Colonic mucosal and serum expression of microRNAs in canine large intestinal inflammatory bowel disease.

BACKGROUND: Canine inflammatory bowel disease (IBD) is a group of chronic gastrointestinal (GI) disorders of still largely unknown etiology. Canine IBD diagnosis is time-consuming and costly as other diseases with similar signs should be initially excluded. In human IBD microRNA (miR) expression changes have been reported in GI mucosa and blood. Thus, there is a possibility that miRs may provide insight into disease pathogenesis, diagnosis and even treatment of canine IBD. The aim of this study was to determine the colonic mucosal and serum relative expression of a miRs panel in dogs with large intestinal IBD and healthy control dogs. RESULTS: Compared to healthy control dogs, dogs with large intestinal IBD showed significantly increased relative expression of miR-16, miR-21, miR-122 and miR-147 in the colonic mucosa and serum, while the relative expression of miR-185, miR-192 and miR-223 was significantly decreased. Relative expression of miR-146a was significantly increased only in the serum of dogs with large intestinal IBD. Furthermore, serum miR-192 and miR-223 relative expression correlated to disease activity and endoscopic score, respectively. CONCLUSION: Our data suggest the existence of dysregulated miRs expression patterns in canine IBD and support the potential future use of serum miRs as useful noninvasive biomarkers.

Bacterial Biogeography of the Colon in Dogs With Chronic Inflammatory Enteropathy.

The intestinal microbiota is believed to play a role in the pathogenesis of inflammatory bowel disease in humans and chronic inflammatory enteropathy (CIE) in dogs. While most previous studies have described the gut microbiota using sequencing methods, it is fundamental to assess the spatial distribution of the bacteria for a better understanding of their relationship with the host. The microbiota in the colonic mucosa of 22 dogs with CIE and 11 control dogs was investigated using fluorescence in situ hybridization (FISH) with a universal eubacterial probe (EUB338) and specific probes for select bacterial groups. The number of total bacteria labeled with EUB338 probe was lower within the colonic crypts of dogs with CIE compared to controls. Helicobacter spp. and Akkermansia spp. were decreased on the colonic surface and in the crypts of dogs with CIE. Dogs with CIE had increased number of Escherichia coli/Shigella spp. on the colonic surface and within the crypts compared to control dogs. In conclusion, the bacterial microbiota in the colonic mucosa differed between dogs with and without CIE, with depletion of the crypt bacteria in dogs with CIE. The crypt bacterial species that was intimately associated with the host mucosa in control dogs was composed mainly of Helicobacter spp.

Derivation of adult canine intestinal organoids for translational research in gastroenterology.

BACKGROUND: Large animal models, such as the dog, are increasingly being used for studying diseases including gastrointestinal (GI) disorders. Dogs share similar environmental, genomic, anatomical, and intestinal physiologic features with humans. To bridge the gap between commonly used animal models, such as rodents, and humans, and expand the translational potential of the dog model, we developed a three-dimensional (3D) canine GI organoid (enteroid and colonoid) system. Organoids have recently gained interest in translational research as this model system better recapitulates the physiological and molecular features of the tissue environment in comparison with two-dimensional cultures. RESULTS: Organoids were derived from tissue of more than 40 healthy dogs and dogs with GI conditions, including inflammatory bowel disease (IBD) and intestinal carcinomas. Adult intestinal stem cells (ISC) were isolated from whole jejunal tissue as well as endoscopically obtained duodenal, ileal, and colonic biopsy samples using an optimized culture protocol. Intestinal organoids were comprehensively characterized using histology, immunohistochemistry, RNA in situ hybridization, and transmission electron microscopy, to determine the extent to which they recapitulated the in vivo tissue characteristics. Physiological relevance of the enteroid system was defined using functional assays such as optical metabolic imaging (OMI), the cystic fibrosis transmembrane conductance regulator (CFTR) function assay, and Exosome-Like Vesicles (EV) uptake assay, as a basis for wider applications of this technology in basic, preclinical and translational GI research. We have furthermore created a collection of cryopreserved organoids to facilitate future research. CONCLUSIONS: We establish the canine GI organoid systems as a model to study naturally occurring intestinal diseases in dogs and humans, and that can be used for toxicology studies, for analysis of host-pathogen interactions, and for other translational applications.

