Proximal Gut Research Articles

Yeast Expressing a Phage Endolysin Reduces Endogenous Clostridium perfringens Ex Vivo in 21-Day-Old Broiler Chicken Intestinal Fluids.

The phage endolysin PlyCP41 when purified from Escherichia coli exhibits lytic activity against Clostridium perfringens (CP) in vitro. The anti-clostridial activity of PlyCP41 endolysin expressed in transgenic yeast (Saccharomyces cerevisiae) was verified in phosphate buffered saline via mixing experiments with cultured CP and transgenic yeast slurries followed by serial dilution plating and colony counts on tryptose sulfite cycloserine (CP indicator) plates. The transgenic yeast containing PlyCP41 resulted in a log10 4.5 reduction (99.997%; P < 0.01) of the cultured CP. In addition, this serial dilution plating assay was used to demonstrate that transgenic yeast slurries could reduce the endogenous CP content in fluids from three different gastrointestinal regions (proximal, medial, and distal) from 21-day-old broiler chickens. The transgenic yeast treatment of gut slurries resulted in a log 10 1.19, 4.53, and 1.28 reduction in proximal, medial, and distal gut slurries (90% to 99.99% of the endogenous CP; P < 0.01), respectively, compared to nontreatment controls. These results indicate that the phage endolysin PlyCP41 expressed in S. cerevisiae is effective at reducing the endogenous CP in gastrointestinal fluids of broiler chickens. Future studies will measure the anti-CP effect in vivo by administering transgenic yeast to broiler chickens in the feed.

Divergent selection for feed efficiency in pigs altered the duodenum transcriptomic response to feed intake and its DNA methylation profiles.

Feed efficiency is a trait of interest in pigs as it contributes to lowering the ecological and economical costs of pig production. A divergent genetic selection experiment from a Large White pig population was performed for 10 generations, leading to pig lines with relatively low- (LRFI) and high- (HRFI) residual feed intake (RFI). Feeding behavior and metabolic differences have been previously reported between the two lines. We hypothesized that part of these differences could be related to differential sensing and absorption of nutrients in the proximal intestine. We investigated the duodenum transcriptome and DNA methylation profiles comparing overnight fasting with ad libitum feeding in LRFI and HRFI pigs (n = 24). We identified 1,106 differentially expressed genes between the two lines, notably affecting pathways of the transmembrane transport activity and related to mitosis or chromosome separation. The LRFI line showed a greater transcriptomic response to feed intake than the HRFI line. Feed intake affected genes from both anabolic and catabolic pathways in the pig duodenum, such as rRNA production and autophagy. Several nutrient transporter and tight junction genes were differentially expressed between lines and/or by short-term feed intake. We also identified 409 differentially methylated regions in the duodenum mucosa between the two lines, while this epigenetic mark was less affected by feeding. Our findings highlighted that the genetic selection for feed efficiency in pigs changed the transcriptome profiles of the duodenum, and notably its response to feed intake, suggesting key roles for this proximal gut segment in mechanisms underlying feed efficiency.NEW & NOTEWORTHY The duodenum is a key organ for the hunger/satiety loop and nutrient sensing. We investigated how the duodenum transcriptome and DNA methylation profiles are affected by feed intakes in pigs. We observed thousands of changes in gene expression levels between overnight-fasted and fed pigs in high-feed efficiency pig lines, but almost none in the related low-feed efficiency pig line.

Effects of Resistant Starch Infusion, Solely and Mixed with Xylan or Cellulose, on Gut Microbiota Composition in Ileum-Cannulated Pigs.

Fermentation of dietary fiber (DF) is beneficial for gut health, but its prebiotic effects are often impeded in the distal large intestine because of the fast degradation of fermentable substrates. One way to enhance the prebiotic effect of DF is to deliver fibers to the lower parts of the gut, which can be achieved by mixing different kinds of fiber. Therefore, in the present study, an ileum-cannulated pig model was employed to investigate the fermentation influence in the large intestine by infusing resistant starch solely (RS, fast fermentable fiber) and mixing with other fibers (xylan or cellulose). Twenty-four ileum-cannulated growing pigs were divided into four groups: one control group receiving saline ileal infusions and three experimental groups infused with RS, RS with xylan, or RS with cellulose. Fecal and plasma samples were analyzed for gut microbiota composition, short-chain fatty acids (SCFAs), and blood biochemistry. Results indicated no significant differences between the RS and control group for the microbiome and SCFA concentration (p > 0.05). However, RS combined with fibers, particularly xylan, resulted in enhanced and prolonged fermentation, marked by an increase in Blautia and higher lactate and acetate production (p < 0.05). In contrast, RS with cellulose infusion enriched bacterial diversity in feces (p < 0.05). Blood biochemistry parameters showed no significant differences across groups (p > 0.05), though a trend of increased glucose levels was noted in the treatment groups (p < 0.1). Overall, RS alone had a limited impact on the distal hindgut microbiota due to rapid fermentation in the proximal gut, whereas combining RS with other fibers notably improved gut microecology by extending the fermentation process.

