, we investigated phage-encoded auxiliary functions that influence bacterial physiology, which revealed an enrichment of genes supporting key steps in sulfate metabolic pathways. Phage and bacterial members of the centenarian microbiome displayed an increased potential for converting methionine to homocysteine, sulfate to sulfide and taurine to sulfide. A greater metabolic output of microbial hydrogen sulfide in centenarians may in turn support mucosal integrity and resistance to pathobionts.

Group 2 innate lymphoid cells (ILC2s) expressing IL-5 and IL-13 are localized at various mucosal tissues and play critical roles in the induction of type 2 inflammation, response to helminth infection, and tissue repair. Here, we reveal a unique ILC2 subset in the mouse intestine that constitutively expresses IL-4 together with GATA3, ST2, KLRG1, IL-17RB, and IL-5. In this subset, IL-4 expression is regulated by mechanisms similar to but distinct from those observed in T cells and is partly affected by IL-25 signaling. Although the absence of the microbiota had marginal effects, feeding mice with a vitamin B1-deficient diet compromised the number of intestinal IL-4+ ILC2s. The decrease in the number of IL-4+ ILC2s caused by the vitamin B1 deficiency was accompanied by a reduction in IL-25–producing tuft cells. Our findings reveal that dietary vitamin B1 plays a critical role in maintaining interaction between tuft cells and IL-4+ ILC2s, a previously uncharacterized immune cell population that may contribute to maintaining intestinal homeostasis.

Emergency myelopoiesis (EM) is a hematopoietic response against systemic infections that quickly supplies innate immune cells. As lymphopoiesis is strongly suppressed during EM, the role of lymphocytes in that process has not received much attention. Here, we found that myeloid-like B cells (M-B cells), which express myeloid markers, emerge in the bone marrow (BM) after the induction of EM. M-B cells were mainly derived from pre-B cells and preferentially expressed IL-10, which directly stimulates hematopoietic progenitors to enhance their survival and myeloid-biased differentiation. Indeed, lacking IL-10 in B cells, blocking IL-10 in the BM with a neutralizing antibody, and deleting the IL-10 receptor in hematopoietic progenitors significantly suppressed EM, which failed to clear microbes in a cecal ligation and puncture model. Thus, a distinct B cell subset generated during infection plays a pivotal role in boosting EM, which suggests the on-demand reinforcement of EM by adaptive immune cells.