Distal Gastrointestinal Tract Research Articles

Oral Sulfate Solution Versus Polyethylene Glycol Solution for Bowel Preparation for Colonoscopy: A Randomised Controlled Trial in a Tertiary Care Centre

Colonoscopy is an indispensable procedure in the armamentarium of an endoscopist to diagnose the pathologies of the distal gastrointestinal tract. A well-prepared bowel is mandatory for an ideal colonoscopy which facilitates in clinching the diagnosis. The aim of the study was to compare the palatability, tolerance, volume and side effects of PEG solution as well as OSS for bowel preparation and also to assess the quality of bowel preparation using Boston Bowel Preparation Scale (BBPS) by the colonoscopist in a controlled, randomised, and double- blinded manner.

Towards an EKG for SBO: A Neural Network for Detection and Characterization of Bowel Obstruction on CT.

A neural network was developed to detect and characterize bowel obstruction, a common cause of acute abdominal pain. In this retrospective study, 202 CT scans of 165 patients with bowel obstruction from March to June 2022 were included and partitioned into training and test data sets. A multi-channel neural network was trained to segment the gastrointestinal tract, and to predict the diameter and the longitudinal position ("longitude") along the gastrointestinal tract using a novel embedding. Its performance was compared to manual segmentations using the Dice score, and to manual measurements of the diameter and longitude using intraclass correlation coefficients (ICC). ROC curves as well as sensitivity and specificity were calculated for diameters above a clinical threshold for obstruction, and for longitudes corresponding to small bowel. In the test data set, Dice score for segmentation of the gastrointestinal tract was 78 ± 8%. ICC between measured and predicted diameters was 0.72, indicating moderate agreement. ICC between measured and predicted longitude was 0.85, indicating good agreement. AUROC was 0.90 for detection of dilated bowel, and was 0.95 and 0.90 for differentiation of the proximal and distal gastrointestinal tract respectively. Overall sensitivity and specificity for dilated small bowel were 0.83 and 0.90. Since obstruction is diagnosed based on the diameter and longitude of the bowel, this neural network and embedding may enable detection and characterization of this important disease on CT.

Structural stability and release properties of emulsion-alginate beads under gastrointestinal conditions

Ca-alginate beads are promising vehicles for targeted release, amongst others for oil. The release pattern is largely determined by the structural stability of such hydrogels (i.e., shrinking, swelling, or even disintegration), albeit that these processes are poorly understood. We designed a range of alginate hydrogel beads with different alginates (M/G ratio 1.2 or 2.1) and oil content (20–54%), and monitored their behaviour under simulated gastrointestinal conditions (INFOGEST protocol).Low M/G ratio emulsion-alginate gels were mechanically stronger than their counterparts prepared with high M/G ratio alginate. Beads prepared with high M/G ratio alginate swelled and disintegrated under gastric and intestinal conditions, with a high oil load contributing to reduced structural stability. This could be mitigated by using a higher Ca2+ concentration (10 mM), confirming that calcium plays a key role in structural stability.The actual release of fatty acids was co-determined by the properties of the hydrogel and their disintegration, which was further enhanced by extended oil digestion. Depending on the release pattern required to achieve a target, both mechanisms can be considered. For release in the more distal gastrointestinal (GI) tract, the structure needs to be maintained for longer, which is possible through the use of low M/G ratio alginate in combination with higher Ca2+ concentration (10 mM). These insights contribute to the development of alginate-based vehicles that can deliver their payload target to the small intestine or colon.

Osteopontin enhances the probiotic viability of Bifidobacteria in pectin-based microencapsulation subjected to in vitro infant gastrointestinal digestion