Correlating Gastrointestinal Histopathologic Changes to Clinical Disease Activity in Dogs With Idiopathic Inflammatory Bowel Disease.

Prior studies have failed to detect a convincing association between histologic lesions of inflammation and clinical activity in dogs with inflammatory bowel disease (IBD). We hypothesized that use of a simplified histopathologic scoring system would improve the consistency of interpretation among pathologists when describing histologic lesions of gastrointestinal inflammation. Our aim was to evaluate the correlation of histopathologic changes to clinical activity in dogs with IBD using this new system. Forty-two dogs with IBD and 19 healthy control dogs were enrolled in this retrospective study. Endoscopic biopsies from the stomach, duodenum, ileum, and colon were independently scored by 8 pathologists. Clinical disease activity was scored using the Canine Inflammatory Bowel Disease Activity Index (CIBDAI) or the Canine Chronic Enteropathy Clinical Activity Index (CCECAI), depending on the individual study center. Summative histopathological scores and clinical activity were calculated for each tissue (stomach, duodenum, ileum, and colon) and each tissue histologic score (inflammatory/morphologic feature). The correlation between CCECAI/CIBDAI and summative histopathologic score was significant ( P < .05) for duodenum ( r = 0.42) and colon ( r = 0.33). In evaluating the relationship between histopathologic scores and clinical activity, significant ( P < .05) correlations were observed for crypt dilation ( r = 0.42), lamina propria (LP) lymphocytes ( r = 0.40), LP neutrophils ( r = 0.45), mucosal fibrosis ( r = 0.47), lacteal dilation ( r = 0.39), and villus stunting ( r = 0.43). Compared to earlier grading schemes, the simplified scoring system shows improved utility in correlating histopathologic features (both summative histology scores and select histologic scores) to IBD clinical activity.

Multilaboratory Survey To Evaluate Salmonella Prevalence in Diarrheic and Nondiarrheic Dogs and Cats in the United States between 2012 and 2014.

Eleven laboratories collaborated to determine the periodic prevalence of Salmonella in a population of dogs and cats in the United States visiting veterinary clinics. Fecal samples (2,965) solicited from 11 geographically dispersed veterinary testing laboratories were collected in 36 states between January 2012 and April 2014 and tested using a harmonized method. The overall study prevalence of Salmonella in cats (3 of 542) was <1%. The prevalence in dogs (60 of 2,422) was 2.5%. Diarrhea was present in only 55% of positive dogs; however, 3.8% of the all diarrheic dogs were positive, compared with 1.8% of the nondiarrheic dogs. Salmonella-positive dogs were significantly more likely to have consumed raw food (P = 0.01), to have consumed probiotics (P = 0.002), or to have been given antibiotics (P = 0.01). Rural dogs were also more likely to be Salmonella positive than urban (P = 0.002) or suburban (P = 0.001) dogs. In the 67 isolates, 27 unique serovars were identified, with three dogs having two serovars present. Antimicrobial susceptibility testing of 66 isolates revealed that only four of the isolates were resistant to one or more antibiotics. Additional characterization of the 66 isolates was done using pulsed-field gel electrophoresis and whole-genome sequencing (WGS). Sequence data compared well to resistance phenotypic data and were submitted to the National Center for Biotechnology Information (NCBI). This study suggests an overall decline in prevalence of Salmonella-positive dogs and cats over the last decades and identifies consumption of raw food as a major risk factor for Salmonella infection. Of note is that almost half of the Salmonella-positive animals were clinically nondiarrheic.

