bluuebunny/crossref_metadata_2025_split huggingface dataset:
A23 IMPAIRMENT OF INTESTINAL CRYPTAL STEM CELL CYCLE AND PANETH CELL PROLIFERATION BY CHRONIC STRESS
Authors: Bai X , Ihara E , Ogawa Y 
Published: February 2024

Abstract

Background
The small intestine’s epithelial barrier is essential for intestinal homeostasis and host defense. It consists of epithelial cells that are constantly renewed by crypt stem cells and their progeny, which have different functions in the gut. Chronic stress can impair the barrier by modulating the gut-brain axis, which involves the hypothalamic-pituitary-adrenal (HPA) axis, the vagus nerve, and the immune system. Chronic stress can also alter the gut microbiota, which can further influence the barrier and immune system.

Aims
The present study examines how chronic stress affects the crypt stem cell niche and function in the small intestine.

Methods
Restraint stress was applied to 6-8 weeks old mice in a plastic container for 1 hour daily for 7 days. The dark/light transition box test was used to evaluate the stress-induced behavior change. The Ussing chamber technique was used to assess the paracellular permeability of the small intestine. RNA was extracted from the mucosa and submucosa layers of the small intestine, after removing the muscle layer, and RNA sequencing analysis was performed. Immunofluorescence analysis was also performed on the small intestinal tissues to detect the expression of various markers. To assess the in vivo effect of cortisol, dexamethasone (a synthetic steroid activating same receptors as cortisol), or metyrapone (an 11β-hydroxylation blocker to inhibit cortisol production), was applied to the mice.

Results
We found that restraint stress induced anxiety-like behavior, increased small intestinal epithelial permeability, and reduced villus and crypt length in mice. RNA sequencing analysis revealed that restraint stress upregulated steroid-driven pathways and downregulated cell cycle and cell mitosis pathways in the small intestinal mucosa, suggesting increased cortisol production and decreased stem cell activity. Immunofluorescence analysis confirmed that restraint stress reduced the expression of stem cell marker Olfm-4 and proliferation markers Ki-67 and p-H3 in the small intestinal crypts. Dexamethasone, a synthetic glucocorticoid, mimicked the effects of restraint stress on crypt stem cell proliferation, while metyrapone, a steroid synthesis inhibitor, attenuated the effects of restraint stress on crypt stem cell proliferation. Restraint stress and dexamethasone also reduced Paneth cell RNA expression and marker in the small intestine.

Conclusions
Chronic stress induces anxiety, barrier dysfunction, and epithelial shortening. Chronic stress also alters crypt stem cell and Paneth cell expression. These findings show how chronic stress impacts intestinal epithelial renewal and differentiation, which may affect intestinal health and disease.

Funding Agencies
JSPS
A24 AN ULCERATIVE COLITIS-ISOLATED PATHOBIONT CAN DEGRADE MUCUS PRODUCED BY UC PATIENT-DERIVED COLONOIDS
Authors: Gilliland A , Chen Y , Tertigas D , Surette M , Bressler B , Vallance B 
Published: February 2024

Abstract

Background
Inflammatory bowel disease (IBD) pathobionts are commensal microbes with pathogenic potential that may cause or exacerbate IBD symptoms. Some pathobionts (ex. Escherichia coli) reside at low levels in the lumen of a healthy gut but can rapidly grow in the inflamed colons of ulcerative colitis (UC) patients. To promote disease, these pathobionts must cross the colonic mucus barrier (comprised of MUC2) that separates the epithelium from luminal microbes. It is currently unclear how bacterial pathobionts cross the mucus barrier of UC patients.

Aims
Using healthy and UC patient biopsy-derived colonic organoids (colonoids) and an air liquid interface (ALI) monolayer model, we investigated how the UC-isolated E. coli pathobiont p19A crosses the mucus barrier.

Methods
Apical out healthy and UC patient biopsy-derived colonoids were infected with p19A to confirm this pathobiont exerts direct cytopathic effects on human colonocytes. Sequencing p19A’s genome, we found it contains two mucus degrading proteins (mucinases). Healthy human and UC colonoids, as well as mouse colonoids, were used to generate mucus-producing ALI monolayers. To detect p19A-mediated mucus degradation, concentrated p19A supernatant was incubated with ALI-derived mucus and degraded MUC2 proteins were detected by protein gel and MUC2 Western blot. MUC2 glycosylation was analyzed by protein gel and PAS staining. ALI monolayers were infected with p19A to evaluate mucus degradation and p19A localization by immunostaining.

