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The esophagus and stomach, joined by a unique transitional zone, contain actively dividing epithelial stem cells required for organ homeostasis. Upon prolonged inflammation, epithelial cells in both organs can undergo a cell fate switch leading to intestinal metaplasia, predisposing to malignancy. Here we discuss the biology of gastroesophageal stem cells and their role as cells of origin in cancer. We summarize the interactions between the stromal niche and gastroesophageal stem cells in metaplasia and early expansion of mutated stem-cell-derived clones during carcinogenesis. Finally, we review new approaches under development to better study gastroesophageal stem cells and advance the field.

Background and Aim Proton pump inhibitors (PPIs) are a potential cause of gastric carcinogenesis after Helicobacter pylori eradication. Thus, appropriate management including chemoprevention is required. The aim of this study was to evaluate the association between nonsteroidal anti‐inflammatory drugs (NSAIDs) and the incidence of post‐eradication gastric cancer in PPI users. Methods A multicenter retrospective cohort study was conducted. Patients who used a PPI (≥30 days) after H. pylori eradication between 2014 and 2019 were analyzed in nine hospital databases. Gastric cancer incidence was a primary outcome, and their association with NSAIDs use and clinical factors was evaluated. Hazard ratios were adjusted by age, sex, smoking, and Charlson Comorbidity Index. Results During the mean follow‐up period of 2.38 years, 1.13% (31/2431) of all patients developed gastric cancer. The cumulative incidence of gastric cancer in PPI users was 0.25% at 1 year, 0.51% at 3 years, and 1.09% at 5 years in the NSAID users and 0.89% at 1 year, 2.32% at 3 years, and 3.61% at 5 years in nonusers. NSAIDs were associated with a lower gastric cancer risk (adjusted hazard ratio = 0.28, P = 0.005). No gastric cancer was observed in the cyclooxygenase‐2 inhibitor users (n = 256). NSAID use with high dose and long duration was significantly associated with a lower incidence of gastric cancer. Conclusion NSAIDs were associated with a 60% decrease in the gastric cancer incidence in H. pylori‐eradicated PPI users, with dose and duration response effects. NSAIDs may be effective for chemoprevention against PPI‐related gastric cancer.

The stem/progenitor cells of the developing intestine are biologically distinct from their adult counterparts. Here, we examine the microenvironmental cues that regulate the embryonic stem/progenitor population, focusing on the role of Notch pathway factor delta-like protein-1 (DLK1). mRNA-seq analyses of intestinal mesenchymal cells (IMCs) collected from embryonic day 14.5 (E14.5) or adult IMCs and a novel coculture system with E14.5 intestinal epithelial organoids were used. Following addition of recombinant DLK1 (rDLK) or

Background & Aims Immune checkpoint inhibitors have limited efficacy in many tumors. We investigated mechanisms of tumor resistance to inhibitors of programmed cell death–1 (PDCD1, also called PD-1) in mice with gastric cancer, and the role of its ligand, PD-L1. Methods Gastrin-deficient mice were given N-methyl-N-nitrosourea (MNU) in drinking water along with Helicobacter felis to induce gastric tumor formation; we also performed studies with H/K-ATPase-hIL1B mice, which develop spontaneous gastric tumors at the antral–corpus junction and have parietal cells that constitutively secrete interleukin 1B. Mice were given injections of an antibody against PD-1 or an isotype control before tumors developed, or anti–PD-1 and 5-fluorouracil and oxaliplatin, or an antibody against lymphocyte antigen 6 complex locus G (also called Gr-1), which depletes myeloid-derived suppressor cells [MDSCs]), after tumors developed. We generated knock-in mice that express PD-L1 specifically in the gastric epithelium or myeloid lineage. Results When given to gastrin-deficient mice before tumors grew, anti–PD-1 significantly reduced tumor size and increased tumor infiltration by T cells. However, anti–PD-1 alone did not have significant effects on established tumors in these mice. Neither early nor late anti–PD-1 administration reduced tumor growth in the presence of MDSCs in H/K-ATPase-hIL-1β mice. The combination of 5-fluorouracil and oxaliplatin reduced MDSCs, increased numbers of intra-tumor CD8+ T cells, and increased the response of tumors to anti–PD-1; however, this resulted in increased tumor expression of PD-L1. Expression of PD-L1 by tumor or immune cells increased gastric tumorigenesis in mice given MNU. Mice with gastric epithelial cells that expressed PD-L1 did not develop spontaneous tumors, but they developed more and larger tumors after administration of MNU and H felis, with accumulation of MDSCs. Conclusions In mouse models of gastric cancer, 5-fluorouracil and oxaliplatin reduce numbers of MDSCs to increase the effects of anti–PD-1, which promotes tumor infiltration by CD8+ T cells. However, these chemotherapeutic agents also induce expression of PD-L1 by tumor cells. Expression of PD-L1 by gastric epithelial cells increases tumorigenesis in response to MNU and H felis, and accumulation of MDSCs, which promote tumor progression. The timing and site of PD-L1 expression is therefore important in gastric tumorigenesis and should be considered in design of therapeutic regimens.

Background and aimsThe gastric epithelium undergoes continuous turnover. Corpus epithelial stem cells located in the gastric isthmus serve as a source of tissue self-renewal. We recently identified the transcription factor Mist1 as a marker for this corpus stem cell population that can give rise to cancer. The aim here was to investigate the regulation of the Mist1+ stem cells in the response to gastric injury and inflammation.MethodsWe used Mist1CreERT;R26-Tdtomato mice in two models of injury and inflammation: the acetic acid-induced ulcer and infection with Helicobacter felis. We analysed lineage tracing at both early (7 to 30 days) and late (30 to 90 days) time points. Mist1CreERT;R26-Tdtomato;Lgr5DTR-eGFP mice were used to ablate the corpus basal Lgr5+ cell population. Constitutional and conditional Wnt5a knockout mice were used to investigate the role of Wnt5a in wound repair and lineage tracing from the Mist1+ stem cells.ResultsIn both models of gastric injury, Mist1+ isthmus stem cells more rapidly proliferate and trace entire gastric glands compared with the normal state. In regenerating tissue, the number of traced gastric chief cells was significantly reduced, and ablation of Lgr5+ chief cells did not affect Mist1-derived lineage tracing and tissue regeneration. Genetic deletion of Wnt5a impaired proliferation in the gastric isthmus and lineage tracing from Mist1+ stem cells. Similarly, depletion of innate lymphoid cells, the main source of Wnt5a, also resulted in reduced proliferation and Mist1+ isthmus cell tracing.ConclusionGastric Mist1+ isthmus cells are the main supplier of regenerated glands and are activated in part through Wnt5a pathway.

The tumor microenvironment favors the growth and expansion of cancer cells. Many cell types are involved in the tumor microenvironment such as inflammatory cells, fibroblasts, nerves, and vascular endothelial cells. These stromal cells contribute to tumor growth by releasing various molecules to either directly activate the growth signaling in cancer cells or remodel surrounding areas. This review introduces recent advances in findings on the interactions within the tumor microenvironment such as in cancer‐associated fibroblasts (CAFs), immune cells, and endothelial cells, in particular those established in mouse gastric cancer models. In mice, myofibroblasts in the gastric stroma secrete R‐spondin and support normal gastric stem cells. Most CAFs promote tumor growth in a paracrine manner, but CAF population appears to be heterogeneous in terms of their function and origin, and include both tumor‐promoting and tumor‐restraining populations. Among immune cell populations, tumor‐associated macrophages, including M1 and M2 macrophages, and myeloid‐derived suppressor cells (MDSCs), are reported to directly or indirectly promote gastric tumorigenesis by secreting soluble factors or modulating immune responses. Endothelial cells or blood vessels not only fuel tumors with nutrients, but also interact with cancer stem cells and immune cells by secreting chemokines or cytokines, and act as a cancer niche. Understanding these interactions within the tumor microenvironment would contribute to unraveling new therapeutic targets., Gastric tumor microenvironment: Cancer‐associated fibroblasts, endothelial cells, gastrin‐expressing cells, and various immune cells including macrophages, MDSCs, and ILC2s serve as tumor‐promoting niche in gastric cancers. There are numerous crosstalks between tumor cells and surrounding stromal cell types, which contribute to tumor development derived from gastric stem cells.