Ultrastructural changes in chronic inflammatory enteropathies-a comparison between dogs and humans.

Chronic inflammatory enteropathies (CIEs) are an important group of diseases in dogs and involve complex pathogenetic aspects. Endoscopy and histopathology are vital for documenting the disease but are less useful for subclassifying CIEs and predicting the response to treatment. However, healing of the mucosal disease process (deep remission) and ultrastructural evaluation of the mucosa have received little attention in canine CIE. Given that canine CIE shares many similarities with inflammatory bowel diseases (IBDs) in human patients-and presents a good spontaneous disease model for human IBD-this perspective article evaluates the literature on ultrastructural lesions in canine CIE and human IBD and offers future directions for the study of ultrastructural mucosal lesions in canine CIE. Such lesions might have a higher sensitivity of detection than structural changes revealed upon light microscopy and may even precede or remain after the resolution of the clinical signs and histologic lesions.

Exploring the role of antioxidants in sepsis-associated oxidative stress: a comprehensive review.

Sepsis is a potentially fatal condition characterized by organ dysfunction caused by an imbalanced immune response to infection. Although an increased inflammatory response significantly contributes to the pathogenesis of sepsis, several molecular mechanisms underlying the progression of sepsis are associated with increased cellular reactive oxygen species (ROS) generation and exhausted antioxidant pathways. This review article provides a comprehensive overview of the involvement of ROS in the pathophysiology of sepsis and the potential application of antioxidants with antimicrobial properties as an adjunct to primary therapies (fluid and antibiotic therapies) against sepsis. This article delves into the advantages and disadvantages associated with the utilization of antioxidants in the therapeutic approach to sepsis, which has been explored in a variety of animal models and clinical trials. While the application of antioxidants has been suggested as a potential therapy to suppress the immune response in cases where an intensified inflammatory reaction occurs, the use of multiple antioxidant agents can be beneficial as they can act additively or synergistically on different pathways, thereby enhancing the antioxidant defense. Furthermore, the utilization of immunoadjuvant therapy, specifically in septic patients displaying immunosuppressive tendencies, represents a promising advancement in sepsis therapy.

Changes in symmetric dimethylarginine and glomerular filtration rates in hyperthyroid cats undergoing radioiodine therapy.

OBJECTIVES: The aims of this study were to evaluate concentrations of symmetric dimethylarginine (SDMA) in hyperthyroid cats before and after radioiodine treatment, and to compare results with other variables used to assess kidney function in cats (creatinine, urine specific gravity [USG] and glomerular filtration rate [GFR] measured by renal scintigraphy). METHODS: Thirteen cats diagnosed with hyperthyroidism based on clinical signs and increased serum total thyroxine (TT4) were included in this prospective study. Study design included physical examination, complete blood count, serum chemistry, TT4, urinalysis and SDMA before treatment (T0) and at 1 month (T1) and 3 months post-treatment (T3). GFR was quantified by renal scintigraphy at T0 and T3. RESULTS: Median GFR decreased significantly from baseline (3.18 ml/kg/min; range 1.35-4.87) at T3 (2.22 ml/kg/min; range 1.81-3.42 [P = 0.005]). While median creatinine and serum urea nitrogen increased post-treatment (creatinine: T0 = 0.8 mg/dl [range 0.4-1.1], T1 = 1.3 mg/dl [range 0.9-2]; T3 = 1.65 mg/dl [range 0.8-2.8]; P <0.001; serum urea nitrogen: T0 = 23 mg/dl [range 15-26]; T1 = 27 mg/dl [range 20-40]; T3 = 27.5 mg/dl [range 20-36]; P <0.001), SDMA and USG did not change significantly (SDMA: T0 = 11 µg/dl [range 7-15]; T1 = 12 µg/dl [range 6-16]; T3 = 10.5 µg/dl [range 8-21]; P = 0.789; USG: T0 = 1.030 [range 1.011-1.059]; T1 = 1.035 [range 1.012-1.044]; T3 = 1.030 [range 1.007-1.055]; P = 0.792). CONCLUSIONS AND RELEVANCE: Our data suggest that factors other than GFR may affect serum SDMA in hyperthyroid cats and that SDMA does not offer an advantage over other biomarkers traditionally used to predict changes in renal function following radioiodine therapy.