DOP37 Noninvasive assessment of gut function using transcriptional recording sentinel cells

Abstract Background Bacteria within the human gut continuously adapt their gene expression to environmental conditions that are associated with diet, health, and disease. Noninvasive measurements of bacterial gene expression patterns throughout the intestine are important to understand in vivo microbiota physiology and pathophysiology. Current methods do not offer sufficient information about transient or proximal events within the intestine without using indirect or invasive approaches that disturb normal physiology and are inapplicable to clinical practice. Methods Here we used transcriptional recording sentinel cells that through a reverse transcriptase-Cas1–Cas2 complex record their own short-lived mRNA expression into long-lived DNA-based CRISPR arrays to report on gut function. We colonized gnotobiotic mice with E. coli sentinel cells and performed Record-seq on fecal samples to reconstruct their cellular histories as they passed along the gastrointestinal tract. Results Upon colonization of the mouse intestine, sentinel cells reported on diet, inflammation, and microbial interactions. Through transcriptome-scale information, Record-seq elucidated E. coli’s adaptations to intraluminal conditions including pH, oxygen levels, and ion availability. Unlike RNA-seq, Record-seq performed on stool samples retained information from proximal gut sections to noninvasively report on the luminal conditions in vivo. In a mouse model of inflammatory bowel disease, Record-seq not only diagnosed the inflammation in a severity-dependent manner but also revealed pathological gut conditions such as mucus depletion and hyperoxygenation. Conclusion Transcriptional recording sentinel cells noninvasively report on gut function and reveal environmental cues such as diet, inflammation, and microbial interactions, even in proximal gut sections that are otherwise unamenable to unbiased diagnostic interrogation.

Live Yeast (Saccharomyces cerevisiae var. boulardii) Supplementation in a European Sea Bass (Dicentrarchus labrax) Diet: Effects on the Growth and Immune Response Parameters.

The present study has been aimed at evaluating the effects of the dietary inclusion of the live yeasts, Saccharomyces cerevisiae var. boulardii (LSB) administered at increasing concentrations (0, 100, and 300 mg kg-1 of feed, here referred to as LSB 0, 100, 300) for 90 days, on the health conditions of European sea bass. The main zootechnical parameters, histological and morphological analyses, innate immunity response parameters (intestinal cytokine expression, lysozyme content, spontaneous hemolytic and hemagglutinating activities, antibacterial activities, and peroxidase activity) were measured as fish welfare parameters. LSB did not impair either growth parameters or the morphometric indexes. LSB down-regulated interleukin-1β transcription in the distal gut of fish treated with 5.4 × 105 CFU g-1 (LSB100) for 21 days. The interleukin-6 mRNA level decreased significantly in the proximal gut for both doses of yeast, after 21 days of feeding; the gene expression of interleukin-6 was significantly lower in the sea bass fed 10.81 × 105 CFU g-1 (LSB300) probiotic. The levels of TNF-α mRNA were not influenced by probiotic supplementation. Increases, although not significant, in the hematological and immunological parameters were also recorded. The data collected in the present study suggests that an LSB-supplemented diet acts on the gut immune system of sea bass by modulating the expression of the key inflammatory genes.