Encapsulation is usually adapted to improve probiotic viability under adverse environmental conditions, and used to ensure adequate amount of probiotics reach the distal gastrointestinal tract and achieve probiotic action. In the present study, sub-species of Bifidobacterium were encapsulated with pectin or alginate by an emulsification/internal gelation method. Bovine milk osteopontin (OPN) was used as an excipient with potential to improve the protection of the probiotics during processing and/or gastrointestinal digestion. Here, we reported that the use of OPN significantly improved probiotic viability during freeze drying and simulated infant gastrointestinal digestion. The viability of Bifidobacterium bifidum R0071 and Bifidobacterium breve M-16V loaded in pectin-based freeze-dried microcapsules with 0.5% OPN addition remained at over 106 CFU g−1 after simulated infant digestion. In comparison, the microcapsules without OPN had a probiotic viability of only 103 CFU g−1. The protective effect of OPN was proven to be dose dependent. For example, the B. bifidum R0071 viability was 6.1 log CFU·g−1 after the gastrointestinal digestion for the pectin-based microencapsulation without OPN addition, while the value remained at 8.4 log CFU·g−1 when OPN concentration reached 5%. In this study, we also demonstrated that extracellular calcium ions could chelate with OPN molecules which could be attributed to preventing damage of the probiotic cell wall and the improved protective effective of microencapsulation. Overall, these results provide new insight into the use of OPN as a microencapsulation excipient to promote probiotic viability and its potential future application in functional foods such as infant formula with enhanced gut health.

PH-dependent pressure-sensitive colonic capsules for the delivery of aqueous bacterial suspensions

Microbiome-based therapies hold great promise for treating various diseases, but the efficient delivery of live bacteria to the colon remains a challenge. Furthermore, current oral formulations, such as lyophilized bacterial capsules or tablets, are produced using processes that can decrease bacterial viability. Consequently, high dosages are required to achieve efficacy. Herein, we report the design of pressure-sensitive colonic capsules for the encapsulation and delivery of aqueous suspensions of live bacteria. The capsules consisted of 2 functional thin-films (hydrophobic and enteric) of ethyl cellulose and Eudragit S100 dip-coated onto hydroxypropyl methylcellulose molds. The capsules could be loaded with aqueous media and provide protection against acidic fluids and, to some extent, oxygen diffusion, suggesting their potential suitability for delivering anaerobic bacterial strains. Disintegration and mechanical studies indicated that the capsules could withstand transit through the stomach and upper/proximal small intestinal segments and rupture in the ileum/colon. In vitro studies showed that bacterial cells (anaerobic and aerobic commensals) remained highly viable (74–98%) after encapsulation and exposure to the simulated GI tract conditions. In vivo studies with a beagle dog model revealed that 67% of the capsules opened after 3.5 h, indicating content release in the distal gastrointestinal tract. These data demonstrate that live aqueous bacterial suspensions comprised of both aerobic and anaerobic commensals can be encapsulated and in the future might be efficiently delivered to the distal gastrointestinal tract, suggesting the practical applications of these capsules in microbiome-based therapies.

Bacillus indicus and Bacillus subtilis as alternative health and colouration promoters to synthetic astaxanthin in cyprinid aquaculture species

One of the largest challenges for the sustainable development of global aquaculture is the threat of infectious diseases. Preventative strategies that reduce antibiotic use are required to ensure fish health, minimise infectious diseases and subsequent pharmaceutical interventions. Recent strategies involve health-promoting feed supplements, such as astaxanthin and probiotic bacteria. Astaxanthin, a widely used carotenoid, offers colouration and antioxidant properties that can improve fish growth and fish survival when challenged with a pathogen. Probiotics can provide fish with a range of health benefits ranging from enhanced feed digestion, synthesis of vitamins, boost of innate immune response and active defence against potential pathogens.In this study, we tested if novel probiotic blends (Bacillus subtilis and/or Bacillus indicus) can be used as alternative health and/or colouration supplements to astaxanthin in two cyprinid species, mirror carp (Cyprinus carpio) and Red Comet goldfish (Carassius auratus auratus). Using experimental feed trials and 16S rRNA microbial profiling, the impact of the probiotic on fish growth and microbial community within the distal gastrointestinal tract was assessed. In addition, in mirror carp, blood samples were tested for immunology and haematological parameters, while in goldfish colouration of the skin was analysed.Mirror carp fed astaxanthin showed significantly increased growth whereas B. substilis /B.indicus supplementation had non-significant effects on growth performance. Our results provide the first insights into how the supplementation of astaxanthin changes the microbial composition in cyprinid species. In mirror carp, astaxanthin and the probiotic blend induce a significant shift in gut microbial communities. Mirror carp fed B. substilis/ B.indicus showed several indices of potential microbial and health benefits such as increased diversity, an abundance of potentially beneficial bacteria and enhancement of the phagocytic activity and creatinine blood levels. However, no effect on colouration, growth or the microbial community was found in goldfish, highlighting substantial species-specific differences in response to probiotics, in two closely related cyprinid species. Further research into the efficacy and site of colonization of supplemented bacteria in fish gastrointestinal tracts, and the mechanisms underlying the observed shifts in the host microbiota, is required to fully understand species-specific responses to probiotic supplementation.