The enteric neurotransmitter acetylcholine governs important intestinal epithelial secretory and immune functions through its actions on epithelial muscarinic Gq-coupled receptors such as M3R. Its role in the regulation of intestinal stem cell function and differentiation, however, has not been clarified. Here, we find that nonselective muscarinic receptor antagonism in mice as well as epithelial-specific ablation of M3R induces a selective expansion of DCLK1-positive tuft cells, suggesting a model of feedback inhibition. Cholinergic blockade reduces Lgr5-positive intestinal stem cell tracing and cell number. In contrast, Prox1-positive endocrine cells appear as primary sensors of cholinergic blockade inducing the expansion of tuft cells, which adopt an enteroendocrine phenotype and contribute to increased mucosal levels of acetylcholine. This compensatory mechanism is lost with acute irradiation injury, resulting in a paucity of tuft cells and acetylcholine production. Thus, enteroendocrine tuft cells appear essential to maintain epithelial homeostasis following modifications of the cholinergic intestinal niche., Acetylcholine regulates intestinal epithelial secretion via muscarinic Gq-coupled receptors but its role in cell differentiation is unclear. Here, the authors show that Prox1-positive endocrine cells are sensors for the cholinergic intestinal niche and can trigger increased differentiation of enteroendocrine DCLK1-positive tuft cells.

Background & Aims Enterochromaffin-like (ECL) cells in the stomach express gastrin/cholecystokinin 2 receptor CCK2R and are known to expand under hypergastrinemia, but whether this results from expansion of existing ECL cells or increased production from progenitors has not been clarified. Methods We used mice with green fluorescent protein fluorescent reporter expression in ECL cells (histidine decarboxylase [Hdc]-green fluorescent protein), as well as Cck2r- and Hdc-driven Tamoxifen inducible recombinase Cre (Cck2r-CreERT2, Hdc-CreERT2) mice combined with Rosa26Sor-tdTomato (R26-tdTomato) mice, and studied their expression and cell fate in the gastric corpus by using models of hypergastrinemia (gastrin infusion, omeprazole treatment). Results Hdc-GFP marked the majority of ECL cells, located in the lower third of the gastric glands. Hypergastrinemia led to expansion of ECL cells that was not restricted to the gland base, and promoted cellular proliferation (Ki67) in the gastric isthmus but not in basal ECL cells. Cck2r-CreERT2 mice marked most ECL cells, as well as scattered cell types located higher up in the glands, whose number was increased during hypergastrinemia. Cck2r-CreERT2+ isthmus progenitors, but not Hdc+ mature ECL cells, were the source of ECL cell hyperplasia during hypergastrinemia and could grow as 3-dimensional spheroids in vitro. Moreover, gastrin treatment in vitro promoted sphere formation from sorted Cck2r+Hdc- cells, and increased chromogranin A and phosphorylated- extracellular signal-regulated kinase expression in CCK2R-derived organoids. Gastrin activates extracellular signal-regulated kinase pathways in vivo and in vitro, and treatment with the Mitogen-activated protein kinase kinase 1 inhibitor U0126 blocked hypergastrinemia-mediated changes, including CCK2R-derived ECL cell hyperplasia in vivo as well as sphere formation and chromogranin A expression in vitro. Conclusions We show here that hypergastrinemia induces ECL cell hyperplasia that is derived primarily from CCK2R+ progenitors in the corpus. Gastrin-dependent function of CCK2R+ progenitors is regulated by the extracellular signal-regulated kinase pathway.

Background/Aim: The entire mechanisms by which epigenetic modifiers contribute to the development of pancreatic cancer remain unknown. Although the histone methyltransferase G9a is a promising target in human cancers, its role in pancreatic carcinogenesis has been under-studied. The aim of the study was to examine the role of G9a in pancreatic carcinogenesis by a gene-targeting mouse model. Materials and Methods: We established pancreas-specific G9a(flox/flox )mice and crossed them with Ptf1a(Cre/); Kras(G12D/+ )(KC) mice, which spontaneously develop pancreatic cancer. The phenotypes of the resulting KC mice with G9a deletion were examined. We analyzed transcriptomic data by microarray and genome-wide chromatin accessibility by transposase-accessible chromatin using sequencing. We established pancreatic organoids from KC mice. Results: G9a deficiency impaired the progression of pancreatic intraepithelial neoplasia (PanIN) and prolonged the survival of KC mice. The number of phosphorylated Erk-positive cells and Dclk1-positive cells, which are reported to be essential for the progression of PanIN, were decreased by G9a deletion. UNC0638, an inhibitor of G9a, suppressed the growth of organoids and increased global chromatin accessibility, especially around the regions including the protein phosphatase 2A genes. Conclusion: Thus, our study suggested the functional interaction of G9a, Dclk1 and Mapk pathway in the Kras-driven pancreatic carcinogenesis. The inhibition of G9a may suppress the initiation of oncogenic Kras-driven pancreatic carcinogenesis.

Background & Aims Liver lobules are typically subdivided into 3 metabolic zones: zones 1, 2, and 3. However, the contribution of zonal differences in hepatocytes to liver regeneration, as well as to carcinogenic susceptibility, remains unclear. Methods We developed a new method for sustained genetic labelling of zone 3 hepatocytes and performed fate tracing to monitor these cells in multiple mouse liver tumour models. Results We first examined changes in the zonal distribution of the Wnt target gene Axin2 over time using Axin2-CreERT2;Rosa26-Lox-Stop-Lox-tdTomato mice (Axin2;tdTomato). We found that following tamoxifen administration at 3 weeks of age, approximately one-third of total hepatocytes that correspond to zone 3 were labelled in Axin2;tdTomato mice; the tdTomato+ cell distribution closely matched that of the zone 3 marker CYP2E1. Cell fate analysis revealed that zone 3 hepatocytes maintained their own lineage but rarely proliferated beyond their liver zonation during homoeostasis; this indicated that our protocol enabled persistent genetic labelling of zone 3 hepatocytes. Using this system, we found that zone 3 hepatocytes generally had high neoplastic potential, which was promoted by constitutive activation of Wnt/β-catenin signalling in the pericentral area. However, the frequency of zone 3 hepatocyte-derived tumours varied depending on the regeneration pattern of the liver parenchyma in response to liver injury. Notably, Axin2-expressing hepatocytes undergoing chronic liver injury significantly contributed to liver regeneration and possessed high neoplastic potential. Additionally, we revealed that the metabolic phenotypes of liver tumours were acquired during tumorigenesis, irrespective of their spatial origin. Conclusions Hepatocytes receiving Wnt/β-catenin signalling from their microenvironment have high neoplastic potential, and Wnt/β-catenin signalling is a potential drug target for the prevention of hepatocellular carcinoma. Lay summary Lineage tracing revealed that zone 3 hepatocytes residing in the pericentral niche have high neoplastic potential. Under chronic liver injury, hepatocytes receiving Wnt/β-catenin signalling broadly exist across all hepatic zones and significantly contribute to liver tumorigenesis as well as liver regeneration. Wnt/β-catenin signalling is a potential drug target for the prevention of hepatocellular carcinoma., Graphical abstract, Highlights • We developed a new method for sustained genetic labelling of Zone 3 hepatocytes. • Lineage tracing revealed that Zone 3 hepatocytes generally have high neoplastic potential. • The frequency of Zone 3 hepatocyte-derived tumours varied depending on the regeneration pattern of liver parenchyma. • Under chronic liver injury, hepatocytes receiving Wnt/β-catenin signalling significantly contributed to tumorigenesis. • Wnt/β-catenin signalling is a potential drug target for the prevention of HCC.