Study Transportation of Drugs within Newly Established Murine Colon Organoid Systems.

The development of 3D organoids of the small intestine is a tremendous breakthrough in drug development and biological research. However, the development of colonic organoids (i.e., colonoids) is particularly challenging due to a lack of simple, cost-effective protocols for colonoid cultivation. Here, intestinal homogenates are described as a supplement to the culture medium for maintaining and replicating colonic stem cells. Colonoids generated by this cultivation protocol demonstrate substantial proliferation and differentiation (3 months). There is a similarity between cultured colonoids and primary colon tissue regarding structure and functionality. To evaluate the functionality of colonoids, permeability testing is performed with suspensions of 4 and 40 kDa fluorescein isothiocyanate-dextran (FITC-DEX). It is observed that neither can permeate the healthy epithelial barrier. The P-glycoprotein receptor, a vital drug efflux pump mitigating potential drug toxicity, is functionally manipulated, as evidenced by its inhibition function by verapamil and monitoring uptake of Rhodamin 123. In addition, Forskolin treatment which affects chloride transport results in organoid swelling; this confirms the functional expression of the CFTR transporter in the colonoids. This protocol to generate colonoids is promising for high-throughput drug screening, toxicity testing, and oral drug development.

Oxidative stress, hormones, and effects of natural antioxidants on intestinal inflammation in inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a chronic, relapsing gastrointestinal (GI) disorder characterized by intestinal inflammation. The etiology of IBD is multifactorial and results from a complex interplay between mucosal immunity, environmental factors, and host genetics. Future therapeutics for GI disorders, including IBD, that are driven by oxidative stress require a greater understanding of the cellular and molecular mechanisms mediated by reactive oxygen species (ROS). In the GI tract, oxidative stressors include infections and pro-inflammatory responses, which boost ROS generation by promoting the production of pro-inflammatory cytokines. Nuclear factor kappa B (NF-κB) and nuclear factor erythroid 2-related factor 2 (Nrf2) represent two important signaling pathways in intestinal immune cells that regulate numerous physiological processes, including anti-inflammatory and antioxidant activities. Natural antioxidant compounds exhibit ROS scavenging and increase antioxidant defense capacity to inhibit pro-oxidative enzymes, which may be useful in IBD treatment. In this review, we discuss various polyphenolic substances (such as resveratrol, curcumin, quercetin, green tea flavonoids, caffeic acid phenethyl ester, luteolin, xanthohumol, genistein, alpinetin, proanthocyanidins, anthocyanins, silymarin), phenolic compounds including thymol, alkaloids such as berberine, storage polysaccharides such as tamarind xyloglucan, and other phytochemicals represented by isothiocyanate sulforaphane and food/spices (such as ginger, flaxseed oil), as well as antioxidant hormones like melatonin that target cellular signaling pathways to reduce intestinal inflammation occurring with IBD.

OrganoidChip facilitates hydrogel-free immobilization for fast and blur-free imaging of organoids.

