Matsuzawa-Ishimoto et al. show that autophagy gene ATG16L1, which is associated with inflammatory diseases of the gastrointestinal tract, is essential for preventing necroptotic cell death and loss of Paneth cells in the intestinal epithelium.

Disruption of intestinal microbial communities appears to underlie many human illnesses, but the mechanisms that promote this dysbiosis and its adverse consequences are poorly understood. In patients who received allogeneic hematopoietic cell transplantation (allo-HCT), we describe a high incidence of enterococcal expansion, which was associated with graft-versus-host disease (GVHD) and mortality. We found that Enterococcus also expands in the mouse gastrointestinal tract after allo-HCT and exacerbates disease severity in gnotobiotic models. Enterococcus growth is dependent on the disaccharide lactose, and dietary lactose depletion attenuates Enterococcus outgrowth and reduces the severity of GVHD in mice. Allo-HCT patients carrying lactose-nonabsorber genotypes showed compromised clearance of postantibiotic Enterococcus domination. We report lactose as a common nutrient that drives expansion of a commensal bacterium that exacerbates an intestinal and systemic inflammatory disease.

The thymus is extremely sensitive to damage but also has a remarkable ability to repair itself. However, the mechanisms underlying this endogenous regeneration remain poorly understood and this capacity diminishes considerably with age. Here we show that thymic endothelial cells (ECs) comprise a critical pathway of regeneration, via their production of BMP4. ECs increased their production of BMP4 after thymic damage, and abrogating BMP4 signalling or production by either pharmacologic or genetic inhibition impaired thymic repair. EC-derived BMP4 acted on thymic epithelial cells (TECs) to increase their expression of Foxn1, a key transcription factor involved in TEC development, maintenance and regeneration; and its downstream targets such as Dll4, itself a key mediator of thymocyte development and regeneration. These studies demonstrate the importance of the BMP4 pathway in endogenous tissue regeneration and offer a potential clinical approach to enhance T cell immunity.

Bone marrow transplantation (BMT) offers curative potential for patients with high-risk hematologic malignancies, but the post-transplantation period is characterized by profound immunodeficiency. Recent studies indicate that the intestinal microbiota not only regulates mucosal immunity, but can also contribute to systemic immunity and hematopoiesis. Using antibiotic-mediated microbiota depletion in a syngeneic BMT mouse model, here we describe a role for the intestinal flora in hematopoietic recovery after BMT. Depletion of the intestinal microbiota resulted in impaired recovery of lymphocyte and neutrophil counts, while recovery of the hematopoietic stem and progenitor compartments and the erythroid lineage were largely unaffected. Depletion of the intestinal microbiota also reduced dietary energy uptake and visceral fat stores. Caloric supplementation through sucrose in the drinking water improved post-BMT hematopoietic recovery in mice with a depleted intestinal flora. Taken together, we show that the intestinal microbiota contribute to post-BMT hematopoietic reconstitution in mice through improved dietary energy uptake.

A goal in precision medicine is to use patient-derived material to predict disease course and intervention outcomes. Here, we use mechanistic observations in a preclinical animal model to design an ex vivo platform that recreates genetic susceptibility to T-cell–mediated damage. Intestinal graft-versus-host disease (GVHD) is a life-threatening complication of allogeneic hematopoietic cell transplantation. We found that intestinal GVHD in mice deficient in Atg16L1, an autophagy gene that is polymorphic in humans, is reversed by inhibiting necroptosis. We further show that cocultured allogeneic T cells kill Atg16L1-mutant intestinal organoids from mice, which was associated with an aberrant epithelial interferon signature. Using this information, we demonstrate that pharmacologically inhibiting necroptosis or interferon signaling protects human organoids derived from individuals harboring a common ATG16L1 variant from allogeneic T-cell attack. Our study provides a roadmap for applying findings in animal models to individualized therapy that targets affected tissues.

Abstract Nuclear factor erythroid-derived 2-like 2 (Nrf2) is a ubiquitously expressed transcription factor that is well known for its role in regulating the cellular redox pathway. Although there is mounting evidence suggesting a critical role for Nrf2 in hematopoietic stem cells and innate leukocytes, little is known about its involvement in T-cell biology. In this study, we identified a novel role for Nrf2 in regulating alloreactive T-cell function during allogeneic hematopoietic cell transplantation (allo-HCT). We observed increased expression and nuclear translocation of Nrf2 upon T-cell activation in vitro, especially in CD4+ donor T cells after allo-HCT. Allo-HCT recipients of Nrf2−/− donor T cells had significantly less acute graft-versus-host disease (GVHD)-induced mortality, morbidity, and pathology. This reduction in GVHD was associated with the persistence of Helios+ donor regulatory T cells in the allograft, as well as defective upregulation of the gut-homing receptor LPAM-1 on alloreactive CD8+ T cells. Additionally, Nrf2−/− donor CD8+ T cells demonstrated intact cytotoxicity against allogeneic target cells. Tumor-bearing allo-HCT recipients of Nrf2−/− donor T cells had overall improved survival as a result of preserved graft-versus-tumor activity and reduced GVHD activity. Our findings characterized a previously unrecognized role for Nrf2 in T-cell function, as well as revealed a novel therapeutic target to improve the outcomes of allo-HCT.

