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The metal ion transporter SLC39A8 is associated with physiological traits and diseases, including blood manganese (Mn) levels and inflammatory bowel diseases (IBD). The mechanisms by which SLC39A8 controls Mn homeostasis and epithelial integrity remain elusive. Here, we generate Slc39a8 intestinal epithelial cell-specific-knockout ( Slc39a8- IEC KO) mice, which display markedly decreased Mn levels in blood and most organs. Radiotracer studies reveal impaired intestinal absorption of dietary Mn in Slc39a8- IEC KO mice. SLC39A8 is localized to the apical membrane and mediates 54 Mn uptake in intestinal organoid monolayer cultures. Unbiased transcriptomic analysis identifies alkaline ceramidase 1 (ACER1), a key enzyme in sphingolipid metabolism, as a potential therapeutic target for SLC39A8-associated IBDs. Importantly, treatment with an ACER1 inhibitor attenuates colitis in Slc39a8- IEC KO mice by remedying barrier dysfunction. Our results highlight the essential roles of SLC39A8 in intestinal Mn absorption and epithelial integrity and offer a therapeutic target for IBD associated with impaired Mn homeostasis. The metal transporter SLC39A8 is linked to blood manganese (Mn) levels and inflammatory bowel diseases (IBD). Here, the authors show that loss of SLC39A8 impairs intestinal Mn absorption and epithelial integrity, suggesting potential therapeutic strategies targeting alkaline ceramidase 1 for IBD patients with SLC39A8 deficiency.

The cellular microenvironment required to sustain adult intestinal stem cells has long been controversial. Cells that release proteins needed for intestinal-tissue renewal have now been defined. The cellular microenvironment required to sustain adult intestinal stem cells has long been controversial. Cells that release proteins needed for intestinal-tissue renewal have now been defined.

ObjectiveLrig1 is a marker of proliferative and quiescent stem cells in the skin and intestine. We examined whether Lrig1-expressing cells are long-lived gastric progenitors in gastric glands in the mouse stomach. We also investigated how the Lrig1-expressing progenitor cells contribute to the regeneration of normal gastric mucosa by lineage commitment to parietal cells after acute gastric injury in mice.DesignWe performed lineage labelling using Lrig1-CreERT2/+;R26R-YFP/+ (Lrig1/YFP) or R26R-LacZ/+ (Lrig1/LacZ) mice to examine whether the Lrig1-YFP-marked cells are gastric progenitor cells. We studied whether Lrig1-YFP-marked cells give rise to normal gastric lineage cells in damaged mucosa using Lrig1/YFP mice after treatment with DMP-777 to induce acute injury. We also studied Lrig1-CreERT2/CreERT2 (Lrig1 knockout) mice to examine whether the Lrig1 protein is required for regeneration of gastric corpus mucosa after acute injury.ResultsLrig1-YFP-marked cells give rise to gastric lineage epithelial cells both in the gastric corpus and antrum, in contrast to published results that Lgr5 only marks progenitor cells within the gastric antrum. Lrig1-YFP-marked cells contribute to replacement of damaged gastric oxyntic glands during the recovery phase after acute oxyntic atrophy in the gastric corpus. Lrig1 null mice recovered normally from acute gastric mucosal injury indicating that Lrig1 protein is not required for lineage differentiation. Lrig1+ isthmal progenitor cells did not contribute to transdifferentiating chief cell lineages after acute oxyntic atrophy.ConclusionsLrig1 marks gastric corpus epithelial progenitor cells capable of repopulating the damaged oxyntic mucosa by differentiating into normal gastric lineage cells in mouse stomach.

Graft-versus-host disease (GVHD) is the main complication of allogeneic bone marrow transplantation. Current strategies to control GVHD rely on global immunosuppression. These strategies are incompletely effective and decrease the anticancer activity of the allogeneic graft. We previously identified Notch signaling in T cells as a new therapeutic target for preventing GVHD. Notch-deprived T cells showed markedly decreased production of inflammatory cytokines, but normal in vivo proliferation, increased accumulation of regulatory T cells, and preserved anticancer effects. Here, we report that γ-secretase inhibitors can block all Notch signals in alloreactive T cells, but lead to severe on-target intestinal toxicity. Using newly developed humanized antibodies and conditional genetic models, we demonstrate that Notch1/Notch2 receptors and the Notch ligands Delta-like1/4 mediate all the effects of Notch signaling in T cells during GVHD, with dominant roles for Notch1 and Delta-like4. Notch1 inhibition controlled GVHD, but led to treatment-limiting toxicity. In contrast, Delta-like1/4 inhibition blocked GVHD without limiting adverse effects while preserving substantial anticancer activity. Transient blockade in the peritransplant period provided durable protection. These findings open new perspectives for selective and safe targeting of individual Notch pathway components in GVHD and other T cell-mediated human disorders.

