Intestinal Epithelial Homeostasis Research Articles

Dysregulation of IL6/IL6R-STAT3-SOCS3 signaling pathway in IBD-associated colorectal dysplastic lesions as compared to sporadic colorectal adenomas in non-IBD patients

BackgroundIL6-IL6R-STAT3-SOCS3 signaling pathway is known to play important roles in regulating intestinal epithelial homeostasis, in pathogenesis of inflammatory bowel disease (IBD), and in tumorigenesis of colorectal neoplasia. We studied the expressions of these factors in IBD-associated dysplasia and compared to sporadic colorectal adenomas in non-IBD individuals. Materials and methodsThe expression of IL6, IL6R, STAT3, and SOCS3 within dysplastic as well as background non-dysplastic epithelial cells was evaluated by immunohistochemistry in 26 sporadic colorectal adenomas in non-IBD patients, 32 adenoma-like and 30 non-adenoma-like dysplastic lesions in IBD (41 ulcerative colitis, 21 Crohn’s disease) patients. The level of expression of each factor was arbitrarily scored as 0, 1, 2, and 3. ResultsIn both IBD and non-IBD lesions, neoplastic epithelium showed a higher expression of all factors, except the IL6R, as compared to non-neoplastic epithelium. For non-neoplastic epithelium between IBD and non-IBD settings, the colitic epithelium showed a similar IL6, lower IL6R, higher STAT3, and higher SOCS3 expression. As compared to non-IBD adenomas, IBD-associated dysplasia showed a significantly lower IL6, lower IL6R, higher STAT3, and lower SOCS3 expression. Most notably, a parallel-elevation pattern of STAT3/SOCS3 expressions was seen in non-IBD adenomas but an inverse-expression pattern of STAT3/SOCS3 seen in IBD dysplastic lesions. No significant differences existed between adenoma-like and non-adenoma-like lesions. ConclusionsIL6/IL6R-STAT3-SOCS3 signaling pathway does not seem to be preferentially associated with IBD-associated dysplasia. However, the STAT3-SOCS3 interaction appears dysregulated in IBD, characterized by a loss of STAT3/SOCS3 balance,i.e., loss of the normal negative regulation of SOCS3.

LSD1 represses a neonatal/reparative gene program in adult intestinal epithelium.

Intestinal epithelial homeostasis is maintained by adult intestinal stem cells, which, alongside Paneth cells, appear after birth in the neonatal period. We aimed to identify regulators of neonatal intestinal epithelial development by testing a small library of epigenetic modifier inhibitors in Paneth cell-skewed organoid cultures. We found that lysine-specific demethylase 1A (Kdm1a/Lsd1) is absolutely required for Paneth cell differentiation. Lsd1-deficient crypts, devoid of Paneth cells, are still able to form organoids without a requirement of exogenous or endogenous Wnt. Mechanistically, we find that LSD1 enzymatically represses genes that are normally expressed only in fetal and neonatal epithelium. This gene profile is similar to what is seen in repairing epithelium, and we find that Lsd1-deficient epithelium has superior regenerative capacities after irradiation injury. In summary, we found an important regulator of neonatal intestinal development and identified a druggable target to reprogram intestinal epithelium toward a reparative state.

SIRT7 Facilitates CENP-A Nucleosome Assembly and Suppresses Intestinal Tumorigenesis.

SummarySIRT7 is a member of the mammalian sirtuins and functions as an NAD+-dependent deacylase. Here we show that SIRT7 deficiency leads to a lowered histone acetyltransferase 1 (HAT1) activity and therefore decreased histone H4K5 and H4K12 acetylation. This in turn causes CENP-A dislocation at the centromere, which further affects chromatin assembly. SIRT7 ablation results in aneuploidy and aging phenotypes, including senescence and nucleolar expansion. Moreover, SIRT7 knockout mice are susceptible to DSS-induced colitis and alcohol-derived epithelial disturbance, revealing a disrupted intestinal epithelial homeostasis. Notably, absence of SIRT7 aggravates the susceptibility of colorectal cancer incidence in APCMin/+ mouse model and elicits further the Wnt signaling. Our findings indicate a tumor suppressive role of SIRT7 in the case of colorectal cancer. Together with the activities in maintaining genome integrity and intestinal homeostasis, activating SIRT7 may serve as a strategy to treat bowel diseases and colorectal cancer.

