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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

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.

Long-Term Recovery of the Fecal Microbiome and Metabolome of Dogs with Steroid-Responsive Enteropathy.

The long-term impact of treatment of dogs with steroid-responsive enteropathy (SRE) on the fecal microbiome and metabolome has not been investigated. Therefore, this study aimed to evaluate the fecal microbiome and metabolome of dogs with SRE before, during, and following treatment with standard immunosuppressive therapy and an elimination diet. We retrospectively selected samples from 9 dogs with SRE enrolled in a previous clinical trial, which received treatment for 8 weeks, and had achieved remission as indicated by the post-treatment clinical scores. Long-term (1 year) samples were obtained from a subset (5/9) of dogs. Samples from 13 healthy dogs were included as controls (HC). We evaluated the microbiome using 16S rRNA sequencing and qPCR. To evaluate the recovery of gut function, we measured fecal metabolites using an untargeted approach. While improvement was observed for some bacterial taxa after 8 weeks of treatment, several bacterial taxa remained significantly different from HC. Seventy-five metabolites were altered in dogs with SRE, including increased fecal amino acids and vitamins, suggesting malabsorption as a component of SRE. One year after treatment, however, all bacterial species were evaluated by qPCR and 16S rRNA gene sequencing, and all but thirteen metabolites were no longer different from healthy controls.

An initial genome-wide investigation of protein-losing enteropathy in Gordon setters: Exploratory observations.

The objective of this preliminary study was to identify genomic regions that may predispose Gordon setters from the United Kingdom to familial protein-losing enteropathy (PLE) at a young age. A total of 106 related Gordon setters was used, including 6 affected dogs from an affected litter, 6 case controls from the same litter, 10 related/affected dogs, and 84 related/unaffected dogs. Genomic DNA was collected from each Gordon setter and extracted from buccal mucosal swabs. Genotyping of affected and unaffected dogs was carried out using the Canine Illumina HD SNP array and data generated were analyzed with PLINK software, using fixation index (Fst) and runs of homozygosity (ROH) methods. Pairwise Fst analyses between the affected and unaffected Gordon setter dogs identified various regions of differentiation on chromosomes 10, 18, 21, and 23 that contained several important genes. These regions revealed 5 candidate genes, including RARB, TTC7A, SOCS5, PIGF, and RHOD, that are associated with human inflammatory bowel disease (IBD) and could potentially be associated with PLE in Gordon setters. Run of homozygosity (ROH) analyses revealed additional unique regions on chromosomes 15 and 17. These regions contained genes SYT1, UCN, and FNDC that could also be potential candidates for PLE in Gordon setters. The biological functions of the identified genes provided initial insights into the pathophysiology of PLE. Further large-scale studies are warranted to investigate the possible causality of these genomic regions and any possible genetic markers that could be used in predicting susceptibility to PLE syndrome.

L'objectif de cette étude préliminaire était d'identifier les régions génomiques susceptibles de prédisposer les chiens Gordon setter du Royaume-Uni à l'entéropathie familiale de perte de protéines (PLE) à un jeune âge. Un total de 106 Gordon setter apparentés a été utilisé, dont six chiens affectés d'une portée affectée, six cas témoins de la même portée, 10 chiens apparentés/affectés et 84 chiens apparentés/non affectés. L'ADN génomique a été obtenu à partir de chaque Gordon setter et extrait des écouvillons de la muqueuse buccale. Le génotypage des chiens affectés et non affectés a été effectué à l'aide de la matrice SNP Canine Illumina HD et les données générées ont été analysées avec le logiciel PLINK, en utilisant des méthodes d'indice de fixation (Fst) et d'homozygotie (ROH). Des analyses Fst par paires entre les chiens Gordon setter affectés et non affectés ont identifié diverses régions de différenciation sur les chromosomes 10, 18, 21 et 23 qui contenaient plusieurs gènes importants. Ces régions ont révélé cinq gènes candidats, dont RARB, TTC7A, SOCS5, PIGF et RHOD, qui sont associés à la maladie inflammatoire de l'intestin (IBD) humaine et pourraient potentiellement être associés à la PLE chez les Gordon setter. Les analyses d'homozygotie (ROH) ont révélé des régions uniques supplémentaires sur les chromosomes 15 et 17. Ces régions contenaient les gènes SYT1, UCN et FNDC qui pourraient également être des candidats potentiels pour la PLE chez les Gordon setter. Les fonctions biologiques des gènes identifiés ont fourni un aperçu initial de la physiopathologie de la PLE. D'autres études à grande échelle sont nécessaires pour étudier la causalité possible de ces régions génomiques et tous les marqueurs génétiques possibles qui pourraient être utilisés pour prédire la sensibilité au syndrome PLE.(Traduit par Docteur Serge Messier).

Colonic mucosal and cytobrush sample cytokine mRNA expression in canine inflammatory bowel disease and their correlation with disease activity, endoscopic and histopathologic score.