Simple Summary Helicobacter pylori is the most important cause of gastric cancer, and its eradication reduces the incidence of gastric cancer after endoscopic resection. However, incidence of metachronous gastric cancer is still high. More studies are needed to identify other chemopreventive drugs that may reduce the incidence of this disease. In this study, we focused on the alteration of the intragastric microbiome and examined the association between the use of antibiotics and probiotic drugs and risk of metachronous gastric cancer. Our findings suggest that the gut microbiome is associated with metachronous gastric cancer development. Abstract Metachronous gastric cancer often occurs after endoscopic resection. Appropriate management, including chemoprevention, is required after the procedure. This study was performed to evaluate the association between medication use and the incidence of metachronous gastric cancer after endoscopic resection. This multicenter retrospective cohort study was conducted with data from nine hospital databases on patients who underwent endoscopic resection for gastric cancer between 2014 and 2019. The primary outcome was the incidence of metachronous gastric cancer. We evaluated the associations of metachronous gastric cancer occurrence with medication use and clinical factors. Hazard ratios were adjusted by age and Charlson comorbidity index scores, with and without consideration of sex, smoking status, and receipt of Helicobacter pylori eradication therapy during the study period. During a mean follow-up period of 2.55 years, 10.39% (140/1347) of all patients developed metachronous gastric cancer. The use of antibiotics other than those used for H. pylori eradication was associated with a lower incidence of metachronous gastric cancer than was non-use (adjusted hazard ratio (aHR) 0.56, 95% confidence interval (CI) 0.38–0.85, p = 0.006). Probiotic drug use was also associated with a lower incidence of metachronous gastric cancer compared with non-use (aHR 0.29, 95% CI 0.091–0.91, p = 0.034). In conclusion, the use of antibiotics and probiotic drugs was associated with a decreased risk of metachronous gastric cancer. These findings suggest that the gut microbiome is associated with metachronous gastric cancer development.

Chronic inflammation and intestinal metaplasia are strongly associated with gastric carcinogenesis. Kras activation and Pten deletion are observed in intestinal-type gastric cancer, and Cdh1 mutation is associated with diffuse-type gastric cancer. Although various mouse models of gastric carcinogenesis have been reported, few mouse lines enable gene manipulation selectively in the stomach. Here we established a Tff1-Cre bacterial artificial chromosome transgenic mouse line in an attempt to induce gene modification specifically in the gastric pit lineage. In the stomach, Tff1-Cre-mediated recombination was most evident in the pit lineage in the corpus and in entire antral glands; recombination was also observed in a few gastric chief and parietal cells. Outside the stomach, recombination was patchy throughout the intestines, and particularly frequently in the duodenum (Brunner glands), cecum, and proximal colon. In the stomachs of Tff1-Cre;LSL-KrasG12D mice, proliferating cell clusters expanded throughout the corpus glands, with foveolar cell expansion with ectopic Alcian blue-positive mucins, oxyntic atrophy, and pseudopyloric changes with spasmolytic polypeptide-expressing metaplasia; however, gastric cancer was not observed even at 12 months of age. Corpus-derived organoids from Tff1-Cre;LSL-KrasG12D mice exhibited accelerated growth and abnormal differentiation with a loss of chief and parietal cell markers. Tff1-Cre;Ptenflox/flox mice displayed similar changes to those seen in Tff1-Cre;LSL-KrasG12D mice, both with aberrant ERK activation within 3 months. In contrast, Tff1-Cre;Cdh1flox/flox mice initially showed signet ring-like cells that were rapidly lost with disruption of the mucosal surface, and later developed gastric epithelial shedding with hyperproliferation and loss of normal gastric lineages. Eventually, the glandular epithelium in Tff1-Cre;Cdh1flox/flox mice was completely replaced by squamous epithelium which expanded from the forestomach. Tff1-Cre mice offer an additional useful tool for studying gastric carcinogenesis both in vivo and in vitro. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

The intestinal stroma provides an important microenvironment for immune cell activation. The perturbation of this tightly regulated process can lead to excessive inflammation. We know that upregulated Toll-like receptor 4 (TLR4) in the intestinal epithelium plays a key role in the inflammatory condition of preterm infants, such as necrotizing enterocolitis (NEC). However, the surrounding stromal contribution to excessive inflammation in the pre-term setting awaits careful dissection. Ex vivo co-culture of embryonic day 14.5 (E14.5) or adult murine intestinal stromal cells with exogenous monocytes was undertaken. We also performed mRNAseq analysis of embryonic and adult stromal cells treated with vehicle control or lipopolysaccharide (LPS), followed by pathway and network analyses of differentially regulated transcripts. Cell characteristics were compared using flow cytometry and pHrodo red phagocytic stain, candidate gene analysis was performed via siRNA knockdown and gene expression measured by qPCR and ELISA. Embryonic stromal cells promote the differentiation of co-cultured monocytes to CD11bhighCD11chigh mononuclear phagocytes, that in turn express decreased levels of CD103. Global mRNAseq analysis of stromal cells following LPS stimulation identified TLR signaling components as the most differentially expressed transcripts in the immature compared to adult setting. We show that CD14 expressed by CD11b+CD45+ embryonic stromal cells is a key inducer of TLR mediated inflammatory cytokine production and phagocytic activity of monocyte derived cells. We utilise transcriptomic analyses and functional ex vivo modelling to improve our understanding of unique molecular cues provided by the immature intestinal stroma.

The mechanism of H. pylori-induced atrophy and metaplasia has not been fully understood. Here, we demonstrate the novel role of Apoptosis signal-regulating kinase 1 (ASK1) and downstream MAPKs as a regulator of host immune responses and epithelial maintenance against H. pylori infection. ASK1 gene deficiency resulted in enhanced inflammation with numerous inflammatory cells including Gr-1+CD11b+ myeloid-derived suppressor cells (MDSCs) recruited into the infected stomach. Increase of IL-1β release from apoptotic macrophages and enhancement of TH1-polarized immune responses caused STAT1 and NF-κB activation in epithelial cells in ASK1 knockout mice. Dysregulated immune and epithelial activation in ASK1 knockout mice led to dramatic expansion of gastric progenitor cells and massive metaplasia development. Bone marrow transplantation experiments revealed that ASK1 in inflammatory cells is critical for inducing immune disorder and metaplastic changes in epithelium, while ASK1 in epithelial cells regulates cell proliferation in stem/progenitor zone without changes in inflammation and differentiation. These results suggest that H. pylori-induced immune cells may regulate epithelial homeostasis and cell fate as an inflammatory niche via ASK1 signaling.