How commensal microbes shape the physiology of Drosophila melanogaster.

The interactions between animals and their commensal microbes profoundly influence the host's physiology. In the last decade, Drosophila melanogaster has been extensively used as a model to study host-commensal microbes interactions. Here, we review the most recent advances in this field. We focus on studies that extend our understanding of the molecular mechanisms underlying the effects of commensal microbes on Drosophila's development and lifespan. We emphasize how commensal microbes influence nutrition and the intestinal epithelium homeostasis; how they elicit immune tolerance mechanisms and how these physiological processes are interconnected. Finally, we discuss the importance of diets and microbial strains and show how they can be confounding factors of microbe mediated host phenotypes.

Gut microbiota maturation during early human life induces enterocyte proliferation via microbial metabolites

BackgroundThe intestinal tract undergoes a period of cellular maturation during early life, primarily characterized by the organization of epithelial cells into specialized crypt and villus structures. These processes are in part mediated by the acquisition of microbes. Infants delivered at term typically harbor a stable, low diversity microbiota characterized by an overrepresentation of various Bacilli spp., while pre-term infants are colonized by an assortment of bacteria during the first several weeks after delivery. However, the functional effects of these changes on intestinal epithelium homeostasis and maturation remain unclear. To study these effects, human neonate feces were obtained from term and pre-term infants. Fecal 16S rDNA sequencing and global untargeted LC-MS were performed to characterize microbial composition and metabolites from each population. Murine enteral organoids (enteroids) were cultured with 0.22 μm filtered stool supernatant pooled from term or pre-term infants.ResultsTerm and pre-term microbial communities differed significantly from each other by principle components analysis (PCoA, PERMANOVA p < 0.001), with the pre-term microbiome characterized by increased OTU diversity (Wilcox test p < 0.01). Term communities were less diverse and dominated by Bacilli (81.54%). Pre-term stools had an increased abundance of vitamins, amino acid derivatives and unconjugated bile acids. Pathway analysis revealed a significant increase in multiple metabolic pathways in pre-term samples mapped to E. coli using the KEGG database related to the fermentation of various amino acids and vitamin biosynthesis. Enteroids cultured with supernatant from pre-term stools proliferated at a higher rate than those cultured with supernatant from term stools (cell viability: 207% vs. 147.7%, p < 0.01), grew larger (area: 81,189μm2 vs. 41,777μm2, p < 0.001), and bud at a higher rate (6.5 vs. 4, p < 0.01). Additionally, genes involved in stem cell proliferation were upregulated in pre-term stool treated enteroid cultures (Lgr5, Ephb2, Ascl2 Sox9) but not term stool treated enteroids.ConclusionsOur findings indicate that microbial metabolites from the more diverse gut microbiome associated with pre-term infants facilitate stem cell proliferation. Therefore, perturbations of the pre-term microbiota may impair intestinal homeostasis.

Arachidonic Acid Promotes Intestinal Regeneration by Activating WNT Signaling.

SummaryIntestinal regeneration is crucial for functional restoration after injury, and nutritional molecules can play an important role in this process. Here, we found that arachidonic acid (AA) serves as a direct proliferation promoter of intestinal epithelial cells that facilitates small intestinal regeneration in both three-dimensional cultured organoids and mouse models. As shown in the study, during post-irradiation regeneration, AA positively regulates intestinal epithelial cell proliferation by upregulating the expression of Ascl2 and activating WNT signaling, but negatively regulates intestinal epithelial cell differentiation. AA acts as a delicate regulator that efficiently facilitates epithelial tissue repair by activating radiation-resistant Msi1+ cells rather than Lgr5+ cells, which are extensively considered WNT-activated crypt base stem cells. Additionally, short-term AA treatment maintains optimal intestinal epithelial homeostasis under physiological conditions. As a result, AA treatment can be considered a potential therapy for irradiation injury repair and tissue regeneration.