CCK and GLP-1 response on enteroendocrine cells of semi-dynamic digests of hydrolyzed and intact casein

A semi-dynamic gastrointestinal device was employed to explore the link between protein structure and metabolic response upon digestion for two different substrates, a casein hydrolysate and the precursor micellar casein. As expected, casein formed a firm coagulum that remained until the end of the gastric phase while the hydrolysate did not develop any visible aggregate. Each gastric emptying point was subjected to a static intestinal phase where the peptide and amino acid composition changed drastically from that found during the gastric phase. Gastrointestinal digests from the hydrolysate were characterized by a high abundancy of resistant peptides and free amino acids. Although all gastric and intestinal digests from both substrates induced the secretion of cholecystokinin (CCK) and glucagon-like peptide-1 (GLP-1) in STC-1 cells, GLP-1 levels were maximum in response to gastrointestinal digests from the hydrolysate. The enrichment of protein ingredients with gastric-resistant peptides by enzymatic hydrolysis is proposed as strategy to deliver protein stimuli to the distal gastrointestinal tract to control food intake or type 2 diabetes.

Use of In Vitro Dynamic Colon Model (DCM) to Inform a Physiologically Based Biopharmaceutic Model (PBBM) to Predict the In Vivo Performance of a Modified-Release Formulation of Theophylline.

A physiologically based biopharmaceutic model (PBBM) of a modified-release formulation of theophylline (Uniphyllin Continus® 200 mg tablet) was developed and implemented to predict the pharmacokinetic (PK) data of healthy male volunteers by integrating dissolution profiles measured in a biorelevant in vitro model: the Dynamic Colon Model (DCM). The superiority of the DCM over the United States Pharmacopeia (USP) Apparatus II (USP II) was demonstrated by the superior predictions for the 200 mg tablet (average absolute fold error (AAFE): 1.1-1.3 (DCM) vs. 1.3-1.5 (USP II). The best predictions were obtained using the three motility patterns (antegrade and retrograde propagating waves, baseline) in the DCM, which produced similar PK profiles. However, extensive erosion of the tablet occurred at all agitation speeds used in USP II (25, 50 and 100 rpm), resulting in an increased drug release rate in vitro and overpredicted PK data. The PK data of the Uniphyllin Continus® 400 mg tablet could not be predicted with the same accuracy using dissolution profiles from the DCM, which might be explained by differences in upper gastrointestinal (GI) tract residence times between the 200 and 400 mg tablets. Thus, it is recommended that the DCM be used for dosage forms in which the main release phenomena take place in the distal GI tract. However, the DCM again showed a better performance based on the overall AAFE compared to the USP II. Regional dissolution profiles within the DCM cannot currently be integrated into Simcyp®, which might limit the predictivity of the DCM. Thus, further compartmentalization of the colon within PBBM platforms is required to account for observed intra-regional differences in drug distribution.

Intestinal GPR119 activation by microbiota-derived metabolites impacts feeding behavior and energy metabolism