Background & Aims Cancer-associated fibroblasts (CAFs), key constituents of the tumor microenvironment, either promote or restrain tumor growth. Attempts to therapeutically target CAFs have been hampered by our incomplete understanding of these functionally heterogeneous cells. Key growth factors in the intestinal epithelial niche, bone morphogenetic proteins (BMPs), also play a critical role in colorectal cancer (CRC) progression. However, the crucial proteins regulating stromal BMP balance and the potential application of BMP signaling to manage CRC remain largely unexplored. Methods Using human CRC RNA expression data, we identified CAF-specific factors involved in BMP signaling, then verified and characterized their expression in the CRC stroma by in situ hybridization. CRC tumoroids and a mouse model of CRC hepatic metastasis were used to test approaches to modify BMP signaling and treat CRC. Results We identified Grem1 and Islr as CAF-specific genes involved in BMP signaling. Functionally, GREM1 and ISLR acted to inhibit and promote BMP signaling, respectively. Grem1 and Islr marked distinct fibroblast subpopulations and were differentially regulated by transforming growth factor β and FOXL1, providing an underlying mechanism to explain fibroblast biological dichotomy. In patients with CRC, high GREM1 and ISLR expression levels were associated with poor and favorable survival, respectively. A GREM1-neutralizing antibody or fibroblast Islr overexpression reduced CRC tumoroid growth and promoted Lgr5+ intestinal stem cell differentiation. Finally, adeno-associated virus 8 (AAV8)–mediated delivery of Islr to hepatocytes increased BMP signaling and improved survival in our mouse model of hepatic metastasis. Conclusions Stromal BMP signaling predicts and modifies CRC progression and survival, and it can be therapeutically targeted by novel AAV-directed gene delivery to the liver.

Mist1 was recently shown to identify a discrete population of stem cells within the isthmus of the oxyntic gland within the gastric corpus. Chief cells at the base of the gastric corpus also express Mist1. The relevance of Mist1 expression as a marker of specific cell populations within the antral glands of the distal stomach, however, is unknown. Using Mist1-CreERT mice, we revealed that Mist1+ antral cells, distinct from the Mist1+population in the corpus, comprise long-lived progenitors that reside within the antral isthmus above Lgr5+or CCK2R+cells. Mist1+ antral progenitors can serve as an origin of antral tumors induced by loss of Apc or MNU treatment. Mist1+ antral progenitors, as well as other antral stem/progenitor population, express Cxcr4, and are located in close proximity to Cxcl12 (the Cxcr4 ligand)-expressing endothelium. During antral carcinogenesis, there is an expansion of Cxcr4+ epithelial cells as well as the Cxcl12+perivascular niche. Deletion of Cxcl12 in endothelial cells or pharmacological blockade of Cxcr4 inhibits antral tumor growth. Cxcl12/Cxcr4 signaling may be a potential therapeutic target., National Institute of Health (U.S.) (grant U54CA126513), National Institute of Health (U.S.) (grant R01CA093405), National Institute of Health (U.S.) (grant R01CA120979), National Institute of Health (U.S.) (grant R01DK052778), Clyde Wu Family Foundation

Dclk1-expressing tuft cells constitute a unique intestinal epithelial lineage that is distinct from enterocytes, Paneth cells, goblet cells, and enteroendocrine cells. Tuft cells express taste-related receptors and distinct transcription factors and interact closely with the enteric nervous system, suggesting a chemosensory cell lineage. In addition, recent work has shown that tuft cells interact closely with cells of the immune system, with a critical role in the cellular regulatory network governing responses to luminal parasites. Importantly, ablation of tuft cells severely impairs epithelial proliferation and tissue regeneration after injury, implicating tuft cells in the modulation of epithelial stem/progenitor function. Finally, tuft cells expand during chronic inflammation and in preneoplastic tissues, suggesting a possible early role in inflammation-associated tumorigenesis. Hence, we outline and discuss emerging evidence that strongly supports tuft cells as key regulatory cells in the complex network of the intestinal microenvironment.

Summary Infection with CagA-producing Helicobacter pylori plays a causative role in the development of gastric cancer. Upon delivery into gastric epithelial cells, CagA deregulates prooncogenic phosphatase SHP2 while inhibiting polarity-regulating kinase PAR1b through complex formation. Here, we show that CagA/PAR1b interaction subverts nuclear translocation of BRCA1 by inhibiting PAR1b-mediated BRCA1 phosphorylation. It hereby induces BRCAness that promotes DNA double-strand breaks (DSBs) while disabling error-free homologous recombination-mediated DNA repair. The CagA/PAR1b interaction also stimulates Hippo signaling that circumvents apoptosis of DNA-damaged cells, giving cells time to repair DSBs through error-prone mechanisms. The DSB-activated p53-p21Cip1 axis inhibits proliferation of CagA-delivered cells, but the inhibition can be overcome by p53 inactivation. Indeed, sequential pulses of CagA in TP53-mutant cells drove somatic mutation with BRCAness-associated genetic signatures. Expansion of CagA-delivered cells with BRCAness-mediated genome instability, from which CagA-independent cancer-predisposing cells arise, provides a plausible “hit-and-run mechanism” of H. pylori CagA for gastric carcinogenesis.

Background and aims Peribiliary glands (PBGs), clusters of epithelial cells residing in the submucosal compartment of extrahepatic bile ducts, have been suggested as biliary epithelial stem/progenitor cell niche; however, evidence to support this claim is limited due to a lack of PBG-specific markers. We therefore sought to identify PBG-specific markers to investigate the potential role of PBGs as stem/progenitor cell niches, as well as an origin of cancer. Methods We examined the expression pattern of the Wnt target gene Axin2 in extrahepatic bile ducts. We then applied lineage tracing to investigate whether Axin2-expressing cells from PBGs contribute to biliary regeneration and carcinogenesis using Axin2-CreERT mice. Results Wnt signaling activation, marked by Axin2, was limited to PBGs located in the periampullary region. Axin2+ cells in periampullary PBGs expressed stem cell markers as well as cell proliferation markers. Lineage tracing revealed that Axin2-expressing periampullary PBG cells are capable of self-renewal and supplying new biliary epithelial cells (BECs) to the luminal surface. Additionally, the expression pattern of Axin2 and CK19 was mutually exclusive in periampullary region, and fate tracing of CK19+ luminal surface BECs revealed gradual replacement by CK19- cells, further supporting the continuous replenishment of new BECs from PBGs to the luminal surface. We also found that R-spondin3 (Rspo3), an enhancer of Wnt signaling, was strongly expressed in the smooth muscle cells adjacent to the lower part of periampullary PBGs. These smooth muscle cells corresponded to human sphincter of Oddi, and similar expression pattern of Axin2 and Rspo3 was observed in human periampullary region. Notably, smooth muscle cell-specific deletion of Rspo3 in mice markedly decreased Axin2 expression in periampullary PBGs and induced significant atrophy of ampulla of Vater. Furthermore, introduction of PTEN deletion into Axin2+ PBG cells (Axin2;PTENΔ/Δ mice), but not CK19+ luminal surface BECs (CK19;PTENΔ/Δ mice), induced ampullary carcinoma. Ampullary carcinoma organoids generated from Axin2;PTENΔ/Δ mice were dependent on Rspo3 for their growth, suggesting that extrinsic stimulation of Wnt signaling is important for the growth of ampullary tumors. In fact, administration of Wnt inhibitor dramatically suppressed ampullary carcinoma development in Axin2;PTENΔ/Δ mice. Conclusion A specific cell population receiving Wnt-activating signal in periampullary PBGs functions as biliary epithelial stem/progenitor cells and also cellular origin of ampullary carcinoma. These new findings can facilitate the development of new therapeutic strategies to induce biliary regeneration, as well as to treat ampullary carcinoma. Download : Download high-res image (147KB) Download : Download full-size image Graphical Abstract