Ileal proteomic changes associated with IL-25-mediated resistance against intestinal trematode infections

BackgroundEchinostoma caproni (Trematoda: Echinostomatidae) is an intestinal trematode, which has been extensively used to investigate the factors that determine the rejection of intestinal helminths. In this sense, several studies have shown that IL-25 is critical for the development of resistance against E. caproni in mice. In fact, treatment of mice with recombinant IL-25 generates resistance against primary E. caproni infection. However, the mechanisms by which IL-25 induces resistance remain unknown.MethodsTo study the mechanisms responsible for resistance elicited by IL-25, we analyzed the ileal proteomic changes induced by IL-25 in mice and their potential role in resistance. To this purpose, we compared the protein expression profiles in the ileum of four experimental groups of mice: naïve controls; E. caproni-infected mice; rIL-25-treated mice; and rIL-25-treated mice exposed to E. caproni metacercariae.ResultsQuantitative comparison by 2D-DIGE showed significant changes in a total of 41 spots. Of these, 40 validated protein spots were identified by mass spectrometry corresponding to 24 proteins.ConclusionsOur results indicate that resistance to infection is associated with the maintenance of the intestinal epithelial homeostasis and the regulation of proliferation and cell death. These results provide new insights into the proteins involved in the regulation of tissue homeostasis after intestinal infection and its transcendence in resistance.

LIF is essential for ISC function and protects against radiation-induced gastrointestinal syndrome

Leukemia inhibitory factor (LIF) is a cytokine essential for maintaining pluripotency of mouse embryonic stem cells. However, its role in adult intestinal stem cells (ISCs) is unclear. The adult intestinal epithelium has a high self-renewal rate driven by ISCs in crypts. Here, we find that LIF is present in the ISC niche in crypts and critical for the function of ISCs in maintaining the intestinal epithelial homeostasis and regeneration. Mechanistically, LIF maintains β-catenin activity through the AKT/GSK3β signaling to regulate ISC functions. LIF deficiency in mice impairs the renewal of the intestinal epithelium under the physiological condition. Further, LIF deficiency in mice impairs the regeneration of intestinal epithelium in response to radiation and shortens the lifespan of mice after high doses of radiation due to gastrointestinal (GI) syndrome, which can be rescued by administering recombinant LIF (rLIF). Importantly, LIF exhibits a radioprotective role in wild-type (WT) mice by protecting mice from lethal radiation-induced GI syndrome; administering rLIF promotes intestinal epithelial regeneration and prolongs survival in WT mice after radiation. These results reveal a previously unidentified and a crucial role of LIF in ensuring ISC function, promoting regeneration of the intestinal epithelium in response to radiation and protecting against radiation-induced GI syndrome.

Ageing, metabolism and the intestine.

The intestinal epithelium serves as a dynamic barrier to the environment and integrates a variety of signals, including those from metabolites, commensal microbiota, immune responses and stressors upon ageing. The intestine is constantly challenged and requires a high renewal rate to replace damaged cells in order to maintain its barrier function. Essential for its renewal capacity are intestinal stem cells, which constantly give rise to progenitor cells that differentiate into the multiple cell types present in the epithelium. Here, we review the current state of research of how metabolism and ageing control intestinal stem cell function and epithelial homeostasis. We focus on recent insights gained from model organisms that indicate how changes in metabolic signalling during ageing are a major driver for the loss of stem cell plasticity and epithelial homeostasis, ultimately affecting the resilience of an organism and limiting its lifespan. We compare findings made in mouse and Drosophila and discuss differences and commonalities in the underlying signalling pathways and mechanisms in the context of ageing.