Organoids are three-dimensional structures of self-assembled cell aggregates that mimic anatomical features of in vivo organs and can serve as in vitro miniaturized organ models for drug testing. The most efficient way of studying drug toxicity and efficacy requires high-resolution imaging of a large number of organoids acquired in the least amount of time. Currently missing are suitable platforms capable of fast-paced high-content imaging of organoids. To address this knowledge gap, we present the OrganoidChip, a microfluidic imaging platform that incorporates a unique design to immobilize organoids for endpoint, fast imaging. The chip contains six parallel trapping areas, each having a staging and immobilization chamber, that receives organoids transferred from their native culture plates and anchors them, respectively. We first demonstrate that the OrganoidChip can efficiently immobilize intestinal and cardiac organoids without compromising their viability and functionality. Next, we show the capability of our device in assessing the dose-dependent responses of organoids' viability and spontaneous contraction properties to Doxorubicin treatment and obtaining results that are similar to off-chip experiments. Importantly, the chip enables organoid imaging at speeds that are an order of magnitude faster than conventional imaging platforms and prevents the acquisition of blurry images caused by organoid drifting, swimming, and fast stage movements. Taken together, the OrganoidChip is a promising microfluidic platform that can serve as a building block for a multiwell plate format that can provide high-throughput and high-resolution imaging of organoids in the future.

Canine Intestinal Organoids as a Novel In Vitro Model of Intestinal Drug Permeability: A Proof-of-Concept Study.

A key component of efforts to identify the biological and drug-specific aspects contributing to therapeutic failure or unexpected exposure-associated toxicity is the study of drug-intestinal barrier interactions. While methods supporting such assessments are widely described for human therapeutics, relatively little information is available for similar evaluations in support of veterinary pharmaceuticals. There is, therefore, a critical need to develop novel approaches for evaluating drug-gut interactions in veterinary medicine. Three-dimensional (3D) organoids can address these difficulties in a reasonably affordable system that circumvents the need for more invasive in vivo assays in live animals. However, a first step in developing such systems is understanding organoid interactions in a 2D monolayer. Given the importance of orally administered medications for meeting the therapeutic need of companion animals, we demonstrate growth conditions under which canine-colonoid-derived intestinal epithelial cells survive, mature, and differentiate into confluent cell systems with high monolayer integrity. We further examine the applicability of this canine-colonoid-derived 2D model to assess the permeability of three structurally diverse, passively absorbed ß-blockers (e.g., propranolol, metoprolol, and atenolol). Both the absorptive and secretive apparent permeability (Papp) of these drugs at two different pH conditions were evaluated in canine-colonoid-derived monolayers and compared with that of Caco-2 cells. This proof-of-concept study provides promising preliminary results with regard to the utility of canine-derived organoid monolayers for species-specific assessments of therapeutic drug passive permeability.

Clinical utility of an immunoglobulin A-based serological panel for the diagnosis of chronic enteropathy in dogs.

BACKGROUND: A panel of IgA-based serologic assays might aid in the diagnosis of chronic enteropathy (CE) in dogs, a syndrome encompassing conditions such as food-responsive enteropathy, immunosuppressant-responsive enteropathy, and inflammatory bowel disease (also referred to as chronic inflammatory enteropathy). However, it is unclear whether these biomarkers discriminate between CE and other types of primary intestinal disorders. OBJECTIVES: To evaluate a diagnostic panel that measures serum concentrations of IgA directed against OmpC (ACA), canine calprotectin (ACNA), and gliadin-derived peptides (AGA) in dogs with well-characterized intestinal diseases. ANIMALS: Fifty-five dogs with primary intestinal disease. METHODS: Serum ACA, ACNA, and AGA concentrations were measured in 30 dogs with CE and 25 dogs with other intestinal diseases (non-CE population), including histoplasmosis, parasitism, E. coli-associated granulomatous colitis, and lymphoma. Serum IgA concentrations were compared among populations, and sensitivities and specificities were calculated using laboratory-provided cut-points. RESULTS: Twenty-six of 30 (87%) CE dogs and 21 of 25 (84%) non-CE dogs had abnormal concentrations (intermediate or high) of at least 2 markers; these proportions were not significantly different (P = .99). A serum ACA concentration ≥15 EU/mL was 86.7% (95% confidence interval [CI], 69.3%-96.2%) sensitive and 24.0% (95% CI, 9.4%-45.1%) specific for CE diagnosis. High AGA concentrations were observed in 16 of 25 (64%) non-CE dogs. CONCLUSIONS AND CLINICAL IMPORTANCE: The evaluated serologic markers were poorly specific for CE diagnosis, which raises concerns that their use in clinical practice might lead to misdiagnoses and delayed or even detrimental treatments in dogs with non-CE intestinal diseases.