Characterization of free fatty acid receptor family in rainbow trout (Oncorhynchus mykiss): towards a better understanding of their involvement in fatty acid signalisation

Since 20 years of research, free fatty acids receptors (FFARs) have received considerable attention in mammals. To date, four FFARs (FFAR1, FFAR2, FFAR3 and FFAR4) are especially studied owing to their physiological importance in various biological processes. This ubiquitist group of G protein-coupled receptors (GPCRs) are majors reports in the key physiological functions such as the regulation of energy balance, metabolism or fatty acid sensing. However, up till date, even some studies were interested in their potential involvement in fatty acid metabolism, no genome investigation of these FFARs have been carried out in teleost fish. Through genome mining and phylogenetic analysis, we identified and characterised 7 coding sequences for ffar2 in rainbow trout whereas no ffar3 nor ffar4 gene have been found. This larger repertoire of ffar2 genes in rainbow trout results from successive additional whole-genome duplications which occurred in early teleosts and salmonids, respectively. A syntenic analysis was used to assign a new nomenclature to the salmonid ffar2 and showed a clear conservation of genomic organisation, further supporting the identity of these genes as ffar2. RT-qPCR was then used to examine, firstly during ontogenesis and secondly on feeding response the expression pattern of ffar1 and ffar2 genes in proximal gut and brain of all trout ffar genes. Overall, this study presents a comprehensive overview of the ffar family in rainbow trout.

A novel drug-like water-soluble small molecule Focal Adhesion Kinase (FAK) activator promotes intestinal mucosal healing

Non-steroidal anti-inflammatory drugs (NSAIDs) injure the proximal and distal gut by different mechanisms. While many drugs reduce gastrointestinal injury, no drug directly stimulates mucosal wound healing. Focal adhesion kinase (FAK), a non-receptor tyrosine kinase, induces epithelial sheet migration.We synthesized and evaluated a water-soluble FAK-activating small molecule, M64HCl, with drug-like properties. Monolayer wound closure and Western blots measured migration and FAK phosphorylation in Caco-2 ​cells, in vitro kinase assays established FAK activation, and pharmacologic tests assessed drug-like properties. 30 ​mg/kg/day M64HCl was administered in two murine small intestine injury models for 4 days.M64HCl (0.1–1000 ​nM) dose-dependently increased Caco-2 FAK-Tyr 397 phosphorylation, without activating Pyk2 and accelerated Caco-2 monolayer wound closure. M64HCl dose-responsively activates the FAK kinase domain vs. the non-salt M64, increasing the Vmax of ATP-binding. Pharmacologic tests suggested M64HCl has drug-like properties and is enterally absorbed. M64HCl 25 ​mg/kg/day continuous infusion promoted healing of ischemic jejunal ulcers and indomethacin-induced small intestinal injury in C57Bl/6 mice. M64HCl-treated mice exhibited smaller ulcers 4 days after ischemic ulcer induction or indomethacin injury. Renal histology and plasma creatinine were normal. Mild hepatic inflammatory changes and ALT elevation were similar among M64HCl-treated mice and controls. M64HCl was concentrated in kidney and gastrointestinal mucosa and functional nephrectomy studies suggested predominantly urinary excretion. Little toxicity was observed in vitro or in single-dose mouse toxicity studies until >1000x higher than effective concentrations. M64HCl, a water-soluble FAK activator, promotes epithelial restitution and intestinal mucosal healing and may be useful to treat gut mucosal injury.

Precursor-derived in-water peracetic acid impacts on broiler performance, gut microbiota, and antimicrobial resistance genes

Past antimicrobial misuse has led to the spread of antimicrobial resistance amongst pathogens, reportedly a major public health threat. Attempts to reduce the spread of antimicrobial resistant (AMR) bacteria are in place worldwide, among which finding alternatives to antimicrobials have a pivotal role. Such molecules could be used as "green alternatives" to reduce the bacterial load either by targeting specific bacterial groups or more generically, functioning as biocides when delivered in vivo. In this study, the effect of in-water peracetic acid as a broad-spectrum antibiotic alternative for broilers was assessed via hydrolysis of precursors sodium percarbonate and tetraacetylethylenediamine. Six equidistant peracetic acid levels were tested from 0 to 50 ppm using four pens per treatment and 4 birds per pen (i.e., 16 birds per treatment and 96 in total). Peracetic acid was administered daily from d 7 to 14 of age whilst measuring performance parameters and end-point bacterial concentration (qPCR) in crop, jejunum, and ceca, as well as crop 16S sequencing. PAA treatment, especially at 20, 30, and 40 ppm, increased body weight at d 14, and feed intake during PAA exposure compared to control (P < 0.05). PAA decreased bacterial concentration in the crop only (P < 0.05), which was correlated to better performance (P < 0.05). Although no differences in alpha- and beta-diversity were found, it was observed a reduction of Lactobacillus (P < 0.05) and Flectobacillus (P < 0.05) in most treatments compared to control, together with an increased abundance of predicted 4-aminobutanoate degradation (V) pathway. The analysis of the AMR genes did not point towards any systematic differences in gene abundance due to treatment administration. This, together with the rest of our observations could indicate that proximal gut microbiota modulation could result in performance amelioration. Thus, peracetic acid may be a valid antimicrobial alternative that could also positively affect performance.