SETDB1 is required for intestinal epithelial differentiation and the prevention of intestinal inflammation

ObjectiveThe intestinal epithelium is a rapidly renewing tissue which plays central roles in nutrient uptake, barrier function and the prevention of intestinal inflammation. Control of epithelial differentiation is essential to...

Abstract A12: L1CAM defines the regenerative origin of metastasis initiating cells in colorectal cancer

Abstract Metastatic cancers relapse due to the emergence of stemlike clones capable of reversible quiescence, tumour reinitiation, and therapy resistance termed metastasis-initiating cells (MICs). The origins of MICs and their relationship to primary tumor-initiating cells are not known, largely due to a lack of representative models of MICS. To directly scrutinize MICs in patient metastases, we established organoid cultures from therapy-resistant, residual colorectal cancer (CRC) liver metastases of patients undergoing cancer surgery. We identify the neuronal cell adhesion molecule L1 (L1CAM) as a critical mediator and marker of colorectal cancer (CRC) MICs. L1CAM+ cells are quiescent in well-structured neoplastic glands in vivo, but when dissociated from their epithelial niche, drive organoid regeneration and xenograft tumor reinitiation. FACS-sorted L1CAM+ cells preferentially regenerate heterogeneous organoids containing both L1CAM+ and L1CAM- progeny. Single-cell mRNA sequencing of 10,000 cells from four patient-derived organoids reveals that L1CAMhigh MICs in human CRC organoids partially overlap with a subset of LGR5high cancer stem cells (CSCs). The number of LGR5-expressing cells decreases, while the number of L1CAM-expressing cells increases in metastases in comparison with primary tumors in patients. We show that L1CAM is required for organoid formation, intestinal epithelial repair following colitis in vivo, regeneration of orthotopic rectal tumors, and metastatic colonization of the liver, but is dispensable for intestinal epithelial homeostasis or tumor initiation. Mechanistically, disruption of intercellular epithelial contacts inhibits an E-cadherin-REST signaling axis and enables transcriptional derepression of L1CAM. Our work underscores the distinct requirements of the CSCs that initiate tumor growth and MICs that drive lethal metastatic relapse. We identify the loss of epithelial integrity, an obligatory step of metastasis, as a crucial molecular driver of the transcriptional plasticity required for the emergence of prometastatic traits. Further, we define L1CAM as a crucial vulnerability of MICs that could be exploited therapeutically to treat patients with metastatic cancer. Citation Format: Karuna Ganesh, Harihar Basnet, Yasemin Kaygusuz, Ashley Laughney, Lan He, Roshan Sharma, Kevin O'Rourke, Vincent Reuter, Yun-Han Huang, Ignas Masilionis, Mesruh Turkekul, Ekrem Er, Katja Manova-Todorova, Leonard Saltz, Martin Weiser, Julio Garcia-Aguilar, Richard Koche, Scott Lowe, Dana Peer, Jinru Shia, Joan Massague. L1CAM defines the regenerative origin of metastasis initiating cells in colorectal cancer [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr A12.

Anthocyanin Improves Glucose Homeostasis in Obese Mice via Regulation of Intestinal Microbiota and Barrier Function