ObjectiveThe gastrointestinal tract affects physiological activities and behavior by secreting hormones and generating signals through the activation of nutrient sensors. GPR119, a lipid sensor, is indirectly involved in the secretion of incretins, such as glucagon-like peptide-1 and glucose-dependent insulinotropic peptide, by enteroendocrine cells, while it directly stimulates insulin secretion by pancreatic beta cells. Since GPR119 has the potential to modulate metabolic homeostasis in obesity and diabetes, it has attracted interest as a therapeutic target. However, previous studies have shown that the deletion of Gpr119 in mice does not affect glucose homeostasis and appetite in either basal or high-fat diet-fed conditions. Therefore, the present study aimed to explore the role of GPR119 signaling system in energy metabolism and feeding behavior in mice. MethodsGpr119 knockout (KO) mice were generated using CRISPR-Cas9 gene-editing technology, and their feeding behavior and energy metabolism were evaluated and compared with those of wild type (WT) mice. ResultsUpon inducing metabolic stress via food deprivation, Gpr119 KO mice exhibited lower blood glucose levels and a higher body weight reduction compared to WT mice. Although food intake in WT and KO mice were similar under free-feeding conditions, Gpr119 KO mice exhibited increased food intake when they were refed after 24 h of food deprivation. Further, food-deprived Gpr119 KO mice presented shorter post-meal intervals and lower satiety for second and later meals during refeeding, resulting in increased food intake. Associated with this meal pattern, levels of oleoylethanolamide (OEA), an endogenous agonist of GPR119, in the luminal contents of the distal gastrointestinal tract were elevated within 2 h after refeeding. The large-intestinal infusion of OEA prolonged post-meal intervals and increased satiety in the first meal, but not the second meal. On the other hand, infusion of oleic acid increased cecal OEA levels at 2 h from the beginning of infusion, while prolonging post-meal intervals and increasing satiety on the meals that occurred approximately 2 h after the infusion. Cecal OEA levels were low in antibiotic-treated mice, suggesting that the gut microbiota partially synthesizes OEA from oleic acid. ConclusionsCollectively, our results indicate that the activation of gastrointestinal GPR119 by microbiota-produced OEA derived from oleic acid is associated with satiety control and energy homeostasis under energy shortage conditions.

The Role of Interleukin-17 in Tissue Protection during Radiation-Induced Oral Mucositis

<h3>Purpose/Objective(s)</h3> Oral mucositis (OM) is a common non-hematological side effect of radiotherapy and chemotherapy for head and neck cancers. Tissue injury associated with OM is painful, and can be difficult to treat. While the clinical progression of OM is understood, the signaling networks involved in the progression and resolution of damage have not been fully elucidated. Interleukin-17 (IL-17) is a proinflammatory cytokine produced by ‘type-17' lymphocytes, including T-helper 17 (Th17) cells. IL-17 is important in host protection against microbes, but it also can be pathogenic in autoimmune conditions. The role of IL-17 in other inflammatory disorders is not straightforward either. Due to the proliferative and inflammatory properties of IL-17 it is implicated in several malignancies, including esophageal cancer and tongue squamous cell carcinoma. Yet, in the distal gastrointestinal tract, IL-17 can be protective in tissue repair and maintenance. We set out to determine the role of IL-17 during severe ulceration of the oral mucosa associated with head and neck irradiation. We hypothesized that IL-17 is beneficial in repair of the oral mucosa after radiation exposure. <h3>Materials/Methods</h3> We approached this question by exposing wild-type (WT) mice and mice deficient in the receptor for IL-17, IL-17RA (<i>Il17ra</i>^−/−), to head and neck irradiation (HNI)-induced OM. Mice received a single dose of 22.5Gy of targeted irradiation using 6 MeV electrons from a linear accelerator. Tongue tissue was excised and processed for RNA-Seq analysis. The tissue was assessed macroscopically and histologically for damage, and the immune cells present in the oral tissue after HNI were determined by flow cytometry. <h3>Results</h3> RNA-Seq of oral tissue showed elevated expression of IL-17 and related immune pathways in response to HNI. Mice lacking IL-17RA exhibited markedly more severe OM. Restoration of the oral mucosa was compromised in the <i>Il17ra</i>^−/- mice and components associated with healing were diminished. The absence of IL-17RA also resulted in excess neutrophils and immunopathology compared to WT mice. <h3>Conclusion</h3> IL-17RA is required to prevent severe mucosal damage, and optimize healing after ionizing radiation. IL-17RA contributes to the proliferative capacity of the oral epithelia and maintains immune cell balance in the oral mucosa creating an environment conducive to restoration of the damaged tissue. This important contribution of IL-17 post-irradiation has implications for other cancer therapies related to the Th17 pathway.