Oral alloantigen exposure promotes donor-specific tolerance in a mouse model of minor-mismatched skin transplantation

Oral antigen exposure is a powerful, non-invasive route to induce immune tolerance to dietary antigens, and has been modestly successful at prolonging graft survival in rodent models of transplantation. To harness the mechanisms of oral tolerance for promoting long-term graft acceptance, we developed a mouse model where the antigen ovalbumin (OVA) was introduced orally prior to transplantation with skin grafts expressing OVA. Oral OVA treatment pre-transplantation promoted permanent graft acceptance and linked tolerance to skin grafts expressing OVA fused to the additional antigen 2W. Tolerance was donor-specific, as secondary donor-matched, but not third-party allografts were spontaneously accepted. Oral OVA treatment promoted an anergic phenotype in OVA-reactive CD4+ and CD8+ conventional T cells (Tconvs) and expanded OVA-reactive Tregs pre-transplantation. However, skin graft acceptance following oral OVA resisted partial depletion of Tregs and blockade of PD-L1. Mechanistically, we revealed a role for the proximal gut draining lymph nodes (gdLNs) in mediating this effect, as an intestinal infection that drains to the proximal gdLNs prevented tolerance induction. Our study extends previous work applying oral antigen exposure to transplantation and serves as proof of concept that the systemic immune mechanisms supporting oral tolerance are sufficient to promote long-term graft acceptance.

Intestinal permeability in patients with IgA nephropathy and other glomerular diseases: an observational study

BackgroundA dysregulated ‘gut-kidney axis’ may contribute to immunoglobulin A nephropathy (IgAN). We studied whether IgAN patients have disturbed intestinal permeability.MethodsIn a prospective, cross sectional, pilot study we assessed intestinal permeability in 35 IgAN patients, 18 patients with non-IgAN glomerulonephritides (GNs) and 19 healthy controls. After an overnight fast, trial participants ingested a multi-sugar solution and samples were obtained from 0 to 2, 2 to 5- and 5 to 24-h urine portions. Urinary sugar concentrations were quantified using isocratic ion-exchange high performance liquid chromatography. Indices of small intestinal permeability (0–2-h lactulose/L-rhamnose (L/R) ratio), distal small intestinal and proximal colonic permeability (2–5-h sucralose/erythritol (S/E) ratio) and colonic permeability (5–24-h sucralose/erythritol (S/E) ratio) were evaluated. Associations between groups and indices of intestinal permeability were investigated by a linear mixed model.ResultsSmall intestinal permeability (0–2 h L/R-ratio) was significantly increased in patients with glomerular diseases versus healthy controls. More precisely, increased small intestinal permeability was exclusively noted in non-IgAN GN patients, whereas IgAN patients exhibited a trend towards elevated small intestinal permeability. In total, 54% of patients with IgAN and 67% of non-IgAN GN patients had increased small intestinal permeability. Neither distal small intestinal and proximal colonic permeability nor colonic gut permeability indices (i.e., 2–5 h and 5–24 h S/E ratios) were significantly different between controls and any of the GN patient groups.ConclusionThe present single center pilot study suggests that disturbed intestinal permeability is common in patients with glomerular diseases and is not specific for IgAN.Trial registration numberGerman Clinical Trials Register DRKS00021533, Date: 24.04.2020.Graphical abstract

Fecal immunoglobulin A (IgA) and its subclasses in systemic lupus erythematosus patients are nuclear antigen reactive and this feature correlates with gut permeability marker levels

Systemic lupus erythematosus (SLE) is characterized by the production of anti-nuclear autoantibodies. Here, for the first time, we show that the abundances of gut permeability marker Zonulin and IgA1- and IgA2- subclasses are significantly higher in the fecal samples of SLE patients compared to HCs. Importantly, IgA-total, and IgA1- and IgA2-subclasses from SLE patients showed higher nAg reactivity titers. Notably, we found that not only the nuclear antigen (nAg) reactive fecal IgA1:IgA2 ratio is higher in SLE patients, but also the abundance and nAg reactivity of fecal IgA and subclasses, IgA1 particularly, correlate with the fecal levels of Zonulin, which is produced primarily in the small intestine. These observations that higher amounts of nAg-reactive IgA and gut permeability marker are produced, particularly, in the proximal gut suggest a compromised epithelial barrier function and pro-inflammatory characteristics of small intestine in SLE patients.

