Risk Factors Associated with Unsuccessful High-Resolution Manometry.

High-resolution manometry (HRM) is used to evaluate the esophageal motor function. Unfortunately, there are times when testing cannot be performed accurately. Our study aimed to quantify the occurrence of failed HRM and identify the associated risk factors. HRM tests were retrospectively collected between September 2021 and August 2022. HRM reports that could not be interpreted based on standard HRM protocol as per Chicago guidelines were classified as failed tests. Information reviewed included testing indications, patient demographics, previous medical/symptom history, and follow-up testing for failed HRM. We then compared patients with successful vs. unsuccessful HRM based on our pre-specified factors. 152 HRM tests were performed, of which 28 tests (18%) were unsuccessful. Factors associated with failed manometry included a history of nausea/vomiting, dyspepsia, and achalasia. Patients who were unable to tolerate the probe during testing were more likely to have a history of dyspepsia (OR 20.3, p = < 0.001) and/or nausea/vomiting (OR 13.8, p = < 0.001). A history of achalasia was found to have an odds ratio of 13.2 when examining failure because of curling of the manometry catheter (p = 0.012). All seven patients who had repeat HRM with endoscopic placement were successful in obtaining diagnostic information. There are two groups that have risk factors for unsuccessful HRM testing. A history of nausea/vomiting and dyspepsia symptoms were associated with being unable to tolerate the manometry probe. The second group comprises patients with a history of achalasia in whom probe curling is more common. Future research targeting these risk factors may minimize diagnostic and treatment delays.

Altered Esophageal Smooth Muscle Phenotype in Achalasia.

Background/Aims: Achalasia is a disorder characterized by impairment in lower esophageal sphincter relaxation and esophageal aperistalsis, caused primarily by loss of inhibitory innervation. However, little is known about associated changes in esophageal smooth muscle. We examined the contractile phenotype and innervation of the circular smooth muscle, as well as inflammatory status, and correlated these with patient-specific parameters. Methods: Circular smooth muscle biopsies were obtained in consecutive patients with achalasia undergoing peroral endoscopic myotomy. Axonal innervation and neurotransmitter subtypes were determined with immunocytochemistry, and this was used with quantitative Polymerase Chain Reaction (qPCR) to characterize smooth muscle proliferation and cellular phenotype, as well as collagen expression. These were compared to control tissue obtained at esophagectomy and correlated with patient demographic factors including age, onset of symptoms, and Eckhardt score. Results: Biopsies of smooth muscle were obtained from 25 patients with achalasia. Overall, there was increased mast cell number and collagen deposition but increased smooth muscle cell proliferation vs control. There was a striking drop in axon density over controls, with no differences among subtypes of achalasia. Immunocytochemical analysis showed increased expression of the contractile marker α-smooth muscle actin, principally in Type 1 achalasia, that increased with disease duration, while qPCR identified increased mRNA for smoothelin with decreased myosin heavy chain and collagen 3a1, but not collagen 1a1. Conclusions: The thickened circular smooth muscle layer in achalasia is largely denervated, with an altered contractile phenotype and fibrosis. Biopsies obtained during peroral endoscopic myotomy provide a means to further study the pathophysiology of achalasia.

The epidemiology and impact of disorders of gut-brain interaction in Canada: Results from the Rome Foundation Global Epidemiologic Study.

BACKGROUND: Using data from the Rome Foundation Global Epidemiology Study examining prevalence of disorders of gut-brain interaction (DGBI) in 33 countries, this study explored the prevalence of all 22 disorders in Canada. It examined differences related to geography and sociodemographic factors, health impact and compared these findings to other countries. METHODS: The Rome Foundation survey was conducted through the Internet, targeting 2000 Canadian participants. The survey used the validated ROME IV diagnostic questionnaire to identify the 22 DGBI and an in-depth supplemental questionnaire that included quality assurance techniques. KEY RESULTS: There were 2029 respondents with a mean age of 48 years and 50% females. Diagnostic criteria for at least one of the 22 DGBI were met by 41.3%, similar to other countries. Functional constipation, functional diarrhea, and unspecified functional bowel disorders were most common. Irritable bowel syndrome prevalence was 4.2% using Rome IV and 10.1% using ROME III criteria. DGBI were associated with poorer quality of life and increased psychological symptom scores. Prevalence of DGBI ranged from 48% in Quebec to 36% in British Columbia. Prescription pain medication was reported by 17% with DGBI compared to 9% without DGBI and correlated negatively with mental health and physical well-being. Quebecers and francophones were in limited number but reported higher DGBI prevalence and symptom severity compared to others. CONCLUSIONS & INFERENCES: A large proportion of Canadians suffer from DGBI which seriously impact their well-being. Findings highlight the need for further research and education, including understanding whether significant regional and cultural differences contribute to DGBI.