Results
The UC-isolated pathobiont p19A infected and exerted cytotoxic effects on apical out healthy and UC patient colonoids. By day 14, ALI monolayers were differentiated and covered by a thick mucus layer as assessed using brightfield microscopy. Mucus removed for mucinase assays was replenished within 7 days. p19A harbours proteins capable of degrading human ALI-, but not mouse ALI-derived mucus, in vitro, suggesting the presence of host-specific mucinases. Mucus produced by UC ALI monolayers showed reduced glycosylation and increased degradation both over time and by p19A proteins. Day 21 ALI-monolayers infected with p19A for 18 hours exhibited overt mucus degradation allowing p19A to infect the underlying epithelium.

Conclusions
Patient-derived ALI monolayers produce a thick mucus layer that can be used to study pathobiont-mucus interactions. The UC pathobiont p19A disrupts apical out organoids and produces proteins that degrade ALI-derived mucus in vitro, with UC mucus being more susceptible to degradation than mucus from healthy controls. The results from our model suggest a potential mechanism for pathobiont-mediated mucosal barrier disruption in UC patients.

Patient-derived ALI monolayer (blue/white) produces a thick mucus layer (green) that can be degraded by the pathobiont p19A (red).

Funding Agencies
CCC, CIHR
A25 DETECTION OF BACTERIAL ADP-HEPTOSE BY INTESTINAL STEM CELLS DRIVES EPITHELIAL REGENERATION FROM PANETH CELL DEDIFFERENTIATION
Authors: Goyal S , Gray-Owen S , Girardin S E 
Published: February 2024

Abstract
NOT PUBLISHED AT AUTHOR’S REQUEST
Please acknowledge all funding agencies listed below
Funding Agencies
CCC, CIHR
A27 ROLE OF HIPPO SIGNALING EFFECTORS YAP & TAZ IN GASTRIC METAPLASIA AND TUMORIGENESIS
Authors: Ju Z , Farag M , Haddad M , Fiset P , Lorenzo L , Gregorieff A 
Published: February 2024

Abstract
NOT PUBLISHED AT AUTHOR’S REQUEST
Please acknowledge all funding agencies listed below
Funding Agencies
CIHR
A28 A SERUM REDOX STATUS-RELATED INDICATOR IS ASSOCIATED WITH THE RISK OF ONSET OF CROHN'S DISEASE
Authors: Mu K , Xue M , Turpin W , Croitoru K 
Published: February 2024

Abstract

Background
Crohn's disease (CD) is a gastrointestinal disorder characterized by chronic inflammation. While increased oxidative stress is observed in established CD patients, it remains unknown whether a shift in redox status is present before the diagnosis of CD and whether it is correlated with changes in immune response and microbial composition. Our hypothesis is that oxidative stress plays a role in the development of CD, and it could be detected before the diagnosis of CD. Furthermore, it is likely to be correlated with systemic inflammation and alterations in gut microbiota composition.

Aims
We aimed to assess the relationship between the serum redox status-related indicators, specifically amino acids ratios, with risk of CD onset and if this is further association with systemic inflammation marker, and gut microbiota composition.

Methods
In the Genetic Environment Microbial Project (CCC-GEM), a cohort of first-degree relatives (FDRs) of CD patients was prospectively followed. Among them, we identified subjects who later developed CD, defined as pre-CD (n=69), and matched them at a 1:4 ratio with FDRs who remained disease-free (n=276). Serum levels at enrollment of cysteineglycine (reduced form) and cystineglycine (oxidized form) were quantified by mass spectrometry, and the cysteineglycine/cystineglycine ratio was used as an indicator of redox status. Conditional logistic regression assessed the association with CD, while partial Spearman regression evaluated its correlation with systemic inflammation, as indicated by c-reactive protein (CRP), and gut microbiota composition (determined by fecal 16S rRNA sequencing).

Results
A decrease in the ratio indicates a shift in redox status toward oxidative stress. The cysteineglycine/cystineglycine ratio was negatively associated with the likelihood of developing CD (coefficient = -0.6188; p =0.0146), and it was also negatively correlated with CRP levels (coefficient = -0.177; p =0.00095). A list of taxa belonging to the phyla Firmicutes and Actinobacteriota were positively correlated with cysteineglycine/cystineglycine ratio (p≤0.05).