Correlation between Targeted qPCR Assays and Untargeted DNA Shotgun Metagenomic Sequencing for Assessing the Fecal Microbiota in Dogs.

DNA shotgun sequencing is an untargeted approach for identifying changes in relative abundances, while qPCR allows reproducible quantification of specific bacteria. The canine dysbiosis index (DI) assesses the canine fecal microbiota by using a mathematical algorithm based on qPCR results. We evaluated the correlation between qPCR and shotgun sequencing using fecal samples from 296 dogs with different clinical phenotypes. While significant correlations were found between qPCR and sequencing, certain taxa were only detectable by qPCR and not by sequencing. Based on sequencing, less than 2% of bacterial species (17/1190) were consistently present in all healthy dogs (n = 76). Dogs with an abnormal DI had lower alpha-diversity compared to dogs with normal DI. Increases in the DI correctly predicted the gradual shifts in microbiota observed by sequencing: minor changes (R = 0.19, DI < 0 with any targeted taxa outside the reference interval, RI), mild-moderate changes (R = 0.24, 0 < DI < 2), and significant dysbiosis (R = 0.54, 0.73, and 0.91 for DI > 2, DI > 5, and DI > 8, respectively), compared to dogs with a normal DI (DI < 0, all targets within the RI), as higher R-values indicated larger dissimilarities. In conclusion, the qPCR-based DI is an effective indicator of overall microbiota shifts observed by shotgun sequencing in dogs.

Canine chronic enteropathy-Current state-of-the-art and emerging concepts.

Over the last decade, chronic inflammatory enteropathies (CIE) in dogs have received great attention in the basic and clinical research arena. The 2010 ACVIM Consensus Statement, including guidelines for the diagnostic criteria for canine and feline CIE, was an important milestone to a more standardized approach to patients suspected of a CIE diagnosis. Great strides have been made since understanding the pathogenesis and classification of CIE in dogs, and novel diagnostic and treatment options have evolved. New concepts in the microbiome-host-interaction, metabolic pathways, crosstalk within the mucosal immune system, and extension to the gut-brain axis have emerged. Novel diagnostics have been developed, the clinical utility of which remains to be critically evaluated in the next coming years. New directions are also expected to lead to a larger spectrum of treatment options tailored to the individual patient. This review offers insights into emerging concepts and future directions proposed for further CIE research in dogs for the next decade to come.

Vertical transmission of attaching and invasive E. coli from the dam to neonatal mice predisposes to more severe colitis following exposure to a colitic insult later in life.

The gastrointestinal microbiota begins to be acquired at birth and continually matures through early adolescence. Despite the relevance for gut health, few studies have evaluated the impact of pathobiont colonization of neonates on the severity of colitis later in life. LF82 is an adherent invasive E. coli strain associated with ileal Crohn's disease. The aim of this study was to evaluate the severity of dextran sodium sulfate (DSS)-induced colitis in mice following E. coli LF82 colonization. Gnotobiotic mice harboring the altered Schaedler flora (ASF) were used as the model. While E. coli LF82 is neither adherent nor invasive, it was been demonstrated that adult ASF mice colonized with E. coli LF82 develop more severe DSS-induced colitis compared to control ASF mice treated with DSS. Therefore, we hypothesized that E. coli LF82 colonization of neonatal ASF mice would reduce the severity of DSS-induced inflammation compared to adult ASF mice colonized with E. coli LF82. To test this hypothesis, adult ASF mice were colonized with E. coli LF82 and bred to produce offspring (LF82N) that were vertically colonized with LF82. LF82N and adult-colonized (LF82A) mice were given 2.0% DSS in drinking water for seven days to trigger colitis. More severe inflammatory lesions were observed in the LF82N + DSS mice when compared to LF82A + DSS mice, and were characterized as transmural in most of the LF82N + DSS mice. Colitis was accompanied by secretion of proinflammatory cytokines (IFNγ, IL-17) and specific mRNA transcripts within the colonic mucosa. Using 16S rRNA gene amplicon sequencing, LF82 colonization did not induce significant changes in the ASF community; however, minimal changes in spatial redistribution by fluorescent in situ hybridization were observed. These results suggest that the age at which mice were colonized with E. coli LF82 pathobiont differentially impacted severity of subsequent colitic events.