Background & Aims Peribiliary glands (PBGs), clusters of epithelial cells residing in the submucosal compartment of extrahepatic bile ducts, have been suggested as biliary epithelial stem/progenitor cell niche; however, evidence to support this claim is limited because of a lack of PBG-specific markers. We therefore sought to identify PBG-specific markers to investigate the potential role of PBGs as stem/progenitor cell niches, as well as an origin of cancer. Methods We examined the expression pattern of the Wnt target gene Axin2 in extrahepatic bile ducts. We then applied lineage tracing to investigate whether Axin2-expressing cells from PBGs contribute to biliary regeneration and carcinogenesis using Axin2-CreERT mice. Results Wnt signaling activation, marked by Axin2, was limited to PBGs located in the periampullary region. Lineage tracing showed that Axin2-expressing periampullary PBG cells are capable of self-renewal and supplying new biliary epithelial cells (BECs) to the luminal surface. Additionally, the expression pattern of Axin2 and the mature ductal cell marker CK19 were mutually exclusive in periampullary region, and fate tracing of CK19+ luminal surface BECs showed gradual replacement by CK19– cells, further supporting the continuous replenishment of new BECs from PBGs to the luminal surface. We also found that Wnt signal enhancer R-spondin3 secreted from Myh11-expressing stromal cells, corresponding to human sphincter of Oddi, maintained the periampullary Wnt signal–activating niche. Notably, introduction of PTEN deletion into Axin2+ PBG cells, but not CK19+ luminal surface BECs, induced ampullary carcinoma whose development was suppressed by Wnt inhibitor. Conclusion A specific cell population receiving Wnt-activating signal in periampullary PBGs functions as biliary epithelial stem/progenitor cells and also the cellular origin of ampullary carcinoma.

The cellular origin of digestive cancers has been a long-standing question in the cancer field. Mouse models have identified long-lived stem cells in most organ systems, including the luminal gastrointestinal tract, and numerous studies have pointed to tissue resident stem cells as the main cellular origin of cancer. During gastric carcinogenesis, chronic inflammation induces genetic and epigenetic alterations in long-lived stem cells, along with expansion of stem cell niches, eventually leading to invasive cancer. The gastric corpus and antrum have distinct stem cells and stem cell niches, suggesting differential regulation of cancer initiation at the 2 sites. In this short review, we discuss recent experimental models and human studies, which provide important insights into the pathogenesis of gastric cancer., National Institutes of Health (U.S.) (grant U54CA126513), National Institutes of Health (U.S.) (grant R01CA093405), National Institutes of Health (U.S.) (grant R01CA120979), National Institutes of Health (U.S.) (grant R01DK052778), National Institutes of Health (U.S.) (grant R35CA210088), National Institutes of Health (U.S.) (grant T32OD010978), National Institutes of Health (U.S.) (grant P30ES002109), National Institutes of Health (U.S.) (grantd P01CA28842), Clyde Wu Family Foundation

Objective Vonoprazan is a potassium-competitive acid blocker, a new type of acid-suppressing drug, and has recently become available for peptic ulcers, gastroesophageal reflux disease, and Helicobacter pylori (H. pylori) eradication. Its efficacy for H. pylori eradication has been reported. However, the evidence for its efficacy and feasibility remains limited. We aimed to compare the feasibility, effectiveness and safety of vonoprazan-based triple therapy with conventional proton pump inhibitor (PPI)-based triple therapy in multicenter clinical practice. Methods We performed a multicenter retrospective study on patients receiving first-line H. pylori eradication therapy between March 2013 and November 2015 with either vonoprazan-based triple therapy or conventional PPI-based triple therapy. Results A total of 2715 patients aged 63.0 ± 12.1 years (1412 [52.0%] males) were analyzed. Eradication rates were 87.2% (368/422) for vonoprazan-based therapy and 72.4% (1661/2293) for conventional PPI-based therapy (P Conclusion Vonoprazan-based triple therapy is feasible, and has a higher rate for H. pylori eradication than conventional PPI as a first-line regimen.

Background and Aims Helicobacter pylori eradication therapy is effective at reducing the incidence of gastric cancer; however, gastric cancer still develops after eradication. We conducted a cohort study to elucidate the risk factors for gastric cancer development after successful H pylori eradication therapy. Methods From June 1998 to December 2012 we assessed histologic and endoscopic findings of gastritis and performed H pylori eradication therapy in 748 patients without a history of gastric cancer. Patients were classified according to the distribution of intestinal metaplasia (IM) as follows: no IM (IM group A), IM in the antrum only (IM group B), and IM in the corpus (IM group C). We assessed atrophy endoscopically according to the Kimura-Takemoto classification system. Gastric cancer incidence was assessed. Results A total of 573 patients underwent follow-up endoscopy; the mean duration of follow-up was 6.2 ± 4.8 years. Gastric cancer developed in 21 (20 intestinal type). The cumulative 5-year incidences of gastric cancer were 3.2% overall; 1.5%, 5.3%, and 9.8% in IM groups A, B, and C; and 0.7%, 1.9%, and 10% in the none/mild, moderate, and severe endoscopic atrophy groups, respectively. Compared with IM group A, the hazard ratio for IM group B was 3.6 (95% confidence interval [CI], 1.2-11), and that for IM group C was 3.7 (95% CI, 1.1-12). Compared with the none/mild endoscopic atrophy group, the hazard ratio for severe atrophy was 9.3 (95% CI, 1.7-174). Conclusions Patients with histologic IM or severe endoscopic atrophy were at increased risk of gastric cancer development after H pylori eradication.

The small intestinal tuft cell-ILC2 circuit mediates epithelial responses to intestinal helminths and protists by tuft cell chemosensory-like sensing and IL-25-mediated activation of lamina propria ILC2s. Small intestine ILC2s constitutively express the IL-25 receptor, which is negatively regulated by A20 (Tnfaip3). A20-deficiency in ILC2s spontaneously triggers the circuit, and, unexpectedly, promotes adaptive small intestinal lengthening and remodeling. Circuit activation occurs upon weaning, and is enabled by dietary polysaccharides that render mice permissive for Tritrichomonas colonization, resulting in luminal accumulation of acetate and succinate, metabolites of the protist hydrogenosome. Tuft cells express GPR91, the succinate receptor, and dietary succinate, but not acetate, activates ILC2s via a tuft-, TRPM5-, and IL-25-dependent pathway. Also induced by parasitic helminths, circuit activation and small intestinal remodeling impairs infestation by new helminths, consistent with the phenomenon of concomitant immunity. We describe a metabolic sensing circuit that may have evolved to facilitate mutualistic responses to luminal pathosymbionts.

In the tumor microenvironment, various stromal and immune cells accumulate and interact with cancer cells to contribute to tumor progression. Among stromal players, nerves have recently been recognized as key regulators of tumor growth. More neurotransmitters, such as catecholamines and acetylcholine (ACh), are present in tumors, as the cells that secrete neurotransmitters accumulate by the release of neurotrophic factors from cancer cells. In this short review, we focus on the role of nerve signaling in gastrointestinal (GI) cancers. Given that muscarinic acetylcholine receptor signaling seems to be a dominant regulator of GI stem cells and cancers, we review the function and mechanism of the muscarinic ACh pathway as a regulator of GI cancer progression. Accumulating evidence suggests that ACh, which is secreted from nerves and tuft cells, stimulates GI epithelial stem cells and contributes to cancer progression via muscarinic receptors.

Summary Cancer is believed to arise from stem cells, but mechanisms that limit the acquisition of mutations and tumor development have not been well defined. We show that a +4 stem cell (SC) in the gastric antrum, marked by expression of Cck2r (a GPCR) and Delta-like ligand 1 (DLL1), is a label-retaining cell that undergoes predominant asymmetric cell division. This +4 antral SC is Notch1low/ Numb+ and repressed by signaling from gastrin-expressing endocrine (G) cells. Chemical carcinogenesis of the stomach is associated with loss of G cells, increased symmetric stem cell division, glandular fission, and more rapid stem cell lineage tracing, a process that can be suppressed by exogenous gastrin treatment. This hormonal suppression is associated with a marked reduction in gastric cancer mutational load, as revealed by exomic sequencing. Taken together, our results show that gastric tumorigenesis is associated with increased symmetric cell division that facilitates mutation and is suppressed by GPCR signaling.