Mortality and cancer in eosinophilic gastrointestinal disorders distal to the esophagus: nationwide cohort study 1990–2017

BackgroundEosinophilic gastrointestinal disorders (EGIDs) include inflammatory conditions with enteric infiltration of eosinophils and resulting symptoms. This study aims to examine a population-based sample of patients for prevalence, mortality, and cancer risk in EGIDs distal to the esophagus.MethodsNationwide, population-based cohort study. EGID was identified through relevant biopsy codes from Sweden’s all 28 pathology departments through the ESPRESSO cohort. Individuals with EGID were then matched to general population reference individuals with similar age and sex. Study participants were linked to Swedish healthcare registers. Through Cox regression, we calculated adjusted hazard ratios (aHRs) adjusting for sex, age, county, calendar period, and education.ResultsIn total, 2429 patients (56% female) were found to have EGID distal to the esophagus, representing a prevalence of about 1/4800 in the Swedish population. Mean age was 44 years with 11% children at the time of diagnosis. Mortality was increased 17% in patients with EGIDs compared to reference individuals (aHR = 1.17; 95%CI = 1.04–1.33). Excess mortality was seen in gastric and small bowel eosinophilic disease, but not colonic disease (aHR = 1.81; 95%CI = 1.32–2.48, aHR = 1.50; 95%CI = 1.18–1.89, and aHR = 0.99; 95%CI = 0.85–1.16, respectively). Cause specific mortality was driven by cancer-related death (aHR = 1.33; 95%CI = 1.05–1.69). However, this study failed to show an increase in incident cancers (aHR = 1.14; 95%CI = 0.96–1.35). Comparison of EGID individuals with their siblings yielded similar aHRs.ConclusionsThis study found an increased risk of death in patients with EGIDs distal to the esophagus, with cancer death driving the increase. Proximal gut disease seems to confer the greatest risk. There was no increase in incident cancers.

Complications of Colostomy Reversal in Patients with Imperforate ANUS

Background: Colostomy reversal after definitive procedure in patients of Imperforate Anus (for example; PSARP) can be difficult and challenging for surgeon due to proximal and distal gut lumen discrepancy, short length of distal loop, difficult mobilization of stoma. These factors affect the anastomosis and anastomosis is done under tension due to these factors. Colostomies which are reversed under tension mostly results in anastomosis leakage and increases the morbidity and mortality rate. The objective of this study to assess postoperative outcome in terms of complications following colostomy closure in patients with imperforate anus. Material and Methods: This is a prospective study done at the Department of Pediatric Surgery, National Institute of Child Health Karachi during the period of 6months from December 2016 to June 2017. A Self- structured performa was used to collect the data of total 84 Patients who Underwent Colostomy Reversal after definitive procedure for imperforate anus . Data analyzed through SPSS version 20. A descriptive statistical analysis of continous and categorical variables were performed. Continuous data (like age) expressed as mean SD. The categorical data like gender, wound infections , anastomosis leakage expressed in frequency and percentages. Results: Out of total 84 children, majority 51 (60.7%) children were presented with ≤4 years of age (Mean age 4.15±3.12 years). Majority 55 (65.5%) children were presented with &gt;8 kg of weight (Mean weight 13.31±6.00 kg). There were 41 (48.8%) males and 43 (51.2%) females. Intra-operative difficulty was found in 55 (65.5%) children. Postoperative complication showed that wound infections were observed in 10 (11.9%) children while frequency of anastomotic leakage was found in 12 (14.3%) children. Conclusion: Anastomotic leakage was found to be higher following colostomy closure in children with intra-operative difficulty due to proximal and distal gut lumen discrepancy, short length of distal loop and dense adhesions making difficult the mobilization of stoma. Keywords: Postoperative outcome, Complications, Colostomy closure, Imperforate anus

Multiple Roles of Ret Signalling During Enteric Neurogenesis.