Abstract Objectives Anthocyanin (ACN) with low bioavailability possess anti-diabetic effect, the mechanism of the action remained unclear. This study aimed to investigate whether the anti-diabetic effects of anthocyanin were related to regulation of gut microbiota and epithelial barrier function. Methods Male C57Bl/6N mice were randomly assigned into 3 groups and fed chow, high fat/high sugar diet (HFHS, 45 kcal% fat, 17 kcal% sucrose) or HFHS + 0.4% anthocyanin for 8 weeks. Furthermore, a separate cohort of HFHS-fed C57Bl/6N mice were oral gavage with fecal microbiota from HFHS, HFHS + ACN and heat-killed fecal microbial from HFHS + ACN-fed donors (HK-ACN) for 4 weeks, The composition of gut microbiota, systemic inflammation, glucose homeostasis and intestinal barrier function were determined. Results The area under the curve (AUC) of the oral glucose tolerance test (GTT) and insulin tolerance test (ITT) in HFHS-fed mice were 61% and 79% higher than those in Chow-fed mice (P &amp;lt; 0.05). ACN supplementation improved HFHS-induced glucose intolerance in the GTT (P &amp;lt; 0.05) and restore insulin sensitivity in the ITT (P &amp;lt; 0.05). ACN supplementation also attenuated HFHS-induced systemic inflammation, intestinal permeability and metabolic endotoxemia (P &amp;lt; 0.05). In addition, ACN administration increased expression of epithelial barrier function genes ZO-1, Occludin and decreased inducible NO-synthase (iNOS) protein levels in intestine of HFHS-fed mice. Gut microbiota analysis showed that ACN greatly altered gut microbiota of HFHS-fed mice. Transplantation of the gut microbiota from ACN-fed mice, but not HFHS-fed or HK-ACN-fed mice, significantly improved intestinal epithelial barrier function, systemic inflammation and glucose homeostasis. Conclusions The present study demonstrated that ACN improves HFHS-induced impairment of glucose metabolism by regulating the gut microbiota and epithelial barrier functions. Funding Sources No funding sources to report.

Proteome changes induced by a short, non-cytotoxic exposure to the mycoestrogen zearalenone in the pig intestine

Intestinal epithelial homeostasis is regulated by a complex network of signaling pathways. Among them is estrogen signaling, important for the proliferation and differentiation of epithelial cells, immune signaling and metabolism. The mycotoxin zearalenone (ZEN) is an estrogen disruptor naturally found in food and feed. The exposure of the intestine to ZEN has toxic effects including alteration of the immune status and is possibly implicated in carcinogenesis, but the molecular mechanisms linked with these effects are not clear. Our objective was to explore the proteome changes induced by a short, non-cytotoxic exposure to ZEN in the intestine using pig jejunal explants. Our results indicated that ZEN promotes little proteome changes, but significantly related with an induction of ERα signaling and a consequent disruption of highly interrelated signaling cascades, such as NF-κB, ERK1/2, CDX2 and HIF1α. The toxicity of ZEN leads also to an altered immune status characterized by the activation of the chemokine CXCR4/SDF-1 axis and an accumulation of MHC-I proteins. Our results connect the estrogen disrupting activity of ZEN with its intestinal toxic effect, associating the exposure to ZEN with cell-signaling disorders similar to those involved in the onset and progression of diseases such as cancer and chronic inflammatory disorders. SignificanceThe proteomics results presented in our study indicate that the endocrine disruptor activity of ZEN is able to regulate a cascade of highly inter-connected signaling events essential for the small intestinal crypt-villus cycle and immune status. These molecular mechanisms are also implicated in the onset and progress of intestinal immune disorders and cancer indicating that exposure to ZEN could play an important role in intestinal pathogenesis.

MON-726 Modifications of FOXO1 and GATA4-NKX2.5 Signaling Induce Human Enteroendocrine Differentiation