We read with interest the recent publication by Cheung et al . The authors conclude that proton pump inhibitor (PPI) use is associated with a 2.4-fold increase in the risk of gastric cancer in patients who received Helicobacter pylori eradication therapy.1 This is an important finding given that the safety of long-term PPI use remains unclear. Notably, no significant association between histamine-2 receptor antagonist (H2RA) use and gastric cancer was observed, which suggests a PPI-specific effect in this context. Cheung et al used a large population-based database to select patients after eradication and performed propensity score adjustment analysis. However, as the authors described in the Discussion section, we have concerns about the limitations of this study, in particular the lack of endoscopic and histological examinations, the results of which could influence the association between PPI use and gastric cancer. The presence …

The discovery of Lgr5+ intestinal stem cells (ISCs) triggered a breakthrough in the field of ISC research. Lgr5+ ISCs maintain the homeostasis of the intestinal epithelium in the steady state, while these cells are susceptible to epithelial damage induced by chemicals, pathogens, or irradiation. During the regeneration process of the intestinal epithelium, more quiescent +4 stem cells and short-lived transit-amplifying (TA) progenitor cells residing above Lgr5+ ISCs undergo dedifferentiation and act as stem-like cells. In addition, several recent reports have shown that a subset of terminally differentiated cells, including Paneth cells, tuft cells, or enteroendocrine cells, may also have some degree of plasticity in specific situations. The function of ISCs is maintained by the neighboring stem cell niches, which strictly regulate the key signal pathways in ISCs. In addition, various inflammatory cytokines play critical roles in intestinal regeneration and stem cell functions following epithelial injury. Here, we summarize the current understanding of ISCs and their niches, review recent findings regarding cellular plasticity and its regulatory mechanism, and discuss how inflammatory cytokines contribute to epithelial regeneration.

Dendritic cells (DCs) mediate host immune responses to gut microbes and play critical roles in inflammatory bowel disease. In this study, we examined the role of TGF-β signaling in DCs in colonic homeostasis. CD11c-cre Tgfbr2fl/fl mice developed spontaneous colitis, and CD11c-cre Tgfbr2fl/+ mice exhibited susceptibility to dextran sulfate sodium–induced colitis. Colitis in these mice was characterized by goblet cell depletion and dysbiosis caused by Enterobacteriaceae enrichment. Wild-type mice gavaged with Enterobacteriaceae from CD11c-cre Tgfbr2fl/fl mice feces showed severe colitis after dextran sulfate sodium treatment, whereas those treated with Notch inhibitor exhibited attenuated colonic injury with increased goblet cell numbers, thickened mucus layer, and fewer fecal Enterobacteriaceae. Wild-type mice transplanted with CD11c-cre Tgfbr2fl/fl bone marrow developed colitis showing increased Jagged1 and Jagged2 in DCs, increased Hes1 levels in epithelium, and goblet cell depletion. These findings suggest that TGF-β signaling in DCs regulates intestinal homeostasis by modulating epithelial cell differentiation and fecal microbiota.

Over recent decades we have witnessed a shift in the anatomical distribution of gastric cancer (GC), which increasingly originates from the proximal stomach near the junction with the oesophagus. In parallel, there has been a dramatic rise in the incidence of oesophageal adenocarcinoma (OAC) in the lower oesophagus, which is associated with antecedent Barrett oesophagus (BO). In this context, there has been uncertainty regarding the characterization of adenocarcinomas spanning the area from the lower oesophagus to the distal stomach. Most relevant to this discussion is the distinction, if any, between OAC and intestinal-type GC of the proximal stomach. It is therefore timely to review our current understanding of OAC and intestinal-type GC, integrating advances from cell-of-origin studies and comprehensive genomic alteration analyses, ultimately enabling better insight into the relationship between these two cancers.

Introduction Cancer-associated fibroblasts (CAFs), a heterogeneous component of the tumor microenvironment, substantially influence tumor progression. Bone morphogenetic proteins (BMP) play a critical part in defining the intestinal epithelial niche and either promote or retard cancer progression in a context-dependent manner. However, the role of BMP signaling in the colorectal cancer (CRC) stroma remains to be fully elucidated. This study investigated the significance of mesenchymal BMP signaling as a potential therapeutic target in CRC progression. Design Using CRC expression array data, we identified two CAF-specific factors involved in BMP signaling, then verified their upregulation in the human CRC stroma by in-situ hybridization (ISH). We took advantage of a preclinical mouse model of CRC hepatic metastasis to test approaches targeting the BMP signaling pathway. Results CRC microarray data identified GREM1 and ISLR as CAF-specific genes involved in BMP signaling. In colonic myofibroblasts, Grem1-overexpression inhibited BMP signaling whereas BMP7 signaling was augmented by Islr overexpression, suggesting opposing roles for GREM1 and ISLR in the regulation of BMP signaling. ISH using human rectal cancer samples revealed that GREM1 and ISLR were expressed in distinct CAF subpopulations and that GREM1 and ISLR expression predicted poor and favorable survival, respectively. Notably, Grem1 and Islr expression was differentially regulated by Foxl1, an intestinal mesenchyme-lineage transcription factor, and TGF-b, indicating a mechanism for generating fibroblast heterogeneity. Finally, adeno-associated virus 8-mediated in-vivo overexpression of Islr in hepatocytes retarded growth and generated more differentiated histology in CRC hepatic metastases. Conclusion These data suggest that increased stromal BMP signaling may ameliorate CRC progression and provide a rationale for targeting stromal BMP signaling to inhibit CRC progression and metastasis. Citation Format: Hiroki Kobayashi, Krystyna A. Gieniec, Tongtong Wang, Josephine A. Wright, Nobumi Suzuki, Tamsin RM Lannagan, Yoku Hayakawa, Simon J. Leedham, Nicholas Arpaia, Siddhartha Mukherjee, Timothy C. Wang, Atsushi Enomoto, Masahide Takahashi, Daniel L. Worthley, Susan L. Woods. Stromal BMP signaling imbalance mediated by GREM1 and ISLR regulates colorectal cancer progression [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3977.

Background & Aims Gastric chief cells, a mature cell type that secretes digestive enzymes, have been proposed to be the origin of metaplasia and cancer through dedifferentiation or transdifferentiation. However, studies supporting this claim have had technical limitations, including issues with the specificity of chief cell markers and the toxicity of drugs used. We therefore sought to identify genes expressed specifically in chief cells and establish a model to trace these cells. Methods We performed transcriptome analysis of Mist1-CreERT–traced cells, with or without chief cell depletion. Gpr30-rtTA mice were generated and crossed to TetO-Cre mice, and lineage tracing was performed after crosses to R26-TdTomato mice. Additional lineage tracing experiments were performed using Mist1-CreERT, Kitl-CreERT, Tff1-Cre, and Tff2-Cre mice crossed to reporter mice. Mice were given high-dose tamoxifen or DMP-777 or were infected with Helicobacter pylori to induce gastric metaplasia. We studied mice that expressed mutant forms of Ras in gastric cells, using TetO-KrasG12D, LSL-KrasG12D, and LSL-HrasG12V mice. We analyzed stomach tissues from GPR30-knockout mice. Mice were given dichloroacetate to inhibit pyruvate dehydrogenase kinase (PDK)–dependent cell competition. Results We identified GPR30, the G-protein–coupled form of the estrogen receptor, as a cell-specific marker of chief cells in gastric epithelium of mice. Gpr30-rtTA mice crossed to TetO-Cre;R26-TdTomato mice had specific expression of GPR30 in chief cells, with no expression noted in isthmus stem cells or lineage tracing of glands. Expression of mutant Kras in GPR30+ chief cells did not lead to the development of metaplasia or dysplasia but, instead, led to a reduction in labeled numbers of chief cells and a compensatory expansion of neck lineage, which was derived from upper Kitl+ clones. Administration of high-dose tamoxifen, DMP-777, or H pylori decreased the number of labeled chief cells. Chief cells were eliminated from epithelia via GPR30- and PDK-dependent cell competition after metaplastic stimuli, whereas loss of GRP30 or inhibition of PDK activity preserved chief cell numbers and attenuated neck lineage cell expansion. Conclusions In tracing studies of mice, we found that most chief cells are lost during metaplasia and therefore are unlikely to contribute to gastric carcinogenesis. Expansion of cells that coexpress neck and chief lineage markers, known as spasmolytic polypeptide-expressing metaplasia, does not occur via dedifferentiation from chief cells but, rather, through a compensatory response from neck progenitors to replace the eliminated chief cells.