Differential Transcriptomic Profiles Following Stimulation with Lipopolysaccharide in Intestinal Organoids from Dogs with Inflammatory Bowel Disease and Intestinal Mast Cell Tumor.

Lipopolysaccharide (LPS) is associated with chronic intestinal inflammation and promotes intestinal cancer progression in the gut. While the interplay between LPS and intestinal immune cells has been well-characterized, little is known about LPS and the intestinal epithelium interactions. In this study, we explored the differential effects of LPS on proliferation and the transcriptome in 3D enteroids/colonoids obtained from dogs with naturally occurring gastrointestinal (GI) diseases including inflammatory bowel disease (IBD) and intestinal mast cell tumor. The study objective was to analyze the LPS-induced modulation of signaling pathways involving the intestinal epithelia and contributing to colorectal cancer development in the context of an inflammatory (IBD) or a tumor microenvironment. While LPS incubation resulted in a pro-cancer gene expression pattern and stimulated proliferation of IBD enteroids and colonoids, downregulation of several cancer-associated genes such as Gpatch4, SLC7A1, ATP13A2, and TEX45 was also observed in tumor enteroids. Genes participating in porphyrin metabolism (CP), nucleocytoplasmic transport (EEF1A1), arachidonic acid, and glutathione metabolism (GPX1) exhibited a similar pattern of altered expression between IBD enteroids and IBD colonoids following LPS stimulation. In contrast, genes involved in anion transport, transcription and translation, apoptotic processes, and regulation of adaptive immune responses showed the opposite expression patterns between IBD enteroids and colonoids following LPS treatment. In brief, the crosstalk between LPS/TLR4 signal transduction pathway and several metabolic pathways such as primary bile acid biosynthesis and secretion, peroxisome, renin-angiotensin system, glutathione metabolism, and arachidonic acid pathways may be important in driving chronic intestinal inflammation and intestinal carcinogenesis.

Synbiotic-IgY Therapy Modulates the Mucosal Microbiome and Inflammatory Indices in Dogs with Chronic Inflammatory Enteropathy: A Randomized, Double-Blind, Placebo-Controlled Study.

Chronic inflammatory enteropathy (CE) is a common cause of persistent gastrointestinal signs and intestinal inflammation in dogs. Since evidence links dysbiosis to mucosal inflammation, probiotics, prebiotics, or their combination (synbiotics) may reduce intestinal inflammation and ameliorate dysbiosis in affected dogs. This study's aim was to investigate the effects of the synbiotic-IgY supplement on clinical signs, inflammatory indices, and mucosal microbiota in dogs with CE. Dogs with CE were enrolled in a randomized prospective trial. Twenty-four client-owned dogs were fed a hydrolyzed diet and administered supplement or placebo (diet) for 6 weeks. Dogs were evaluated at diagnosis and 2- and 6-week post-treatment. Outcome measures included clinical activity, endoscopic and histologic scores, inflammatory markers (fecal calprotectin, C-reactive protein), and composition of the mucosal microbiota via FISH. Eleven supplement- and nine placebo-treated dogs completed the trial. After 6 weeks of therapy, clinical activity and endoscopic scores decreased in both groups. Compared to placebo-treated dogs, dogs administered supplement showed decreased calprotectin at 2-week post-treatment, decreased CRP at 2- and 6-week post-treatment increased mucosal Clostridia and Bacteroides and decreased Enterobacteriaceae in colonic biopsies at trial completion. Results suggest a beneficial effect of diet and supplements on host responses and mucosal microbiota in dogs with CE.