Enteroendocrine (EE) cells are the most abundant hormone-producing cells in the human body and are vital for metabolism, as well as intestinal and pancreatic function. They have been implicated in the pathogenesis of multiple diseases including diabetes mellitus. Although recent studies have identified multiple signaling pathways (including Wnt, MAPK, BMP and Notch) that can induce low levels of EE cell differentiation, the production of functional human EE cells in vitro remains challenging, making their study and therapeutic utilization difficult. To improve this, we employed the human intestinal organoid culturing system, as it mimics intestinal epithelial homeostasis, allowing for differentiation of multiple epithelial cell types. Using a small scale, directed screen, we targeted multiple transcriptional regulators, using small molecules known to control pancreatic and intestinal development, and hormone production. We chose small molecules instead of gene editing tools to avoid the potential pitfall of off-target mutagenesis.We found that inhibition of FoxO1 in our organoid culture led to an increase in EE cell differentiation as assessed by EE-specific gene expression, with a 5-10 fold upregulation in expression of ChgA, NeuroD1, and Neurog3 compared to whole mucosal biopsies (P<0.01 for all targets, n=3 per group). Flow cytometry data showed 6-8% of cells produced CHGA, compared to 0.2% in undifferentiated organoids (P<0.0001, n=3 per group), and the 1% typically seen in the duodenum. We also noted a corresponding increase in the production of EE hormones, including glucose-dependent insulinotropic peptide (GIP), serotonin and somatostatin, by qPCR and immunofluorescence. Analysis of conditioned media using ELISA, compared to undifferentiated organoids, revealed increased serotonin (362.6±52.3 vs 167.5±5.1 ng/mL, P=.0037, n=3 per group) and GIP (5.76±1.31 pg/mL vs undetectable, n=3 per group). Independently, upregulation of GATA4-Nkx2.5 also induced EE cell differentiation and hormone production, although to a lesser extent than FoxO1 inhibition. The exception to this was GIP, which showed increased expression and production with GATA4-Nkx2.5 compared to FoxO1 inhibition (20.8±7.4 vs 5.8±1.3 pg/mL, n=3 per group), with a much larger increase when FoxO1 inhibition was followed by GATA4-Nkx2.5 activation (53.4±4.8 pg/mL, n=3). Of note, all experiments were performed in a minimum of three human lines.Taken together, our data have identified multiple factors, including inhibition of FoxO1 and activation of GATA4-Nkx2.5, that can drive ex vivo human EE cell differentiation, with unique hormone production profiles, when targeted via small molecules. This is a critical first step towards understanding the role of enteroendocrine cells in disease and the development of EE cell-based therapies.

Mitochondria regulate intestinal stem cell proliferation and epithelial homeostasis through FOXO.

A metabolic transition from glycolysis to oxidative phosphorylation is often associated with differentiation of many types of stem cells. However, the link between mitochondrial respiration and stem cells’ behavior is not fully understood. We genetically disrupted electron transport chain (ETC) complexes in the intestinal stem cells (ISCs) of Drosophila. We found that ISCs carrying impaired ETC proliferated much more slowly than normal and produced very few enteroblasts, which failed to further differentiate into enterocytes. One of the main impediments to ISC proliferation and lineage specification appeared to be abnormally elevated forkhead box O (FOXO) signaling in the ETC-deficient ISCs, as genetically suppressing the signaling pathway partially restored the number of enterocytes. Contrary to common belief, reactive oxygen species (ROS) accumulation did not appear to mediate the ETC mutant phenotype. Our results demonstrate that mitochondrial respiration is essential for Drosophila ISC proliferation and lineage specification in vivo and acts at least partially by repressing endogenous FOXO signaling.

Canonical and non-canonical Wnt signaling are simultaneously activated by Wnts in colon cancer cells

The Wnt signaling pathway is a crucial regulator of the intestinal epithelium homeostasis and is altered in most colon cancers. While the role of aberrant canonical, β-catenin-dependent Wnt signaling has been well established in colon cancer promotion, much less is known about the role played by noncanonical, β-catenin-independent Wnt signaling in this type of cancer. This work aimed to characterize the noncanonical signal transduction pathway in colon cancer cells. To this end, we used the prototype noncanonical ligand, Wnt5a, in comparison with Wnt3a, the prototype of a canonical β-catenin activating ligand. The analysis of the expression profile of Wnt receptors in colon cancer cell lines showed a clear increase in both level expression and variety of Frizzled receptor types expressed in colon cancer cells compared with non-malignant cells. We found that Wnt5a activates a typical Wnt/Ca++ - noncanonical signaling pathway in colon malignant cells, inducing the hyperphosphorylation of Dvl1, Dvl2 and Dvl3, promoting Ca++ mobilization as a result of phospholipase C (PLC) activation via pertussis toxin-sensitive G-protein, and inducing PLC-dependent cell migration. We also found that while the co-receptor Ror2 tyrosine kinase activity is not required for Ca++ mobilization-induced by Wnt5a, it is required for the inhibitory effects of Wnt5a on the β-catenin-dependent transcriptional activity. Unexpectedly, we found that although the prototype canonical Wnt3a ligand was unique in stimulating the β-catenin-dependent transcriptional activity, it also simultaneously activated PLC, promoted Ca++ mobilization, and induced Rho kinase and PLC-dependent cell migration. Our data indicate, therefore, that a Wnt ligand can activate at the same time the so-called Wnt canonical and noncanonical pathways inducing the formation of complex signaling networks to integrate both pathways in colon cancer cells.