ObjectiveSerrated colorectal cancer (CRC) accounts for approximately 25% of cases and includes tumours that are among the most treatment resistant and with worst outcomes. This CRC subtype is associated with activating mutations in the mitogen-activated kinase pathway gene, BRAF, and epigenetic modifications termed the CpG Island Methylator Phenotype, leading to epigenetic silencing of key tumour suppressor genes. It is still not clear which (epi-)genetic changes are most important in neoplastic progression and we begin to address this knowledge gap herein.DesignWe use organoid culture combined with CRISPR/Cas9 genome engineering to sequentially introduce genetic alterations associated with serrated CRC and which regulate the stem cell niche, senescence and DNA mismatch repair.ResultsTargeted biallelic gene alterations were verified by DNA sequencing. Organoid growth in the absence of niche factors was assessed, as well as analysis of downstream molecular pathway activity. Orthotopic engraftment of complex organoid lines, but not BrafV600E alone, quickly generated adenocarcinoma in vivo with serrated features consistent with human disease. Loss of the essential DNA mismatch repair enzyme, Mlh1, led to microsatellite instability. Sphingolipid metabolism genes are differentially regulated in both our mouse models of serrated CRC and human CRC, with key members of this pathway having prognostic significance in the human setting.ConclusionWe generate rapid, complex models of serrated CRC to determine the contribution of specific genetic alterations to carcinogenesis. Analysis of our models alongside patient data has led to the identification of a potential susceptibility for this tumour type.

Small bowel adenocarcinoma (SBA) is a rare, aggressive malignancy with a poor prognosis, and the mechanisms of carcinogenesis in SBA remain unclear. Our aims were to investigate the molecular mechanisms underlying SBA and to identify treatments by establishing and characterizing an SBA cell line and performing anti-cancer drug screening. SIAC1 cells, established from jejunal SBA, showed epithelial characteristics and formed organoids in 3D culture. SIAC1 cells had a heterozygous β-catenin deletion mutation, resulting in a stable β-catenin protein with enhanced Wnt/β-catenin activity. SIAC1 cells lacked MLH1 and MSH6 expression, and target genes such as TGFBR2 and ACVR2 showed frameshift mutations. Among 10 clinical SBA samples, 2 (20%) had interstitial deletions in β-catenin, expression of mismatch repair protein was aberrant in 4 (40%), and heterozygous frameshift mutations of three target genes were found in all 10 samples. On screening assay using 140 compounds, eribulin significantly inhibited SIAC1 cell growth both in vitro and in vivo by inhibition of the Wnt/β-catenin pathway via enhanced degradation of β-catenin. In conclusion, we established an SBA cell line with molecular characteristics similar to those of clinical SBA samples, including β-catenin deletion and mismatch repair protein deficiency, that will be useful for SBA research. Eribulin might be a candidate for SBA treatment due to its inhibitory effect on Wnt/β-catenin signaling.

The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs)., National Institutes of Health (U.S.) (Grant R01 RHL115145A)

Despite a significant decrease in the incidence of gastric cancer in Western countries over the past century, gastric cancer is still one of the leading causes of cancer-related deaths worldwide. Most human gastric cancers develop after long-term Helicobacter pylori infection via the Correa pathway: the progression is from gastritis, atrophy, intestinal metaplasia, dysplasia, to cancer. However, it remains unclear whether metaplasia is a direct precursor of gastric cancer or merely a marker of high cancer risk. Here, we review human studies on the relationship between metaplasia and cancer in the stomach, data from mouse models of metaplasia regarding the mechanism of metaplasia development, and the cellular responses induced by H. pylori infection.

Background Recent studies show that some Escherichia coli strains possessing a gene cluster named the pks island might have a causative role in the development of human colorectal cancer (CRC). In several reports from Europe, they are found more prevalently in colon tissue specimens derived from CRC patients compared to those from controls. In this study we sought to clarify the difference in pks prevalence between CRC patients and non-CRC controls in the Japanese population, by using non-invasive sample collection technique during colonoscopy. Methods Colonic lavage samples were collected during diagnostic colonoscopy, and bacterial DNA within each sample was extracted. Fecal DNA samples were then examined for pks island genes using conventional qualitative PCR and real-time quantitative PCR. In some patients biopsy samples were also collected in the same session of colonoscopy, and the correlation between the pks status of the colonic lavage sample and the biopsy sample of the same patients was evaluated. Results Twelve out of thirteen patients (92%) showed the same pks status by colonic lavage sample and biopsy sample, suggesting the usefulness of colonic lavage samples as a surrogate for biopsy samples. A total of 98 colonic lavage samples were collected, which included 35 from CRC patients, 37 from adenoma patients, and 26 from controls. The pks-positive bacterial DNA was detected in 43, 51, and 46% of colonic lavage samples from CRC, adenoma, and control patients, respectively, and there was no significant difference among diseases. Real-time quantitative PCR showed no significant difference in the relative concentrations of pks-positive bacterial DNA among diseases. Age, gender, location of CRC, CRC staging, or k-ras gene status was not associated with pks prevalence. Conclusions Although the method of collecting fecal DNA from colonic lavage samples was safe and technically feasible, factors other than pks-positive bacteria appear to play more important roles in CRC development in this cohort.

The carcinogenic mechanism of extrahepatic cholangiocarcinoma (ECC) is unclear, due at least in part to the lack of an appropriate mouse model. Because human studies have reported frequent genetic alterations in the Ras- and TGFβ/SMAD-signaling pathways in ECC, mice with tamoxifen-inducible, duct-cell-specific Kras activation and a TGFβ receptor type 2 (TGFβR2) deletion were first generated by crossing LSL-KrasG12D , Tgfbr2flox/flox , and K19CreERT mice (KT-K19CreERT ). However, KT-K19CreERT mice showed only mild hyperplasia of biliary epithelial cells (BECs) in the extrahepatic bile duct (EHBD) and died within 7 wk, probably a result of lung adenocarcinomas. Next, to analyze the additional effect of E-cadherin loss, KT-K19CreERT mice were crossed with CDH1flox/flox mice (KTC-K19CreERT ). Surprisingly, KTC-K19CreERT mice exhibited a markedly thickened EHBD wall accompanied by a swollen gallbladder within 4 wk after tamoxifen administration. Histologically, invasive periductal infiltrating-type ECC with lymphatic metastasis was observed. Time-course analysis of EHBD revealed that recombined BECs lining the bile duct lumen detached due to E-cadherin loss, whereas recombined cells could survive in the peribiliary glands (PBGs), which are considered a BEC stem-cell niche. Detached dying BECs released high levels of IL-33, as determined by microarray analysis using biliary organoids, and stimulated inflammation and a regenerative response by PBGs, leading eventually to ECC development. Cell lineage tracing suggested PBGs as the cellular origin of ECC. IL-33 cooperated with Kras and TGFβR2 mutations in the development of ECC, and anti-IL-33 treatment suppressed ECC development significantly. Thus, this mouse model provided insight into the carcinogenic mechanisms, cellular origin, and potential therapeutic targets of ECC.