Does Point-of-Care Ultrasound Change the Needle Insertion Location During Routine Bedside Paracentesis?

BACKGROUND: Paracentesis is a bedside procedure to obtain ascitic fluid from the peritoneum. Point-of-care ultrasound (POCUS) improves the safety of some medical procedures. However, the evidence supporting its utility in paracentesis is limited. OBJECTIVE: We aimed to assess if POCUS would yield a user-preferred site for needle insertion compared to conventional landmarking, defined as a ≥ 5 cm change in location. DESIGN: This was a prospective non-randomized trial comparing a POCUS-guided site to the conventional anatomic site in the same patient. PARTICIPANTS: Adult patients at Kingston Health Sciences Centre undergoing paracentesis were included. INTERVENTIONS: Physicians landmarked using conventional technique and compared this to a POCUS-guided site. The paracentesis was performed at whatever site was deemed optimal, if safe to do so. MAIN MEASURES: Data collected included the distance from the two sites, depth of fluid pockets, and anatomic considerations. KEY RESULTS: Forty-five procedures were performed among 30 patients and by 24 physicians, who were primarily in their PGY 1 and 2 years of training (33% and 31% respectively). Patients' ascites was mostly due to cirrhosis (84%) predominantly due to alcohol (47%) and NAFLD (34%). Users preferred the POCUS-guided site which resulted in a change in needle insertion ≥ 5 cm from the conventional anatomic site in 69% of cases. The average depth of fluid was greater at the POCUS site vs. the anatomic site (5.4±2.8 cm vs. 3.0±2.5 cm, p < 0.005). POCUS deflected the needle insertion site superiorly and laterally to the anatomic site. The POCUS site was chosen (1) to avoid adjacent organs, (2) to optimize the fluid pocket, and (3) due to abdominal wall considerations, such as pannus. Six cases landmarked anatomically were aborted when POCUS revealed inadequate ascites. CONCLUSIONS: POCUS changes the needle insertion site from the conventional anatomic site for most procedures, due to optimizing the fluid pocket and safety concerns, and helped avoid cases where an unsafe volume of ascites was present.

Are psychological interventions effective in treating functional dyspepsia? A systematic review and meta-analysis.

BACKGROUND AND AIM: Functional dyspepsia (FD) is a common gastrointestinal disorder, characterized primarily by postprandial fullness or early satiety and/or pain in the epigastrium with no endoscopic evidence of disease. Psychological therapies have been adapted to the treatment of disordered gut-brain interaction such as FD. We sought to determine if psychological interventions were efficacious in providing symptom management and improving health-related quality of life in patients suffering from FD. METHODS: Data were sorted that belonged to Embase (1947 to January 2020), PsychINFO (1806 to January 2020), and Ovid MEDLINE (1946 to January 2020). Randomized controlled trials using a psychological intervention in adults meeting relevant diagnostic criteria for FD were included. Data including symptom scores and quality of life measures were extracted. A random-effect model meta-analysis with standardized mean differences was used. RESULTS: Nine randomized controlled trials were identified that met our inclusion criteria. These were small, single-centered studies and used varying psychological therapies. Three studies had a sham treatment arm, leading to a high risk of bias in the remaining studies. All the studies reported beneficial effects of psychological treatment on patient's symptoms, some of which persisted up to 1 year. Psychological intervention was associated with an improvement in global FD symptom scores (standardized differences in means -1.33, 95% confidence interval -1.97 to -0.68). CONCLUSIONS: Despite the limited data, the available evidence suggests that psychological therapy is beneficial in treating patients with FD and should be considered by treating physicians if available and patients are willing. Large well-designed, sham controlled trials are needed for this extremely common disorder.

Microbial Disruption of Autophagy Alters Expression of the RISC Component AGO2, a Critical Regulator of the miRNA Silencing Pathway.