Conclusions
This study is the first to report that when redox status shifts towards oxidative stress, as indicated by the cysteineglycine/cystineglycine ratio, the likelihood of CD increases. Furthermore, these markers also correlate with CRP levels and gut microbiota composition, indicating a loss of various taxa when the redox status shifts towards oxidative stress.
Submitted on behalf of the CCC-GEM consortium.

Funding Agencies
CCC, CIHRHCT
A29 INVESTIGATING THE IMPACT OF PARKINSON’S DISEASE-ASSOCIATED GENES ON INTESTINAL HOMEOSTASIS
Authors: Pei J , Recinto S , MacDonald A , Kazanova A , Gavino C , Trudeau L , Desjardins M , Stratton J , Gruenheid S 
Published: February 2024

Abstract

Background
Intestinal epithelial cells (IECs) provide an essential physical barrier between harsh luminal contents and underlying host tissue. The maintenance of intestinal homeostasis must be intricately regulated through the proliferation and differentiation of intestinal stem cells (ISCs). Dysregulation of this system results in the loss of barrier function, causing pathologies in both intestinal and extra-intestinal diseases. While Parkinson’s Disease (PD) is primarily a neurodegenerative disorder, there is increasing evidence linking PD progression and gastrointestinal (GI) dysfunction. Constipation and increased bowel permeability are often observed years prior to development of motor dysfunction in PD, and people with inflammatory bowel disease are more likely to develop PD. Our group developed a model to investigate the role of the gut in PD, demonstrating that mice with genetic ablation of the PD-associated gene Pink1 exhibited motor phenotypes only when previously infected with Gram-negative Citrobacter rodentium intestinal bacteria. As Pink1 and other PD-associated genes are expressed in IECs, we hypothesize that PD-associated gene mutations directly affect the epithelium and impact early PD pathophysiology.

Aims
1. Determine how Pink1 mutation affects the transcriptional profiles in IECs at steady state and in vivo C. rodentium infection.
2. Use colonoid systems to study the effect of Pink1 mutation on epithelial activity.

Methods
ScRNAseq was performed on IECs isolated from uninfected and infected Pink1 WT and KO mice, sacrificed at day 7 and 14 (early and peak infection) to elucidate transcriptional differences between epithelial lineages of each genotype. Additionally, colonoids were derived from primary mouse tissue and treated with lipopolysaccharide (LPS) to determine how Pink1 KO affects the inflammatory response of the epithelium.

Results
Our data revealed that Pink1 KO profoundly affected several epithelial lineages. ISCs from infected Pink1 KO mice had dysregulated cell cycle genes, transit amplifying cells showed dysregulated expression of tight junction genes, and enterocytes displayed dysregulation in oxidative damage and apoptotic genes. Preliminary data from colonoids showed that Pink1 KO mice, when stimulated with LPS, had altered pro-inflammatory cytokine gene expression. We are also evaluating colon motility differences between genotypes through transit time assays, counting fecal pellets per hour, and fecal water content.

Conclusions
In Pink1 KO IECs, there is indeed an altered cellular response upon infection, but more information is needed to decern the mechanistic role of IECs in PD. By investigating the role of PD genes in the GI tract, these studies carry important implications for understanding the initiation and progression of PD.

Funding Agencies
CIHR
A30 MICROBIOTA AND NOD2 REGULATE SMALL INTESTINAL RESTITUTION THROUGH FETAL-LIKE INTESTINAL STEM CELLS
Authors: Tsang D , Maisonneuve C , Ayyaz A , Foerster E , Nissan M , Baerg L , Trcka D , Streutker C , Wrana J L , Girardin S E , Philpott D 
Published: February 2024

Abstract

Background
Inflammatory bowel disease is characterized by chronic inflammation of the gastrointestinal tract, resulting in recurrent injury to the intestinal epithelium. Restitution of the small intestinal epithelium is a coordinated response that involves the dedifferentiation of epithelial cell lineages, proliferation of Lgr5+ intestinal stem cells, and fetal-like stem cell reversion. While gut microbiota are critical mediators of intestinal inflammation, their impact on epithelial restitution remains unclear.

Aims
We aim to identify the how microbes regulate small intestinal epithelial restitution following damage. Our hypothesis is that gut microbiota accelerate restitution through pattern recognition receptor-driven signals following fetal-like stem cell reversion.