The majority of the enteric nervous system is formed by vagal neural crest cells which enter the foregut and migrate rostrocaudally to colonise the entire length of the gastrointestinal tract. Absence of enteric ganglia from the distal colon are the hallmark of Hirschsprung disease, a congenital disorder characterised by severe intestinal dysmotility. Mutations in the receptor tyrosine kinase RET have been identified in approximately 50% of familial cases of Hirschsprung disease but the cellular processes misregulated in this condition remain unclear. By lineage tracing neural crest cells in mice homozygous for a knock-in allele of Ret (Ret51/51), we demonstrate that normal activity of this receptor is required in vivo for the migration of enteric nervous system progenitors throughout the gut. In mutant mice, progenitors of enteric neurons fail to colonise the distal colon, indicating that failure of colonisation of the distal intestine is a major contributing factor for the pathogenesis of Hirschsprung disease. Enteric nervous system progenitors in the ganglionic proximal guts of mutant mice are also characterised by reduced proliferation and differentiation. These findings suggest that the functional abnormalities in Hirschsprung disease result from a combination of colonic aganglionosis and deficits in neuronal circuitry of more proximal gut segments. The reduced neurogenesis in the gut of Ret51/51 mutants was reproduced in the multilineage enteric nervous system progenitors isolated from these animals. Correction of the molecular defects of such progenitors fully restored their neurogenic potential in culture. These observations enhance our understanding of the pathogenesis of Hirschsprung disease and highlight potential approaches for its treatment.

Taurocholic acid, a primary 12α-hydroxylated bile acid, induces leakiness in the distal small intestine in rats

A high-fat diet increases 12α-hydroxylated (12αOH) bile acid (BA) secretion in rats, and secondary BAs are responsible for the leaky gut. This study aimed to examine the role of primary 12αOH BAs in gut barrier impairment in rats using dietary cholic acid (CA) supplementation (0.5 g/kg diet). The CA diet increased the 12αOH BAs concentrations in the small and large intestine, accompanied by gut barrier impairment. Based on the luminal 12αOH BAs concentrations, ex vivo gut leakiness was determined. Deoxycholic acid increased permeability in the large intestine, whereas taurocholic acid (TCA) increased the ileal permeability, but not jejunal permeability. A Rho kinase inhibitor attenuated TCA-induced ileal permeability. Administration of vancomycin, which abolishes secondary BAs, did not influence the gut leakiness induced by the CA diet. Changes in the gut permeation marker in the tail vein blood suggested the possibility that the CA-induced leakiness occurred in the small intestine. The CA diet enhanced the phosphorylation of myosin light chain 2 and reduced claudins expressions in rat ileal epithelia. Reductions in barrier function-related genes were observed in the ileum, but not in the colon of the CA-fed rats. Overall, the present study demonstrated the significance of TCA in proximal gut leakiness.

Oral alloantigen exposure promotes donor-specific tolerance in a mouse model of minor mismatched skin transplantation

Abstract Oral tolerance is a powerful, non-invasive route to induce immune tolerance, yet it has been unsuccessful to promote graft acceptance in rodent models of transplantation. To this end, we developed a mouse model where the model antigen ovalbumin (OVA) was introduced orally, in a regimen designed to improve bioavailability, duodenal lymph node drainage and prolong duration of antigen exposure, prior to transplantation with skin grafts constitutively expressing membrane-bound OVA. We found that oral OVA treatment pre-transplantation promoted permanent graft acceptance as well as linked tolerance to skin grafts expressing OVA coupled to the additional model antigen 2W. Tolerance was donor-specific, as secondary donor-matched, but not third-party grafts were spontaneously accepted in the absence of additional immunosuppression. Treatment with this oral OVA regimen promoted an anergic phenotype in OVA-reactive CD8+ T cells, generation of iTregs, and dysfunction of OVA-reactive CD4+ T cells pre-transplantation. The tolerance induced by oral OVA was robust, as anti-CD25/anti-PD-L1 at the time of transplantation failed to prevent skin graft acceptance. Mechanistically, we revealed a critical role for the proximal gut draining lymph nodes (gLNs) in mediating this effect, as an intestinal infection that drains to the proximal gLNs prevented the induction of transplantation tolerance. Our study extends previous work applying oral antigen exposure to transplantation and serves as proof of concept that the systemic immune mechanisms supporting oral tolerance are sufficient to promote long-term graft acceptance. Regimens that improve bioavailability and duration of oral alloantigen administration may hold promise in the clinic. This work was supported by a NIAID grant to ASC and MLA (P01AI-97113). Peter Wang was supported by undergraduate research grants from the Katen Scholars Program and the Dean's Fund for Undergraduate Research.