Gastrin is secreted following a rise in gastric pH, leading to gastric acid secretion. Sleeve gastrectomy (SG), a bariatric surgery where 80% of the gastric corpus is excised, presents a challenge for gastric pH homeostasis. Using histology, and single-cell RNA sequencing of the gastric epithelium in 12 women, we observed that SG is associated with an increase in a sub-population of acid-secreting parietal cells that overexpress respiratory enzymes and an increase in histamine-secreting enterochromaffin-like cells (ECLs). ECLs of SG-operated patients overexpressed genes coding for biosynthesis of neuropeptides and serotonin. Mathematical modeling showed that pH homeostasis by gastrin is analogous to non-linear proportional and integral control, that drives adaptation of the epithelium to acid-secretion demand. Quantitative model predictions were validated in patients. The results demonstrate human gastric epithelium remodeling following SG at the molecular and cellular levels, and more generally how trophic hormones enable robust adaptation of tissue function to meet physiological demand. Sleeve gastrectomy is a common bariatric surgery in which most of the gastric corpus is removed. Here, the authors show the adaptation of the gastric mucosa to surgery in patients, and how it facilitates maintenance of gastric pH homeostasis through a proportional-integral feedback control.

ObjectiveWe tested the ability of Notch pathway receptors Notch1 and Notch2 to regulate stem and epithelial cell homoeostasis in mouse and human gastric antral tissue.DesignMice were treated with the pan-Notch inhibitor dibenzazepine (DBZ) or inhibitory antibodies targeting Notch1 and/or Notch2. Epithelial proliferation, apoptosis and cellular differentiation were measured by histological and molecular approaches. Organoids were established from mouse and human antral glands; growth and differentiation were measured after treatment with Notch inhibitors.ResultsNotch1 and Notch2 are the predominant Notch receptors expressed in mouse and human antral tissue and organoid cultures. Combined inhibition of Notch1 and Notch2 in adult mice led to decreased epithelial cell proliferation, including reduced proliferation of LGR5 stem cells, and increased apoptosis, similar to the response to global Notch inhibition with DBZ. Less pronounced effects were observed after inhibition of individual receptors. Notch pathway inhibition with DBZ or combined inhibition of Notch1 and Notch2 led to increased differentiation of all gastric antral lineages, with remodelling of cells to express secretory products normally associated with other regions of the GI tract, including intestine. Analysis of mouse and human organoids showed that Notch signalling through Notch1 and Notch2 is intrinsic to the epithelium and required for organoid growth.ConclusionsNotch signalling is required to maintain gastric antral stem cells. Notch1 and Notch2 are the primary Notch receptors regulating epithelial cell homoeostasis in mouse and human stomach.

Epithelial tuft cells are named after their characteristic microtubule bundles located at the cell apex where these are exposed to the luminal environment. As such, tuft cells are found in multiple organs, including the gastrointestinal (GI) tract where the apical “tuft” is hypothesized to detect and transmit environmental signals. Thus, the goal of our study was to characterize gastric tuft cells during GI tract development, then subsequently in the normal and metaplastic adult stomach. GI tracts from mouse embryos, and newborn and postnatal mice were analyzed. Tuft cells were identified by immunohistochemistry using acetylated-α-tubulin (acTub) antibody to detect the microtubule bundle. Additional tuft cell markers, e.g., doublecortin-like kinase 1 (DCLK1), were used to co-localize with acTub. Tuft cells were quantified in human gastric tissue arrays and in mouse stomachs with or without inflammation. In the developing intestine, tuft cells in both the crypts and villi expressed all markers by E18.5. In the stomach, acTub co-localized with DCLK1 and other established tuft cell markers by E18.5 in the antrum, but not until postnatal day 7 in the corpus, with the highest density of tuft cells clustered at the forestomach ridge. Tuft cell numbers increased in hyperplastic human and mouse stomachs. In the adult GI tract, the tuft cell marker acTub co-expressed with DCKL1 and chemosensory markers, e.g.,TRPM5. In summary, tuft cells appear in the gastric antrum and intestine at E18.5, but their maximal numbers in the corpus are not achieved until after weaning. Tuft cell numbers increase with inflammation, hyperplasia, and metaplasia.