Methods
Irradiation (IR, 12Gγ) was used to induce a synchronized small intestinal epithelial restitution response in mice. Intestinal restitution kinetics were assessed transcriptionally (scRNA-Seq, qPCR) and histologically. Small intestinal organoids were used to assess epithelial restitution kinetics in vitro.

Results
ScRNA-Seq of small intestinal epithelial cells following IR from germ-free mice (GF), and specific pathogen-free mice (SPF) mice revealed that microbiota induced greater expression of fetal-like stem cell reversion markers, Ly6a and Clu, and greater expression of the proliferation marker, Pcna. These results were supported histologically as irradiated SPF mice observed an increase in fetal-like stem cells marked by Ly6a and Clu and an increase BrdU+ proliferating cells. ScRNA-seq and in situ hybridization highlighted that fetal-like intestinal stem cells upregulate expression of Nod2, a bacterial pattern recognition receptor. Using intestinal organoids to assess the function of Nod2, we observed that muramyl dipeptide driven Nod2-signlaing potentiates an interferon gene signature following IFNγ and TNFα co-stimulation. Further supporting the role for Nod2 in intestinal restitution, intestinal epithelium specific Nod2KO mice decreased BrdU+ proliferating cells post-IR compared to littermate controls.

Conclusions
Microbiota promote small intestinal restitution following IR through Nod2-signaling in fetal-like intestinal stem cells.

Funding Agencies
CIHR
A31 DYSPLASIA IN COLONIC POLYPS: PREPARING FOR A RESECT AND DISCARD STRATEGY IN CANADA
Authors: Patel V , Bechara R , Rai M S 
Published: February 2024

Abstract

Background
While diminutive colorectal polyps have a negligible cancer risk, current management involves resecting and submitting all polyps for histological assessment. This is a substantial burden and cost to the healthcare system. The European Society of Gastrointestinal Endoscopy (ESGE) has recommended a “resect-and-discard strategy” without histological evaluation as an acceptable strategy when high-confidence endoscopic characterization of colorectal polyps is achieved. However, a resect and discard strategy has not been adopted yet in Canada.

Aims
The objective of this study was to determine the prevalence and characteristics of polyps removed at our center based on size.

Methods
We retrospectively reviewed colonoscopies and pathology reports at Kingston Health Sciences Center from January to December 2020. Based on the pathology report, polyps were classified as diminutive (1-5mm), small (6-9mm) or large (ampersand:003E or = 10mm). We recorded histology and the presence of high-grade dysplasia (HGD) or cancer.

Results
Out of 2218 colonoscopies, 2945 polyps were removed. 1703 (57.8%) polyps were diminutive with only two (0.1%) having focal HGD and none having cancer. 699 (23.7%) polyps were classified as small with three (0.4%) having HGD and none having cancer. The large polyp group had 543 (18.4%) polyps, of which 87 (16%) showed HGD and 15 (2.8%) exhibited cancer. The specific histologic findings are shown in Table 1. Endoscopy reports specifically mentioned concern of dysplasia in one out of the five polyps with HGD in the diminutive and small groups.

Conclusions
As expected, most polyps were either diminutive or small (81.5%). Neither of these groups had cancer and only 5 had HGD. Adopting a resect and discard strategy at our center for diminutive polyps has the potential for significant cost savings with negligible risk of missing a high-risk polyp. The next steps would involve assessing optical diagnosis sensitivity and specificity for diminutive and small polyps.
Histological characterization of polyps within each size range

Funding Agencies
None
A32 CATHELICIDIN REGULATES GOBLET CELL MUCUS SECRETION DURING CITROBACTER RODENTIUM-INDUCED COLITIS
Authors: Cobo E R , Blyth G , Fiorani F , Lahiri P , Herik A , Dufour A , Chadee K 
Published: February 2024

Abstract

Background
Colonic goblet cells by secreting Muc2 mucin and specific proteins is critical for physically entrapping and expelling invading enteropathogens. Thus, is not surprising that Muc2-/- littermates exhibit increased susceptibility to attaching/effacing Citrobacter rodentium colonization. The colonic epithelium also secretes small cathelicidin peptide, which potentially interacts intimately with goblet cells and was presumed to accumulate within the sterile inner mucus layer as a simple antimicrobial peptide defense.