BACKGROUND: Autophagy is implicated in Crohn's disease (CD) pathogenesis. Recent evidence suggests autophagy regulates the microRNA (miRNA)-induced silencing complex (miRISC). Therefore, autophagy may play a novel role in CD by regulating expression of miRISC, thereby altering miRNA silencing. As microbes associated with CD can alter autophagy, we hypothesized that microbial disruption of autophagy affects the critical miRISC component AGO2. METHODS: AGO2 expression was assessed in epithelial and immune cells, and intestinal organoids with disrupted autophagy. Microarray technology was used to determine the expression of downstream miRNAs in cells with defective autophagy. RESULTS: Increased AGO2 was detected in autophagy-deficient ATG5-/- and ATG16-/- mouse embryonic fibroblast cells (MEFs) in comparison with wild-type MEFs. Chemical agents and VacA toxin, which disrupt autophagy, increased AGO2 expression in MEFs, epithelial cells lines, and human monocytes, respectively. Increased AGO2 was also detected in ATG7-/- intestinal organoids, in comparison with wild-type organoids. Five miRNAs were differentially expressed in autophagy-deficient MEFs. Pathway enrichment analysis of the differentially expressed miRNAs implicated signaling pathways previously associated with CD. CONCLUSIONS: Taken together, our results suggest that autophagy is involved in the regulation of the critical miRISC component AGO2 in epithelial and immune cells and primary intestinal epithelial cells. We propose a mechanism by which autophagy alters miRNA expression, which likely impacts the regulation of CD-associated pathways. Furthermore, as enteric microbial products can manipulate autophagy and AGO2, our findings suggest a novel mechanism by which enteric microbes could influence miRNA to promote disease.

Probiotics normalize the gut-brain-microbiota axis in immunodeficient mice.

The gut-brain-microbiota axis is increasingly recognized as an important regulator of intestinal physiology. Exposure to psychological stress causes activation of the hypothalamic-pituitary-adrenal (HPA) axis and causes altered intestinal barrier function, intestinal dysbiosis, and behavioral changes. The primary aim of this study was to determine whether the effects of psychological stress on intestinal physiology and behavior, including anxiety and memory, are mediated by the adaptive immune system. Furthermore, we wanted to determine whether treatment with probiotics would normalize these effects. Here we demonstrate that B and T cell-deficient Rag1(-/-) mice displayed altered baseline behaviors, including memory and anxiety, accompanied by an overactive HPA axis, increased intestinal secretory state, dysbiosis, and decreased hippocampal c-Fos expression. Both local (intestinal physiology and microbiota) and central (behavioral and hippocampal c-Fos) changes were normalized by pretreatment with probiotics, indicating an overall benefit on health conferred by changes in the microbiota, independent of lymphocytes. Taken together, these findings indicate a role for adaptive immune cells in maintaining normal intestinal and brain health in mice and show that probiotics can overcome this immune-mediated deficit in the gut-brain-microbiota axis.

Studies on the toxicity of an aqueous suspension of C60 nanoparticles using a bacterium (gen. Bacillus) and an aquatic plant (Lemna gibba) as in vitro model systems.

The increasing use of C60 nanoparticles and the diversity of their applications in industry and medicine has led to their production in a large scale. C60 release into wastewaters and the possible accumulation in the environment has raised concerns about their ecotoxicological impact. In the present study, an aqueous suspension of C60 nanoparticles was prepared and its potential toxicity studied in laboratory, using a bacterium (Bacillus stearothermophilus) and an aquatic plant (Lemna gibba) as model systems. C60 nanoparticles inhibited the growth of L. gibba, in contrast to that of the bacterium. Consistently, the ultrastructure and respiratory activity of bacterial cells were not affected by C60, but the contents of chlorophylls a and b and chloroplast oxygen production decreased considerably in L. gibba. Altogether, our results suggest that C60 aqueous dispersions must be viewed as an environmental pollutant, potentially endangering the equilibrium of aquatic ecosystems.

Alternative splicing results in RET isoforms with distinct trafficking properties.

RET encodes a receptor tyrosine kinase that is essential for spermatogenesis, development of the sensory, sympathetic, parasympathetic, and enteric nervous systems and the kidneys, as well as for maintenance of adult midbrain dopaminergic neurons. RET is alternatively spliced to encode multiple isoforms that differ in their C-terminal amino acids. The RET9 and RET51 isoforms display unique levels of autophosphorylation and have differential interactions with adaptor proteins. They induce distinct gene expression patterns, promote different levels of cell differentiation and transformation, and play unique roles in development. Here we present a comprehensive study of the subcellular localization and trafficking of RET isoforms. We show that immature RET9 accumulates intracellularly in the Golgi, whereas RET51 is efficiently matured and present in relatively higher amounts on the plasma membrane. RET51 is internalized faster after ligand binding and undergoes recycling back to the plasma membrane. This differential trafficking of RET isoforms produces a more rapid and longer duration of signaling through the extracellular-signal regulated kinase/mitogen-activated protein kinase pathway downstream of RET51 relative to RET9. Together these differences in trafficking properties contribute to some of the functional differences previously observed between RET9 and RET51 and establish the important role of intracellular trafficking in modulating and maintaining RET signaling.

Matrix metalloproteinase 9 contributes to gut microbe homeostasis in a model of infectious colitis.