Myeloid-biased hematopoietic stem cells (MB-HSCs) play critical roles in recovery from injury, but little is known about how they are regulated within the bone marrow niche. Here we describe an auto-/paracrine physiologic circuit that controls quiescence of MB-HSCs and hematopoietic progenitors marked by histidine decarboxylase (Hdc). Committed Hdc+ myeloid cells lie in close anatomical proximity to MB-HSCs and produce histamine, which activates the H2 receptor on MB-HSCs to promote their quiescence and self-renewal. Depleting histamine-producing cells enforces cell cycle entry, induces loss of serial transplant capacity, and sensitizes animals to chemotherapeutic injury. Increasing demand for myeloid cells via lipopolysaccharide (LPS) treatment specifically recruits MB-HSCs and progenitors into the cell cycle; cycling MB-HSCs fail to revert into quiescence in the absence of histamine feedback, leading to their depletion, while an H2 agonist protects MB-HSCs from depletion after sepsis. Thus, histamine couples lineage-specific physiological demands to intrinsically primed MB-HSCs to enforce homeostasis. Chen et al. show that histidine decarboxylase (Hdc) marks quiescent myeloid-biased HSCs (MB-HSCs). Daughter myeloid cells form a spatial cluster with Hdc+ MB-HSCs and secrete histamine to enforce their quiescence and protect them from depletion, following activation by a variety of physiologic insults.

The acquisition of genetic/epigenetic mutations in long-lived gastrointestinal stem cells leads to the development of cancer, as well as precancerous lesions such as metaplasia and dysplasia. In the proximal stomach corpus, this model of progression from stem cells has been supported by studies in mice and human beings, showing abundant proliferation in the isthmus and clonal expansion of mutated cells from the stem cell region. An alternative theory proposes that gastric metaplasia arises from mature differentiated chief cells. Despite reports of low levels of proliferation in chief cells in acute injury models, there is little evidence for reprogramming of chief cells into long-lived stem cells that continuously supply progeny over time. Critical flaws in the chief cell transdifferentiation theory include the definition of acute SPEM, the chief cell-damaging effect of chemical reagents, and the specificity of chief cell lineage tracing. In contrast, there is now strong evidence regarding the stem cell origins of gastric metaplasia that refutes the transdifferentiation theory. Here, we briefly review the history and definition of gastric metaplasia, and outline in detail the evidence that supports the stem cell origin of metaplasia.

Doublecortin-like kinase 1 protein (DCLK1) is a gastrointestinal tuft cell marker that has been proposed to identify quiescent and tumor growth-sustaining stem cells. DCLK1+ tuft cells are increased in inflammation-induced carcinogenesis; however, the role of these cells within the gastrointestinal epithelium and their potential as cancer-initiating cells are poorly understood. Here, using a BAC-CreERT-dependent genetic lineage-tracing strategy, we determined that a subpopulation of DCLK1+ cells is extremely long lived and possesses rare stem cell abilities. Moreover, genetic ablation of Dclk1 revealed that DCLK1+ tuft cells contribute to recovery following intestinal and colonic injury. Surprisingly, conditional knockdown of the Wnt regulator APC in DCLK1+ cells was not sufficient to drive colonic carcinogenesis under normal conditions; however, dextran sodium sulfate-induced (DSS-induced) colitis promoted the development of poorly differentiated colonic adenocarcinoma in mice lacking APC in DCLK1+ cells. Importantly, colonic tumor formation occurred even when colitis onset was delayed for up to 3 months after induced APC loss in DCLK1+ cells. Thus, our data define an intestinal DCLK1+ tuft cell population that is long lived, quiescent, and important for intestinal homeostasis and regeneration. Long-lived DCLK1+ cells maintain quiescence even following oncogenic mutation, but are activated by tissue injury and can serve to initiate colon cancer.

E-cadherin is an important adhesion molecule whose loss is associated with progression and poor prognosis of liver cancer. However, it is unclear whether the loss of E-cadherin is a real culprit or a bystander in liver cancer progression. In addition, the precise role of E-cadherin in maintaining liver homeostasis is also still unknown, especially in vivo. Here we demonstrate that liver-specific E-cadherin knockout mice develop spontaneous periportal inflammation via an impaired intrahepatic biliary network, as well as periductal fibrosis, which resembles primary sclerosing cholangitis. Inducible gene knockout studies identified E-cadherin loss in biliary epithelial cells as a causal factor of cholangitis induction. Furthermore, a few of the E-cadherin knockout mice developed spontaneous liver cancer. When knockout of E-cadherin is combined with Ras activation or chemical carcinogen administration, E-cadherin knockout mice display markedly accelerated carcinogenesis and an invasive phenotype associated with epithelial-mesenchymal transition, up-regulation of stem cell markers, and elevated ERK activation. Also in human hepatocellular carcinoma, E-cadherin loss correlates with increased expression of mesenchymal and stem cell markers, and silencing of E-cadherin in hepatocellular carcinoma cell lines causes epithelial-mesenchymal transition and increased invasiveness, suggesting that E-cadherin loss can be a causal factor of these phenotypes. Thus, E-cadherin plays critical roles in maintaining homeostasis and suppressing carcinogenesis in the liver.

Within the gastrointestinal stem cell niche, nerves help to regulate both normal and neoplastic stem cell dynamics. Here, we reveal the mechanisms underlying the cancer-nerve partnership. We find that Dclk1+ tuft cells and nerves are the main sources of acetylcholine (ACh) within the gastric mucosa. Cholinergic stimulation of the gastric epithelium induced nerve growth factor (NGF) expression, and in turn NGF overexpression within gastric epithelium expanded enteric nerves and promoted carcinogenesis. Ablation of Dclk1+ cells or blockade of NGF/Trk signaling inhibited epithelial proliferation and tumorigenesis in a muscarinic acetylcholine receptor-3 (M3R)-dependent manner, in part through suppression of Yes-Associated Protein (YAP) function. This feed-forward ACh-NGF axis activates the gastric cancer niche and offers a compelling target for tumor treatment and prevention.

Background: Helicobacter pylori (H. pylori) eradication therapy may improve gastric atrophy and intestinal metaplasia, but the results of previous studies have not always been consistent. The aim of this study was to compare the histological changes of intestinal metaplasia and gastric atrophy among the use of acid-suppressing drugs after H. pylori eradication. Methods: A cohort of 242 patients who underwent successful eradication therapy for H. pylori gastritis and surveillance endoscopy examination from 1996 to 2015 was analyzed. Changes in the histological scores of intestinal metaplasia and atrophy according to drug use (proton-pump inhibitors (PPIs), H2 receptor antagonists (H2RAs), and non-acid suppressant use) were evaluated in biopsies of the antrum and corpus using a generalized linear mixed model in all patients. Results: The mean follow-up period and number of biopsies were 5.48 &plusmn, 4.69 years and 2.62 &plusmn, 1.67 times, respectively. Improvement in the atrophy scores of both the antrum (p = 0.042) and corpus (p = 0.020) were significantly superior in patients with non-acid suppressant drug use compared with those of PPI and H2RA use. Metaplasia scores in both the antrum and corpus did not improve in all groups, and no significant differences were observed among groups in the antrum (p = 0.271) and corpus (p = 0.077). Conclusions: Prolonged acid suppression by PPIs or H2RAs may limit the recovery of gastric atrophy following H. pylori eradication.