Endoscopic diagnostics and treatment of “early” anal cancer

Anal cancer is a rare malignancy of the distal gastrointestinal tract, often associated with human papillomavirus, the most common sexually transmitted infection worldwide. Currently available screening methods for anal intraepithelial neoplasia, a precursor for anal cancer, combine anal Papanicolaou cytology and high resolution anoscopy with biopsy of suspicious lesions. It is well known that the anal canal is an area examined by proctologists, however, with endoscopic examination, cancer of the anal canal can also be detected, including in the early stages of the tumor process. When conducting an endoscopic examination of the lower gastrointestinal tract, the most common cause of incomplete examination of the colon during colonoscopy is poor preparation, less often—the anatomical features of the colon. An endoscopist, who examined the colon often skip the changes related to the lower ampullar part of the rectum and the anal canal, because of quickly removing the endoscope and not using the technique of examining this area in retrofl exed mode. This review describes the epidemiology of anal intraepithelial neoplasia and anal cancer and carcinogenesis in the population of the Russian Federation, presents an algorithm for examining patients at risk, diagnostic and therapeutic endoscopic techniques that allow timely diagnosis of pathological conditions of the lower ampulla of the rectum and anal canal, also presents modern possibilities of endoscopic treatment of this pathology.

Colonic Delivery of Nutrients for Sustained and Prolonged Release of Gut Peptides: A Novel Strategy for Appetite Management.

Obesity is one of the major global threats to human health and risk factors for cardiometabolic diseases and certain cancers. Glucagon-like peptide-1 (GLP-1) plays a major role in appetite and glucose homeostasis and recently the USFDA approved GLP-1 agonists for the treatment of obesity and type 2 diabetes. GLP-1 is secreted from enteroendocrine L-cells in the distal part of the gastrointestinal (GI) tract in response to nutrient ingestion. Endogenously released GLP-1 has a very short half-life of &lt;2min and most of the nutrients are absorbed before reaching the distal GI tract and colon, which hinders the use of nutritional compounds for appetite regulation. The review article focuses on nutrients that endogenously stimulate GLP-1 and peptide YY (PYY)secretion via their receptors in order to decrease appetite as preventive action. In addition, various delivery technologies such as pH-sensitive, mucoadhesive, time-dependent, and enzyme-sensitive systems for colonic targeting of nutrients delivery are described. Sustained colonic delivery of nutritional compounds could be one of the most promising approaches to prevent obesity and associated metabolic diseases by, e.g., sustained GLP-1 release.

Irinotecan decreases intestinal UDP-glucuronosyltransferase (UGT) 1A1 via TLR4/MyD88 pathway prior to the onset of diarrhea.

Irinotecan is a first-line treatment for colorectal cancer and the prodrug of 7-ethyl-10-hydroxy-camptothecin (SN-38). However, its fatal gastrointestinal (GI) toxicity raises serious concern. In liver, irinotecan generates its inactive metabolite, SN-38G via UDP-glucuronosyltransferase (UGT)1A1. Subsequently, SN-38G is excreted into GI tract where it is reactivated by microbiome to yield the toxic metabolite, SN-38. Activation of toll-like receptor (TLR)/myeloid differentiation primary response 88 (MyD88) by bacterial endotoxin decreases drug-metabolizing enzymes. In this study, we treated C57BL6/J mice with 50 mg/kg irinotecan once daily until observing grade 4 diarrhea. Mice were sacrificed on day0, day2 and day8. Based on the finding in C57BL6/J mice, we repeated the treatment in Tlr2-/-, Tlr4-/- and Myd88-/- mice to determine the impact of inflammation on UGT metabolism. Our toxicity study in C57BL6/J mice showed that mice started bloody diarrhea after 6 days' injection of irinotecan. Ugt1a1 expression in GI tract started decreasing after 24h since first dose, before the onset of diarrhea. In Tlr4-/- and Myd88-/- mice, no Ugt1a1 reduction was observed in distal GI tract after irinotecan injection. In Tlr2-/- mice, intestinal Ugt1a1 expression was down-regulated. Our results indicate that after two doses of irinotecan, mice started losing capability of detoxifying SN-38. TLR4 plays more important role in Ugt1a1 reduction than TLR2, despite that TLR2 and TLR4 share MyD88 as common adaptor protein. We concluded that irinotecan reduced intestinal Ugt1a1 via TLR4/MyD88 pathway, which eventually triggers the onset of diarrhea. Our finding unveils a novel mechanism underlying irinotecan-induced diarrhea and provides a new direction to prevent chemotherapy side effect.