SREBPs in the control of intestinal homeostasis and tumorigenesis

Lipids are key regulators of the growth and differentiation of intestinal epithelial cells, including intestinal stem cells (ISC). High‐fat diets and obesity are serious risk factors for gastrointestinal cancers in humans. Sterol regulatory element‐binding proteins (SREBPs) are transcription factors that control lipid synthesis and uptake in the intestine. Our recent studies have shown that SREBPs and their major products, sterols (from SREBP‐2) and unsaturated fatty acids (from SREBP‐1) have a critical role in sustaining growth of the intestinal epithelia: (1) The loss of Scap, which controls intracellular cleavage events required for SREBP activation, reduces lipid synthesis and causes a lethal intestinal injury in mice characterized by a loss of intestinal crypts, which house ISCs. (2) When SREBP‐2 is inactivated in intestinal epithelium, hyperplasia of intestinal crypts and hypertrophy of the intestine results. In SREBP‐2 knockouts, sterol synthesis falls while unsaturated fatty acid synthesis does not, owing to an increase in SREBP‐1. (3) Despite this overgrowth phenotype, ablation of SREBP‐2 in intestinal epithelia of mice blocks the formation of adenomas caused by mutant APC. These findings indicate that SREBPs govern intestinal epithelial homeostasis in complex ways. We propose that SREBPs maintain homeostasis of intestinal epithelia by producing specific, yet‐to‐be identified lipid metabolites that sustain the proliferation of ISCs. These pathways control intestinal organ size and malignant potential, which bears on human diseases of under‐ or over‐proliferation of the intestine, such as short gut syndrome and colorectal cancer, the 3rd most common cause of cancer death in the U.S.Support or Funding InformationNIH‐R01DK121842, NIH‐K08DK102652

Stromal Cells in the Pathogenesis of Inflammatory Bowel Disease.

Up till now, research on inflammatory bowel disease [IBD] has mainly been focused on the immune cells present in the gastrointestinal tract. However, recent insights indicate that stromal cells also play an important and significant role in IBD pathogenesis. Stromal cells in the intestines regulate both intestinal epithelial and immune cell homeostasis. Different subsets of stromal cells have been found to play a role in other inflammatory diseases [e.g. rheumatoid arthritis], and these various stromal subsets now appear to carry out also specific functions in the inflamed gut in IBD. Novel potential therapies for IBD utilize, as well as target, these pathogenic stromal cells. Injection of mesenchymal stromal cells [MSCs] into fistula tracts of Crohn’s disease patients is already approved and used in clinical settings. In this review we discuss the current knowledge of the role of stromal cells in IBD pathogenesis. We further outline recent attempts to modify the stromal compartment in IBD with agents that target or replace the pathogenic stroma.