Helicobacter bilis colonization enhances susceptibility to Typhlocolitis following an inflammatory trigger.

BACKGROUND: Aberrant mucosal immune responses to antigens of the resident microbiota are a significant cause of inflammatory bowel diseases (IBD), as are genetic and environmental factors. Previous work from our laboratory demonstrated that Helicobacter bilis colonization of immunocompetent, defined microbiota mice induced antigen-specific immune responses to the resident microbiota, yet these mice failed to develop colitis, suggesting that the immunological provocation induced by H. bilis alone was insufficient to induce disease. AIM: The purpose of this study was to test the hypothesis that the introduction of a bacterial provocateur such as H. bilis enhances the host's susceptibility to IBD following an inflammatory event. METHODS: Defined microbiota (DM) mice colonized with H. bilis were administered low dose (1.5%) dextran sodium sulfate (DSS) in drinking water for 5 days followed by a 4-day restitution period. Severity of lesions was assessed grossly and microscopically. Differential expression of select mucosal genes and histopathologic lesions was characterized. RESULTS: Helicobacter bilis colonization increased the severity of intestinal inflammation induced by an inflammatory trigger in the form of low-dose DSS. An analysis of the molecular and cellular mechanisms associated with H. bilis colonization revealed significant increases in expression of mucosal genes associated with lymphocyte activation and inflammatory cell chemotaxis as well as increased infiltration of mucosal macrophages and T cells in mice colonized with H. bilis prior to DSS treatment versus DSS treatment alone. CONCLUSIONS: These results indicate that prior colonization with H. bilis heightens the host's sensitivity to enteric inflammation by altering mucosal homeostasis and initiating immune cell activation and migration.

Rules of Engagement: Epithelial-Microbe Interactions and Inflammatory Bowel Disease.

Inflammatory bowel diseases (IBD), including Crohn's disease (CD) and ulcerative colitis (UC), are complex, multifactorial disorders that lead to chronic and relapsing intestinal inflammation. The exact etiology remains unknown, however multiple factors including the environment, genetic, dietary, mucosal immunity, and altered microbiome structure and function play important roles in disease onset and progression. Supporting this notion that the gut microbiota plays a pivotal role in IBD pathogenesis, studies in gnotobiotic mice have shown that mouse models of intestinal inflammation require a microbial community to develop colitis. Additionally, antimicrobial therapy in some IBD patients will temporarily induce remission further demonstrating an association between gut microbes and intestinal inflammation. Finally, a dysfunctional intestinal epithelial barrier is also recognized as a key pathogenic factor in IBD. The intestinal epithelium serves as a barrier between the luminal environment and the mucosal immune system and guards against harmful molecules and microorganisms while being permeable to essential nutrients and solutes. Beneficial (i.e., mutualists) bacteria promote mucosal health by strengthening barrier integrity, increasing local defenses (mucin and IgA production) and inhibiting pro-inflammatory immune responses and apoptosis to promote mucosal homeostasis. In contrast, pathogenic bacteria and pathobionts suppress expression and localization of tight junction proteins, cause dysregulation of apoptosis/proliferation and increase pro-inflammatory signaling that directly damages the intestinal mucosa. This review article will focus on the role of intestinal epithelial cells (IECs) and the luminal environment acting as mediators of barrier function in IBD. We will also share some of our translational observations of interactions between IECs, immune cells, and environmental factors contributing to maintenance of mucosal homeostasis, as it relates to GI inflammation and IBD in different animal models.