Cutaneous Manifestations of Inflammatory Bowel Disease: A Basic Overview.

Inflammatory bowel disease (IBD) is a chronic inflammatory condition of the gastrointestinal (GI) tract that is subdivided into Crohn's disease (CD) and ulcerative colitis (UC). CD is characterized by involvement of the entire GI tract, while UC mainly affects the distal GI tract. Moreover, both CD and UC can present with extraintestinal manifestations (EIMs) of the disease affecting multiple organ systems including the hepatobiliary tract, kidney, bones, eyes, joints, and skin. These complications can cause significant morbidity and negatively impact the quality of life for IBD patients. Although the pathogenesis of EIMs is not clearly elucidated, it is postulated that the diseased GI mucosa similarly stimulates excess immune responses at the extraintestinal sites. Cutaneous EIMs occur in up to 15% of patients with IBD, often predating their IBD diagnosis. They are categorized into (1) specific, (2) reactive, (3) associated, and (4) treatment-induced. Here, we review the epidemiological, clinical, diagnostic, and histologic features of the most commonly described cutaneous EIMs of IBD along with their respective treatment options.

Mid-term outcomes of corrective surgery for anorectal malformations at Komfo Anokye Teaching Hospital, Ghana

BackgroundAnorectal malformations (ARM) are a spectrum of congenital defects of the distal gastrointestinal tract. Although faecal continence and the absence of constipation are recognised measures of outcome of curative surgery for ARM, evaluation of these measures can be challenging particularly in resource-poor countries. The difficulty of long-term follow- up in such settings and the other plausible causes of faecal incontinence other than the surgical procedure performed are the usual reasons. The Krickenbeck score for faecal incontinence is one objective way of measuring the surgical outcome following the surgical correction of ARM. We present the medium-term outcomes of the patients who had ARM surgically corrected at our hospital.&#x0D; Methods&#x0D; The folders of all patients who had curative surgery for ARM at our hospital (Komfo Anokye Teaching Hospital, Kumasi) from 2012-2016 were retrieved from the records department. Those who had had colostomy closure after curative surgery were identified. Data on age, sex, diagnosis and type of corrective surgery for these patients were documented. Telephone interviews of their mothers were then conducted and the incidence of faecal soiling and&#x0D; constipation recorded.&#x0D; Results&#x0D; A total of 46 interviews were concluded out of the 53 patients identified. The ages ranged from 8 - 107months with an interquartile range of 9-24months (SD 16.93) and a male-female ratio of 1:1. Most patients (61%, n=28) had voluntary bowel movements without faecal soiling (Krickenbeck grade 1) and 1 had persistent constipation. Urinary disturbances were experienced by 4 patients (incontinence-3, straining at micturiction-1).&#x0D; Conclusion&#x0D; Our study reveals acceptable medium-term outcomes for most patients who had corrective surgery for ARM at our centre. Long term follow-up and further evaluation of patients with faecal incontinence is necessary.

Genetic Background Influences Severity of Colonic Aganglionosis and Response to GDNF Enemas in the Holstein Mouse Model of Hirschsprung Disease.

Hirschsprung disease is a congenital malformation where ganglia of the neural crest-derived enteric nervous system are missing over varying lengths of the distal gastrointestinal tract. This complex genetic condition involves both rare and common variants in dozens of genes, many of which have been functionally validated in animal models. Modifier loci present in the genetic background are also believed to influence disease penetrance and severity, but this has not been frequently tested in animal models. Here, we addressed this question using Holstein mice in which aganglionosis is due to excessive deposition of collagen VI around the developing enteric nervous system, thereby allowing us to model trisomy 21-associated Hirschsprung disease. We also asked whether the genetic background might influence the response of Holstein mice to GDNF enemas, which we recently showed to have regenerative properties for the missing enteric nervous system. Compared to Holstein mice in their original FVB/N genetic background, Holstein mice maintained in a C57BL/6N background were found to have a less severe enteric nervous system defect and to be more responsive to GDNF enemas. This change of genetic background had a positive impact on the enteric nervous system only, leaving the neural crest-related pigmentation phenotype of Holstein mice unaffected. Taken together with other similar studies, these results are thus consistent with the notion that the enteric nervous system is more sensitive to genetic background changes than other neural crest derivatives.