Metabolic growth, plankton selectivity, haemato-biochemical and intestinal morphometry of Nile tilapia (Oreochromis niloticus) fed a lysine-deficient diet in earthen ponds

Sustainable aquaculture requires the efficient use of natural pond resources as well as supplements to meet the dietary requirements of fish species. However, aquaculture diets are often produced without considering the lysine content of the natural production in the pond. As such, two plant-based diets, a control diet (L-Con) supplemented with 5.00 g lysine kg−1 diet and a lysine deficient diet (L-Def) without supplementation were fed to Nile tilapia raised in ponds with five replicates ponds for each diet. The aim was to investigate the effect of L-Def diet on plankton selectivity, metabolic growth, haemato-biochemical parameters, gut content and intestinal morphometry. Fish were fed a fixed ration of 18 g kg−0.8 d−1 for 10 weeks. During the experimental period, plankton species in the fish guts were identified and categorised to phylum level: phytoplankton phyla – Euglenophyta, Bacillariophyta, Cyanobacteria, and Chlorophyta; zooplankton phyla - Copepoda, Rotifera, and Cladocera. Fish in ponds receiving the L-Def diet selected positively for all phyto and zooplankton phyla but the selectivity was only significantly higher for Bacillariophyta with fish in ponds receiving the L-Con diet (P < 0.05). The numerical index (N%) of plankton in the gut content of Nile tilapia was higher in the L-Def treatment (48%) than in the L-Con treatment (38%). Fish growth and mortality were not significantly different between the two dietary treatments. The organo-somatic indices and haemato-biochemical parameters were similar between diets. The fish intestinal cross-sections from the L-Con and L-Def treatments showed numerous goblet cells in the proximal, middle, and distal gut, while the distal gut showed a partial serosal surface. Meanwhile, intestine morphometry sections of the distal villi length, submucosal layer, muscular layer and mid villi width, proximal / mid goblet cell counts and proximal lamina propria, were significantly higher in the L-Def treatment than in the L-Con treatment. The adverse impacts of dietary lysine deficiency on growth and fish health were reduced through plankton selectivity by Nile tilapia reared in ponds.

Kinetics of iron absorption from ferrous fumarate with and without galacto-oligosaccharides determined from stable isotope appearance curves in women

ABSTRACTBackgroundPrebiotic galacto-oligosaccharides (GOS) are novel enhancers of iron absorption from ferrous fumarate (FeFum). However, the mechanism(s) of this effect, and whether it occurs in the proximal or distal gut, are uncertain.ObjectivesWe studied: 1) in vitro, the effect of GOS on iron solubility and dialyzability from FeFum; 2) in volunteers, the absorption kinetics of FeFum given with and without GOS using stable isotope appearance curves (SIAC).MethodsWe measured iron solubility at various pH and dialyzability from FeFum with and without GOS. In crossover design, iron-depleted women [n = 11; median serum ferritin (SF) 15.2; IQR: 12.6–21.2 µg/L] received 2 14-mg iron doses as labeled (57Fe,58Fe) FeFum 14 d apart with and without 15 g GOS in randomly assigned order. Multiple blood samples were collected over a time period of 24 h and 14 d later to determine SIAC and fractional iron absorption (FIA), respectively. SIAC data were fitted using nonlinear mixed effects modeling to a 1-compartment model with first-order absorption, and AUC and time of peak serum isotope concentration (tmax) were calculated.ResultsIron dialyzability was 75% higher with GOS (P < 0.001) and iron solubility was more than doubled at pH 4 and 6 with GOS [both P < 0.001]. Mean ± SD AUC (5830.9 ± 4717.3 μg/min with GOS, 4454.0 ± 3260.7 μg/min for control), and median (IQR) FIA (20.3% (8.6%–38.7%) with GOS, and 15.6% (10.6%–24.8% f)or control) were not different with compared to without GOS (P = 0.064; P = 0.080). Mean ±SD tmax was not altered with GOS (3.08 ± 0.47 h with GOS; 2.80 ±0.50 h for control; P = 0.096). Iron bioavailability significantly increased with decreasing SF and this effect was significantly enhanced by GOS (P = 0.037, interaction of GOS with SF).ConclusionsGOS increases iron solubility from FeFum at physiological pH characteristic of the proximal duodenum. The absorption kinetics in vivo are consistent with effects on iron absorption in the proximal, rather than distal, parts of the gut. There was no overall effect of GOS on FIA in vivo, but the interaction of GOS and SF on FIA might benefit iron-deficient women, an effect potentially mediated by the higher solubility shown in vitro. This study was registered at clinicaltrials.gov as NCT03996421.