Gastrointestinal infection is often associated with hypophagia and weight loss; however, the precise mechanisms governing these responses remain poorly defined. Furthermore, the possibility that alterations in feeding during infection may be beneficial to the host requires further study. We used the nematode Trichinella spiralis, which transiently inhabits the small intestine before migrating to skeletal muscle, as a biphasic model of infection to determine the cellular and molecular pathways controlling feeding during enteric and peripheral inflammation. Through the infection of genetically modified mice lacking cholecystokinin, Tumor necrosis factor α receptors and T and B-cells, we observed a biphasic hypophagic response to infection resulting from two separate immune-driven mechanisms. The enteroendocrine I-cell derived hormone cholecystokinin is an essential mediator of initial hypophagia and is induced by CD4+ T-cells during enteritis. In contrast, the second hypophagic response is extra-intestinal and due to the anorectic effects of TNFα during peripheral infection of the muscle. Moreover, via maintaining naive levels of the adipose secreted hormone leptin throughout infection we demonstrate a novel feedback loop in the immunoendocrine axis. Immune driven I-cell hyperplasia and resultant weight loss leads to a reduction in the inflammatory adipokine leptin, which in turn heightens protective immunity during infection. These results characterize specific immune mediated mechanisms which reduce feeding during intestinal or peripheral inflammation. Importantly, the molecular mediators of each phase are entirely separate. The data also introduce the first evidence that I-cell hyperplasia is an adaptively driven immune response that directly impinges on the outcome to infection.

Biliary obstruction and cholangiocyte hyperproliferation are important features of cholangiopathies affecting the large extrahepatic bile duct (EHBD). The mechanisms underlying obstruction-induced cholangiocyte proliferation in the EHBD remain poorly understood. Developmental pathways, including WNT signaling, are implicated in regulating injury responses in many tissues, including the liver. To investigate the contribution of WNT signaling to obstruction-induced cholangiocyte proliferation in the EHBD, we used complementary in vivo and in vitro models with pharmacologic interventions and transcriptomic analyses. To model obstruction, we used bile duct ligation (BDL) in mice. Human and mouse biliary organoids and mouse biliary explants were used to investigate the effects of WNT activation and inhibition in vitro. We observed an upregulation of WNT ligand expression associated with increased biliary proliferation following obstruction. Cholangiocytes were identified as both WNT ligand–expressing and WNT-responsive cells. Inhibition of WNT signaling decreased cholangiocyte proliferation in vivo and in vitro, while activation increased proliferation. WNT effects on cholangiocyte proliferation were β-catenin dependent, and we showed a direct effect of WNT7B on cholangiocyte growth. Our studies suggested that cholangiocyte-derived WNT ligands can activate WNT signaling to induce proliferation after obstructive injury. These findings implicate the WNT pathway in injury-induced cholangiocyte proliferation within the EHBD.

Germline adenomatous polyposis coli ( APC ) mutation in patients with familial adenomatous polyposis (FAP) promotes gastrointestinal polyposis, including the formation of frequent gastric fundic gland polyps (FGPs). In this study, we investigated how dysregulated Wnt signaling promotes FGPs and why they localize to the corpus region of the stomach. We developed a biobank of FGP and surrounding nonpolyp corpus biopsies and organoids from patients with FAP for comparative studies. Polyp biopsies and polyp-derived organoids exhibited enhanced Wnt target gene expression. Polyp-derived organoids with intrinsically upregulated Wnt signaling showed poor tolerance to further induction, suggesting that high Wnt restricts growth. Targeted genomic sequencing revealed that most gastric polyps did not arise via APC loss of heterozygosity. Studies in genetic mouse models demonstrated that heterozygous Apc loss increased epithelial cell proliferation in the corpus but not the antrum, while homozygous Apc loss was not maintained in the corpus yet induced hyperproliferation in the antrum. Our findings suggest that heterozygous APC mutation in patients with FAP may be sufficient to drive polyp formation in the corpus region while subsequent loss of heterozygosity to further enhance Wnt signaling is not tolerated. This finding contextualizes the abundant yet benign nature of gastric polyps in FAP patient corpus compared with the rare, yet adenomatous polyps in the antrum.