Antagonistic Effects of Lipids Against the Anti-Escherichia coli and Anti-Salmonella Activity of Thymol and Thymol-β-d-Glucopyranoside in Porcine Gut and Fecal Cultures In Vitro.

Strategies are sought to reduce the carriage and dissemination of zoonotic pathogens and antimicrobial-resistant microbes within food-producing animals and their production environment. Thymol (an essential oil) is a potent bactericide in vitro but in vivo efficacy has been inconsistent, largely due to its lipophilicity and absorption, which limits its passage and subsequent availability in the distal gastrointestinal tract. Conjugation of thymol to glucose to form thymol-β-d-glucopyranoside can decrease its absorption, but in vivo passage of effective concentrations to the lower gut remains suboptimal. Considering that contemporary swine diets often contain 5% or more added fat (to increase caloric density and reduce dustiness), we hypothesized that there may be sufficient residual fat in the distal intestinal tract to sequester free or conjugated thymol, thereby limiting the availability and subsequent effectiveness of this biocide. In support of this hypothesis, the anti-Salmonella Typhimurium effects of 6 mM free or conjugated thymol, expressed as log10-fold reductions of colony-forming units (CFU) ml−1, were diminished 90 and 58%, respectively, following 24-h in vitro anaerobic fecal incubation (at 39°C) with 3% added vegetable oil compared to reductions achieved during culture without added oil (6.1 log10 CFU ml−1). The antagonistic effect of vegetable oil and the bactericidal effect of free and conjugated thymol against Escherichia coli K88 tested similarly were diminished 86 and 84%, respectively, compared to reductions achieved in cultures incubated without added vegetable oil (5.7 log10 CFU ml−1). Inclusion of taurine (8 mg/ml), bile acids (0.6 mg/ml), or emulsifiers such as polyoxyethylene-40 stearate (0.2%), Tween 20, or Tween 80 (each at 1%) in the in vitro incubations had little effect on vegetable oil-caused inhibition of free or conjugated thymol. Based on these results, it seems reasonable to suspect that undigested lipid in the distal gut may limit the effectiveness of free or conjugated thymol. Accordingly, additional research is warranted to learn how to overcome obstacles diminishing bactericidal activity of free and conjugated thymol in the lower gastrointestinal tract of food-producing animals.

Toll Like Receptors as Sensors of the Tumor Microbial Dysbiosis: Implications in Cancer Progression.

The microbiota is a complex ecosystem of active microorganisms resident in the body of mammals. Although the majority of these microorganisms resides in the distal gastrointestinal tract, high-throughput DNA sequencing technology has made possible to understand that several other tissues of the human body host their own microbiota, even those once considered sterile, such as lung tissue. These bacterial communities have important functions in maintaining a healthy body state, preserving symbiosis with the host immune system, which generates protective responses against pathogens and regulatory pathways that sustain the tolerance to commensal microbes. Toll-like receptors (TLRs) are critical in sensing the microbiota, maintaining the tolerance or triggering an immune response through the direct recognition of ligands derived from commensal microbiota or pathogenic microbes. Lately, it has been highlighted that the resident microbiota influences the initiation and development of cancer and its response to therapies and that specific changes in the number and distribution of taxa correlate with the existence of cancers in various tissues. However, the knowledge of functional activity and the meaning of microbiome changes remain limited. This review summarizes the current findings on the function of TLRs as sensors of the microbiota and highlighted their modulation as a reflection of tumor-associated changes in commensal microbiota. The data available to date suggest that commensal “onco-microbes” might be able to break the tolerance of TLRs and become complicit in cancer by sustaining its growth.