EP.FRI.719 Gastric Perforation Leading to The Diagnosis of Classic Ehlers Danlos syndrome: A Case Report of an Unusual Presentation

Abstract Background Ehlers-Danlos syndromes (EDS) are clinically and genetically heterogeneous group of heritable connective tissue disorders caused by defective collagen synthesis or structure. Vascular subtype (EDS IV) is reported to be associated with a higher incidence of gastrointestinal ruptures. The Most reported site of perforation was the colon particularly the sigmoid colon followed by small bowel. It is very rare to have stomach perforation. There were no reported cases among classic type I and II. In addition, this patient presented with Marfanoid habitus which may develop acute gastric volvulus in combined with pre-existing EDS, perforation can occur. Case description A 14-year-old girl attended our Emergency Department (ED) with abdominal pain and vomiting. Initially diagnosed with gastroenteritis and discharged once her condition improved. 24 hours later, she developed severe abdominal pain with recurrent vomiting with peritonitis evident on clinical examination. Initial KUB failed to show any free air, however, enhanced Computed Tomography (CT) revealed free air and proximal gut contrast extravasation. During exploratory laparotomy, an ischemia anterior and posterior gastric wall with gastric perforation was encountered. A free-hand partial gastrectomy was done. Her post-operative period was complicated with wound infection that managed successfully with Vacuum-assisted closure (VAC) dressing. She recovered well without gastrointestinal sequelae in 4 years follow up. Conclusions A high level of suspicion must be maintained for heritable systemic connective tissue diseases in any young patient with unusual spontaneous perforation. As these patients can develop life-threatening conditions, immediate intervention is required in addition to prepare for anticipated complications.

Gastric perforation leading to the diagnosis of classic Ehlers\u2013Danlos syndrome: a case report

BackgroundEhlers–Danlos syndrome is a clinically and genetically heterogeneous group of heritable connective tissue disorders caused by a defect in collagen synthesis and structure. The vascular subtype (Ehlers–Danlos syndrome IV) is reported to be associated with a higher incidence of gastrointestinal perforations. The most reported site of perforation is the colon, followed by the small bowel. Perforation of the stomach is very rare, and there are no reported cases to date of classic types I and II.Case presentationWe present the case of a 14-year-old Saudi girl who visited our emergency department with abdominal pain and vomiting. Initially, she was diagnosed with gastroenteritis and discharged once her condition stabilized. After 48 hours, she developed severe abdominal pain with recurrent vomiting and peritonitis evident on clinical examination. Initial abdominal x-ray failed to show any free air; however, enhanced computed tomography revealed free air and contrast extravasation in the proximal gut. During exploratory laparotomy, a large perforation was found on the anterior wall of the stomach due to the underlying ischemia. The posterior wall had ischemic mucosa with an intact healthy serosa. A free-hand partial gastrectomy was performed to resect all ischemic parts of the stomach. Detailed examinations and laboratory workup were carried out after the surgery to figure out the possible underlying cause. The clinical findings during the physical examination supported marfanoid features. Marfan’s syndrome and related disorders sequencing panel was requested, and Deoxyribonucleic acid (DNA) samples were sent. Given results were supporting the diagnosis of classical Ehlers–Danlos syndrome, the patient was labeled as a case of Ehlers–Danlos syndrome. During the postoperative period, she developed a wound infection that was managed successfully with vacuum-assisted closure dressing. She recovered well without gastrointestinal sequelae in the 4 years of follow-up.ConclusionsHeritable systemic connective tissue diseases must be given serious consideration in young patients with unusual spontaneous perforation. Such patients might develop life-threatening conditions that require immediate intervention. Hence, correct and timely diagnosis is important to prepare for the anticipated complications.