Canine inflammatory bowel disease (IBD) is a group of chronic gastrointestinal disorders, the pathogenesis of which remains elusive, but it possibly involves the interaction of the intestinal immune system with luminal microbiota and food-derived antigens. Mucosal cytokines profiles in canine IBD have been investigated mainly in small intestinal disease, while data on cytokine profiles in large intestinal IBD are limited. The objective of this study was to measure colonic mucosal and cytobrush sample messenger (m)RNA expression of interleukin (IL)-1ß, IL-2, IL-12p40, IL-23p19, tumor necrosis factor-alpha (TNF-α) and chemokine C-C motif ligand (CCL28) in dogs with IBD and healthy controls using quantitative real-time polymerase chain reaction (PCR), and assess their correlation with clinical disease activity, endoscopic and histopathologic score. Dogs with IBD had a significantly increased mRNA expression of IL-1ß, IL-23p19 and CCL28 in the colonic mucosa, compared to healthy controls. None of the selected cytokines had significantly different mRNA expression in the colonic cytobrush samples between the two groups or between the colonic mucosa and cytobrush samples of dogs with IBD. Finally, there was a statistically significant correlation of clinical disease activity with endoscopic activity score and fibrosis and atrophy of the colonic mucosa in dogs with large intestinal IBD. IL-1ß, IL-23p19 and CCL28 could play a role in the pathogenesis of canine large intestinal IBD. Colonic cytokine expression does not correlate with clinical disease activity and/or endoscopic score. However, clinical signs reflect the severity of endoscopic lesions.

Harnessing the Biology of Canine Intestinal Organoids to Heighten Understanding of Inflammatory Bowel Disease Pathogenesis and Accelerate Drug Discovery: A One Health Approach.

In a recent issue of the Lancet, the prevalence of Inflammatory Bowel Disease (IBD) was estimated at 7 million worldwide. Overall, the burden of IBD is rising globally, with direct and indirect healthcare costs ranging between $14.6 and $31.6 billion in the U.S. alone in 2014. There is currently no cure for IBD, and up to 40% of patients do not respond to medical therapy. Although the exact determinants of the disease pathophysiology remain unknown, the prevailing hypothesis involves complex interplay among host genetics, the intestinal microenvironment (primarily bacteria and dietary constituents), and the mucosal immune system. Importantly, multiple chronic diseases leading to high morbidity and mortality in modern western societies, including type II diabetes, IBD and colorectal cancer, have epidemiologically been linked to the consumption of high-calorie, low-fiber, high monosaccharide, and high-fat diets (HFD). More specifically, data from our laboratory and others have shown that repeated consumption of HFD triggers dysbiotic changes of the gut microbiome concomitant with a state of chronic intestinal inflammation and increased intestinal permeability. However, progress in our understanding of the effect of dietary interventions on IBD pathogenesis has been hampered by a lack of relevant animal models. Additionally, current in vitro cell culture systems are unable to emulate the in vivo interplay between the gut microbiome and the intestinal epithelium in a realistic and translatable way. There remains, therefore, a critical need to develop translatable in vitro and in vivo models that faithfully recapitulate human gut-specific physiological functions to facilitate detailed mechanistic studies on the impact of dietary interventions on gut homeostasis. While the study of murine models has been pivotal in advancing genetic and cellular discoveries, these animal systems often lack key clinical signs and temporal pathological changes representative of IBD. Specifically, some limitations of the mouse model are associated with the use of genetic knockouts to induce immune deficiency and disease. This is vastly different from the natural course of IBD developing in immunologically competent hosts, as is the case in humans and dogs. Noteworthily, abundant literature suggests that canine and human IBD share common clinical and molecular features, such that preclinical studies in dogs with naturally occurring IBD present an opportunity to further our understanding on disease pathogenesis and streamline the development of new therapeutic strategies. Using a stepwise approach, in vitro mechanistic studies investigating the contribution of dietary interventions to chronic intestinal inflammation and "gut leakiness" could be performed in intestinal organoids and organoid derived monolayers. The biologic potential of organoids stems from the method's ability to harness hard-wired cellular programming such that the complexity of the disease background can be reflected more accurately. Likewise, the effect of therapeutic drug candidates could be evaluated in organoids prior to longitudinal studies in dog and human patients with IBD. In this review, we will discuss the value (and limitations) of intestinal organoids derived from a spontaneous animal disease model of IBD (i.e., the dog), and how it can heighten understanding of the interplay between dietary interventions, the gut microbiota and intestinal inflammation. We will also review how intestinal organoids could be used to streamline the preclinical development of therapeutic drug candidates for IBD patients and their best four-legged friends.

Effects of the Probiotic Mixture Slab51® (SivoMixx®) as Food Supplement in Healthy Dogs: Evaluation of Fecal Microbiota, Clinical Parameters and Immune Function.

The gut microbiota plays a crucial role in several physiologic functions of the host. In humans and animals, manipulation of the intestinal microbiota by oral administration of probiotic lactic acid bacteria plays a significant role in modulating the immune system. The aim of this study was to evaluate the safety of the probiotic mixture Slab51® and the capacity of this mixture to stimulate immune function in healthy dogs. Twenty dogs were divided in two groups and received a control diet or the same diet supplemented with a dose of 400 billion cfu of lyophilized bacteria for a period of 60 days. Body weight, food intake, body condition score (BCS), fecal score (FSS), fecal immunoglobulin IgA concentration, plasma IgG concentration, and fecal microbiota composition were monitored. Weight, food intake, BCS, FSS, and biochemical parameters remained unchanged during the treatment in both groups of animals. The fecal microbiota showed a significant decrease in the abundance of Clostridium perfringens and a significant increase in the abundance of beneficial Bifidobacterium and Lactobacillus organisms (p < 0.05). Fecal IgA and plasma IgG levels were significantly higher in the group receiving the probiotic compared to healthy controls. These data show that dietary supplementation with the probiotic mixture Slab51® is safe and well-tolerated, modulating the composition of the intestinal microbiota, and enhancing specific immune functions in healthy dogs.