Lactobacillus reuteri maintains intestinal epithelial regeneration and repairs damaged intestinal mucosa

ABSTRACT Little is known about the regulatory effect of microbiota on the proliferation and regeneration of ISCs. Here, we found that L. reuteri stimulated the proliferation of intestinal epithelia by increasing the expression of R-spondins and thus activating the Wnt/β-catenin pathway. The proliferation-stimulating effect of Lactobacillus on repair is further enhanced under TNF -induced intestinal mucosal damage, and the number of Lgr5+ cells is maintained. Moreover, compared to the effects of C. rodentium on the induction of intestinal inflammation and crypt hyperplasia in mice, L. reuteri protected the intestinal mucosal barrier integrity by moderately modulating the Wnt/β-catenin signaling pathway to avoid overactivation. L. reuteri had the ability to maintain the number of Lgr5+ cells and stimulate intestinal epithelial proliferation to repair epithelial damage and reduce proinflammatory cytokine secretion in the intestine and the LPS concentration in serum. Moreover, activation of the Wnt/β-catenin pathway also induced differentiation toward Paneth cells and increased antimicrobial peptide expression to inhibit C. rodentium colonization. The protective effect of Lactobacillus against C. rodentium infection disappeared upon application of the Wnt antagonist Wnt-C59 in both mice and intestinal organoids. This study demonstrates that Lactobacillus is effective at maintaining intestinal epithelial regeneration and homeostasis as well as at repairing intestinal damage after pathological injury and is thus a promising alternative therapeutic method for intestinal inflammation.

A259 HIGH-FAT/HIGH-CHOLESTEROL DIETS PREDISPOSE THE HOST TO EXACERBATED ENTERIC INFECTIONS

Abstract Background High-fat/high-cholesterol diets are a well established risk factor for cardiovascular and metabolic diseases, given their propensity to trigger perturbations ranging from altering whole body lipid profile to the induction of intestinal dysbiosis. However much less is known about their effects on the host’s susceptibility to enteric infections. Aims To determine the effects of high-fat/high-cholesterol diets over the host’s susceptibility to enteric bacterial infections and identify the underlying molecular mechanisms. Methods C57BL/6 mice were given two different high-fat/high-cholesterol diets; HFHC (40% kcal fat, 1.25% cholesterol) or HFHCC (40% kcal fat, 1.25% cholesterol, 0.5% sodium cholate) and a control, normal diet (ND, 10% kcal fat, 0% cholesterol, 0% sodium cholate). After four weeks of administration, animals were euthanized and colonic tissue samples taken for histology, immunofluorescence, gene expression analyses, total protein lysates and microbiome sequencing (16S). A separated group of animals was gavaged with FITC-dextran to measure intestinal permeability. Mice fed with the diets for three weeks were infected with ~5x108 cfu of Citrobacter rodentium DBS100/StrpR by oral gavage, and kept on the corresponding diets after the infections. Bacterial shedding in the feces was followed for up to 30 days after infection. Results Administration of the HFHC and HFHCC diets caused an increase in intestinal permeability. Colonic sections stained with H&amp;E and alcian blue evidenced a decreased in the number of mucin-filled goblet cells and a thinner mucus layer, suggesting a defect in the assembly and/or stability of the mucus layer. Expression analyses revealed a drop in the mRNA levels of Muc1 and Muc2, suggesting reduced mucin production. The concentration of IgA was slightly reduced in colon lysates and the transcript levels of the antimicrobial peptide genes Ang4, Leap2 and Cramp were also significantly reduced. Immunofluorescent microscopy showed that goblet cell granules of HFHC- and HFHCC-fed mice were devoid of Relmβ and Tff3, indicating defective production of those two factors critical for intestinal epithelial defense and homeostasis. Collectively, our results suggest that HFHC and HFHCC diets induce differentiation and functional defects in goblet cells. Fecal shedding of C. rodentium showed an increased bacterial burden in HFHC- and HFHCC-fed animals, indicating a more aggressive bacterial infection, accompanied by increased epithelial damage. Conclusions Consumption of high-fat/high cholesterol diets perturb the colonic homeostasis and alter intestinal defenses and the integrity of the intestinal barrier, predisposing the host to a higher susceptibility to enteric infections. Funding Agencies CIHRNSERC