There is a substantial unmet clinical need for new strategies to protect the hematopoietic stem cell (HSC) pool and regenerate hematopoiesis after radiation injury, either from cancer therapy or accidental exposure1,2. In addition to their role in promoting sexual dimorphisms, increasing evidence suggests that sex hormones regulate HSC self-renewal, differentiation, and proliferation3–5. We and others previously reported that sex steroid ablation promotes bone marrow (BM) lymphopoiesis and HSC recovery in aged and immunodepleted mice5–7. Here we show that a luteinizing hormone-releasing hormone-antagonist (LHRH-Ant), currently used widely in the clinic for sex steroid inhibition, promoted hematopoietic recovery and mouse survival when administered 24 h after an otherwise lethal dose of total body irradiation (L-TBI). Unexpectedly, this protective effect was independent of sex steroids, but instead relied on suppression of luteinizing hormone (LH) levels. Human and mouse long-term self-renewing HSCs (LT-HSCs) expressed high levels of the luteinizing hormone/choriogonadotropin receptor (LHCGR) and expand ex vitro when stimulated with LH. In contrast, suppression of LH after L-TBI inhibited entry of HSCs into the cell cycle, thus promoting quiescence of HSCs and protecting them from exhaustion. These findings reveal a role for LH in regulating HSC function and offer a new therapeutic approach for hematopoietic regeneration after injury.

INTRODUCTION: The intestinal microbiota is essential for the fermentation of fibers into the short-chain fatty acids (SCFA) butyrate, acetate and propionate. SCFA can bind to G-protein-coupled receptors GPR41, GPR43 and GPR109a to activate downstream anti-inflammatory signaling pathways. In colitis or graft versus host disease (GVHD), GPR43 signaling has been reported as an important regulator of intestinal homeostasis by increasing the pool of regulatory T cells. In contrast to GPR43, which binds preferentially propionate and acetate, GPR109a is the major receptor for butyrate. We and others have demonstrated that butyrate can ameliorate gastrointestinal injury during GVHD through enterocyte protection. Therefore, we hypothesized that GPR109a plays an important role in the pathophysiology of intestinal GVHD, focusing specifically on alloreactive T cells. METHODS AND RESULTS: Using mouse models of GVHD, we examined the role of the butyrate/niacin receptor, GPR109a in allogeneic hematopoietic cell transplantation (allo-HCT). First, we studied whether a genetic knock-out (KO) of GPR109a in transplant recipient mice affected GVHD, but GPR109a-KO recipient mice did not exhibit increased mortality from GVHD compared to wild type (WT) mice. We next investigated the role of GPR109a in the donor compartment by transplanting either BM or T cells from WT or GPR109a-KO mice into major MHC mismatched BALB/c host mice. Mice transplanted with B6 BM, with T cells from a GPR109a-KO mouse into BALB/c hosts displayed a lower incidence of lethal GVHD (Fig. 1A). To determine whether the attenuation of GVHD is intrinsic to GPR109a-KO T cells, we established BM chimeras and performed a secondary transplant by transplanting B6 BM + (B6 à Ly5.1) or (GPR109a à Ly5.1) T cells into BALB/c hosts. We observed the same improvement in survival in mice that received GPR109a-KO T cells. This indicates an intrinsic role for GPR109a specifically in the donor hematopoietic compartment. Having identified a T-cell specific requirement for GPR109a we next examined expression of GPR109a on WT T cells in vitro at baseline and following stimulation with CD3/28 and found GPR109a significantly upregulated on both CD4+ and CD8+ T cells after 72 hours of stimulation (Fig 1B). At steady state in vivo, we observed the same numbers and percentages of splenic effector memory, central memory, and naïve CD4+ T cells as well as regulatory T cells in WT B6 mice and GPR109a-KO mice, suggesting normal T cell development in the knockout mice. In an in vitro mixed lymphocyte reaction (MLR), GPR109a-KO CD4+ T cells become activated, proliferate, polarize and secrete cytokine (specifically IFNg) to the same level as WT CD4+ T cells, suggesting normal functional capacity. However, after allo-HCT in mice we observed significantly fewer CD4+ and CD8+ T cells, and specifically fewer effector memory CD4+ T cells (Fig. C), in the small and large intestines of mice that received GPR109a-KO T cells at day 7 post transplant. In contrast, we found significantly more regulatory T cells in the intestines (Fig. 1D) and the spleen of GPR1091-KO T cell recipients, while numbers and percentages of polarized Th1 and Th17 T cells were similar between the two groups. We further 16S rRNA sequenced the gut microbiota of mice at day 7 after transplant and observed an increased relative abundance of bacteria from the genus Clostridium (Fig. 1D) along with an increased concentration of cecal butyrate as measured by GC-MS (Fig. 1E). In a preliminary graft versus tumor (GVT) experiment, we found that mice that received A20 tumor cells and GPR109a-KO T cells exhibited increased survival compared to mice that received A20 tumor cells and WT T cells. These preliminary findings suggest that GPR109a-KO T cells maintain their graft versus tumor response while causing less GVHD, and exclude a defective functional capacity. CONCLUSIONS: We report a novel role of the butyrate/niacin receptor GPR109a on donor T cells in allo-HCT as a genetic knock-out on T cells attenuates lethal GVHD. As these T cells are tested as functionally intact, we propose that the reduction in overall T cells of KO T cell recipients may underlie the attenuation in GVHD. Furthermore, such a reduction in allograft-induced gut injury is accompanied by maintenance of the gut commensal Clostridium and butyrate production, which is known to protect the intestinal epithelium and increases the regulatory T cell pool. Disclosures No relevant conflicts of interest to declare.