109 Is the equine gut colonised in utero by amniotic bacteria?

Until very recently it was believed that the uterus was sterile, with microbial colonisation of the foal's gastrointestinal tract (GIT) commencing at birth. GIT colonisation is an integral developmental process, critical for an optimal functioning immune system and ongoing health of the foal. The in utero colonisation paradigm has now challenged this concept, indicating that the amniotic fluid (AF) composition contains non-pathogenic microbiota that potentially commence colonisation of the GIT during the process of fetal swallowing. In turn, preparing the foal for life outside the womb. This study examined the presence of microbiota in the AF of healthy pregnant mares, identifying microorganisms that have the potential to colonise the fetal GIT in utero. AF samples were obtained from 14 healthy mares in the second stage of parturition, before the rupture of the amnion. When the amnion was clearly visible, the surface was seared with a sterile spatula and AF extracted with a sterile needle and syringe. The sample had no contact with any other surface of the mare and was immediately snap-frozen and stored at −80°C for microbiota analysis. Genomic DNA was extracted and processed for 16S rRNA gene amplicon sequencing, followed by bioinformatic analysis of the results using QIIME2 and CalypsoV8.84 software packages. Microbiota was detected in all AF samples. Communities were of low diversity, as indicated by the α diversity parameters Chao1, Evenness, Richness and Simpson's Index. Operational Taxonomic Units (OTUs) of the top 20 taxa showed some host variation. The most abundant OTUs across the majority of samples were classified to the phyla Opisthokonta (100% of samples), Bacteroidetes (79%), Actinobacteria (57%) and Firmicutes (57%). At the family level, Staphylococcacea (79%) and Microbacteriaceae (71%) were the most abundant. These microbes have been previously detected across other locations of the equine microbiome, including the oral, esophageal, distal GIT and vaginal ecosystems. Our results have demonstrated the presence of a complex microbial ecosystem of low diversity in the AF of healthy equine pregnancies, likely of maternal origin. The environmental and gastrointestinal microbes detected in the AF displayed similarities to other equine ecosystems, suggesting microbial translocation from the mare to the AF. Further research would be required to determine if these AF microbes are being ingested by the fetus, thus commencing GIT colonisation in utero . Deciphering microbial colonisation of the foal's GIT has important implications for maternal management before parturition and could lead to the development of strategies to enhance foal health outcomes.

NOX5 is expressed aberrantly but not a critical pathogenetic gene in Hirschsprung disease

BackgroundHirschsprung disease (HSCR) is a congenital disorder characterized by the absence of intramural ganglion cells in the distal gastrointestinal tract (GI), which results in tonic contraction of the aganglionic gut segment and functional intestinal obstruction. Recent studies have suggested NADPH oxidase 5 (NOX5) as a candidate risk gene for HSCR. In this study, we examined the function of NOX5 to verify its role in the development of the enteric nervous system (ENS).MethodsHSCR tissue specimens (n = 10) were collected at the time of pull-through surgery and control specimens (n = 10) were obtained at the time of colostomy closure in patients. The NOX5 expression in aganglionic and ganglionic segments of HSCR colon and normal colon were analyzed by immunohistochemistry (IHC), western blot and real-time quantitative PCR (qPCR). The gene expression levels and spatiotemporal expression spectrum of NOX5 in different development stages of zebrafish embryo were determined using qPCR and in-situ hybridization (ISH). The enteric nervous system in NOX5 Morpholino (MO) knockdown and wild type (WT) zebrafish embryo was analyzed by whole-mount immunofluorescence (IF). Intestinal transit assay was performed to analyze the gastrointestinal motility in NOX5 knockdown and control larvae.ResultsNOX5 is strongly expressed in the ganglion cells in the proximal segment of HSCR colons and all segments of normal colons. Moreover, the expression of NOX5 is markedly decreased in the aganglionic segment of HSCR colon compared to the ganglionic segment. In zebrafish, NOX5 mRNA level is the highest in the one cell stage embryos and it is decreased overtime with the development of the embryos. Interestingly, the expression of NOX5 appears to be enriched in the nervous system. However, the number of neurons in the GI tract and the GI motility were not affected upon NOX5 knockdown.ConclusionsOur study shows that NOX5 markedly decreased in the aganglionic segment of HSCR but didn’t involve in the ENS development of zebrafish. It implies that absence of intestinal ganglion cells may lead to down-regulation of NOX5.