Aims
To determine the effects of cathelicidin on mucin secretion in goblet cells during C. rodentium-induced colitis and the impact on the mucus barrier defense.

Methods
We used cathelicidin-deficient (Camp-/-) mice, mouse colonoids and human colonic LS174T like-goblet epithelial cells to elucidate the mechanisms by which cathelicidin regulates goblet cell secretions.

Results
Camp -/- littermates infected with C. rodentium displayed increased fecal shedding and epithelial colonization. Camp-/- littermates at the peak of C. rodentium infection (7 dpi) showed a deficient mucin layer with fewer Alcian blue/PAS filled goblet cells and a reduction in fucose (UEA-1+) and N-acetylglucosamine (WGA+) glycoproteins. By transmission electron microscopy (TEM), goblet cells in Camp-/- colons were swollen and retained a large number of mucus granules during C. rodentium infection. C. rodentium infected Camp-/- littermates showed impaired reactive oxygen species (ROS) production and a transcriptomic profiling associated with decreased ROS biosynthesis and an increase in ROS negative regulators. In mucin producing LS174T colonic epithelial cells, human cathelicidin LL-37 promptly induced the secretion of goblet cell-associated TFF3 and RELMβ, via a ROS-dependent mechanism.

Conclusions
These findings revealed that mice lacking cathelicidin (Camp-/-) were more susceptible to C. rodentium colonization caused by defective goblet cell mucus and mucin-associated protein secretion via a ROS-dependent mechanism. Importantly, cathelicidin regulated mucus secretion revealing a non microbicidal action of this peptide with homeostatic properties on the colonic mucus barrier, critical in excluding luminal microbiota away from the epithelia to clear bacterial infections and restore gut homeostasis.

Funding Agencies
NSERC
A33 EOSINOPHILS PROMOTE SALMONELLA TYPHIMURIUM COLONIZATION IN THE MURINE SMALL INTESTINE
Authors: FitzPatrick R D , Noone J R , Cartwright R A , Gatti D M , Brosschot T , Lane J M , Zafarullah M , Kroker Kimber I , Reynolds L 
Published: February 2024

Abstract

Background
During inflammatory bowel disease (IBD) both the number of intestinal eosinophils and susceptibility to enteric bacterial infections can drastically increase. It is unknown, however, whether eosinophils recruited to the tissues lining the gastrointestinal (GI) tract during severe disease help to control pathogenic bacterial colonization or whether their presence promotes bacterial infection. No studies to date have examined the role of eosinophils during an acute bacterial infection in the small intestine.

Aims
Our research aims to uncover the role of eosinophils during an enteric Salmonella infection in mice and we hypothesize that eosinophils undergo phenotypic changes in the presence of Salmonella and aid in the clearance of this enteric bacterial pathogen.

Methods
To assess how the presence of Salmonella impacts the frequency and phenotype of eosinophils we performed flow cytometry on eosinophils isolated from mice 3-days post-oral infection with Salmonella enterica serovar Typhimurium and compared these findings to naïve BALB/c controls. To determine if eosinophils are essential for controlling Salmonella infection we quantified levels of Salmonella colonization along the GI tract, liver and spleen of wild-type and eosinophil-deficient (ΔdblGATA) littermate mice by plating homogenized tissues on LB agar.

Results
Following an oral Salmonella infection we found higher frequencies of eosinophils in the gut-draining mesenteric lymph nodes compared to uninfected mice. Intestinal lamina propria-resident eosinophils in the small intestine and colon displayed altered phenotypes following infection in an intestinal-region specific manner: during infection, in the small intestine eosinophils increased their expression of MHCII, and in the colon eosinophils displayed elevated granularity. When wild-type and ΔdblGATA mice were pre-treated with the antibiotic streptomycin 1-day prior to infection to confer robust Salmonella burdens in the small intestine we found a significant reduction in Salmonella colonization in the ileum of the small intestine in ΔdblGATA mice compared to their wild-type littermates. These data suggest eosinophils have a role in promoting colonization by this enteric bacterial pathogen.

Conclusions
Our data contribute to the growing evidence that microbes can influence the phenotype of intestinal eosinophils and demonstrate that the influence of and requirements for eosinophils during bacterial infection is highly context-dependent. We found that eosinophils are not essential for controlling an acute enteric Salmonella infection and instead they promote Salmonella colonization in the small intestine.

Funding Agencies
CIHRTRIANGLE