BACKGROUND: Inflammatory bowel diseases are associated with increased expression of zinc-dependent Matrix Metalloproteinase 9 (MMP-9). A stark dysregulation of intestinal mucosal homeostasis has been observed in patients with chronic inflammatory bowel diseases. We therefore sought to determine the contribution of MMP-9 to the pathogenesis of Citrobacter rodentium-induced colitis and its effects on gut microbiome homeostasis. RESULTS: Wild-type and MMP-9-/- mice aged 5-6 weeks were challenged with C. rodentium by orogastric gavage and sacrificed either 10 or 30 days post-infection. Disease severity was assessed by histological analysis of colonic epithelial hyperplasia and by using an in vivo intestinal permeability assay. Changes in the inflammatory responses were measured by using qPCR, and the composition of the fecal microbiome evaluated with both qPCR and terminal restriction fragment length polymorphism. Activation and localization of MMP-9 to the apical surface of the colonic epithelium in response to C. rodentium infection was demonstrated by both zymography and immunocytochemistry. The pro-inflammatory response to infection, including colonic epithelial cell hyperplasia and barrier dysfunction, was similar, irrespective of genotype. Nonmetric multidimensional scaling of terminal restriction fragments revealed a different fecal microbiome composition and C. rodentium colonization pattern between genotypes, with MMP-9-/- having elevated levels of protective segmented filamentous bacteria and interleukin-17, and lower levels of C. rodentium. MMP-9-/- but not wild-type mice were also protected from reductions in fecal microbial diversity in response to the bacterial enteric infection. CONCLUSIONS: These results demonstrate that MMP-9 expression in the colon causes alterations in the fecal microbiome and has an impact on the pathogenesis of bacterial-induced colitis in mice.

Probiotics are effective for the prevention and treatment of Citrobacter rodentium-induced colitis in mice.

BACKGROUND: Probiotics prevent disease induced by Citrobacter rodentium, a murine-specific enteric pathogen. Whether probiotics can be used to interrupt the infectious process following initiation of infection was determined. METHODS: C57BL/6 adult and neonatal mice were challenged with C. rodentium, and a probiotic mixture containing Lactobacillus helveticus and Lactobacillus rhamnosus was provided 1 week before bacterial challenge, concurrently with infection, or 3 days and 6 days after infection. Mice were sacrificed 10 days after infection, and disease severity was assessed by histological analysis and in vivo intestinal permeability assay. Inflammatory pathways and the composition of the fecal microbiome were assessed in adult mice. RESULTS: Preadministration and coadministration of probiotics ameliorated C. rodentium-induced barrier dysfunction, epithelial hyperplasia, and binding of the pathogen to host colonocytes in adults, with similar findings in neonatal mice. Upregulated tumor necrosis factor α and interferon γ transcripts were suppressed in the pretreated probiotic group, whereas interleukin 17 transcription was suppressed with probiotics given up to 3 days after infection. Probiotics promoted transcription of interleukin 10 and FOXP3, and increased follicular T-regulatory cells in pretreatment mice. C. rodentium infection resulted in an altered fecal microbiome, which was normalized with probiotic intervention. CONCLUSIONS: This study provides evidence that probiotics can prevent illness and treat disease in an animal model of infectious colitis.

Bacterial infection causes stress-induced memory dysfunction in mice.

BACKGROUND: The brain-gut axis is a key regulator of normal intestinal physiology; for example, psychological stress is linked to altered gut barrier function, development of food allergies and changes in behaviour. Whether intestinal events, such as enteric bacterial infections and bacterial colonisation, exert a reciprocal effect on stress-associated behaviour is not well established. OBJECTIVE: To determine the effects of either acute enteric infection or absence of gut microbiota on behaviour, including anxiety and non-spatial memory formation. METHODS: Behaviour was assessed following infection with the non-invasive enteric pathogen, Citrobacter rodentium in both C57BL/6 mice and germ-free Swiss-Webster mice, in the presence or absence of acute water avoidance stress. Whether daily treatment with probiotics normalised behaviour was assessed, and potential mechanisms of action evaluated. RESULTS: No behavioural abnormalities were observed, either at the height of infection (10 days) or following bacterial clearance (30 days), in C rodentium-infected C57BL/6 mice. When infected mice were exposed to acute stress, however, memory dysfunction was apparent after infection (10 days and 30 days). Memory dysfunction was prevented by daily treatment of infected mice with probiotics. Memory was impaired in germ-free mice, with or without exposure to stress, in contrast to conventionally reared, control Swiss-Webster mice with an intact intestinal microbiota. CONCLUSIONS: The intestinal microbiota influences the ability to form memory. Memory dysfunction occurs in infected mice exposed to acute stress, while in the germ-free setting memory is altered at baseline.