A human autoimmune organoid model reveals IL-7 function in coeliac disease.

In vitro models of autoimmunity are constrained by an inability to culture affected epithelium alongside the complex tissue-resident immune microenvironment. Coeliac disease (CeD) is an autoimmune disease in which dietary gluten-derived peptides bind to the major histocompatibility complex (MHC) class II human leukocyte antigen molecules (HLA)-DQ2 or HLA-DQ8 to initiate immune-mediated duodenal mucosal injury1-4. Here, we generated air-liquid interface (ALI) duodenal organoids from intact fragments of endoscopic biopsies that preserve epithelium alongside native mesenchyme and tissue-resident immune cells as a unit without requiring reconstitution. The immune diversity of ALI organoids spanned T cells, B and plasma cells, natural killer (NK) cells and myeloid cells, with extensive T-cell and B-cell receptor repertoires. HLA-DQ2.5-restricted gluten peptides selectively instigated epithelial destruction in HLA-DQ2.5-expressing organoids derived from CeD patients, and this was antagonized by blocking MHC-II or NKG2C/D. Gluten epitopes stimulated a CeD organoid immune network response in lymphoid and myeloid subsets alongside anti-transglutaminase 2 (TG2) autoantibody production. Functional studies in CeD organoids revealed that interleukin-7 (IL-7) is a gluten-inducible pathogenic modulator that regulates CD8+ T-cell NKG2C/D expression and is necessary and sufficient for epithelial destruction. Furthermore, endogenous IL-7 was markedly upregulated in patient biopsies from active CeD compared with remission disease from gluten-free diets, predominantly in lamina propria mesenchyme. By preserving the epithelium alongside diverse immune populations, this human in vitro CeD model recapitulates gluten-dependent pathology, enables mechanistic investigation and establishes a proof of principle for the organoid modelling of autoimmunity.

The integrated relaxation pressure may not be an appropriate gold standard for deglutitive relaxation due to reliance on a single intragastric reference sensor.

BACKGROUND: Integrated relaxation pressure (IRP) calculation depends on the selection of a single gastric reference sensor. Variable gastric pressure readings due to sensor selection can lead to diagnostic uncertainty. This study aimed to examine the effect of gastric reference sensor selection on IRP measurement and diagnosis. METHODS: We identified high-resolution manometry (HRM) conducted between January and November 2017 with at least six intragastric reference sensors. IRP measurements and Chicago Classification 3.0 (CCv3) diagnoses were obtained for each of six gastric reference sensors. Studies were categorized as "stable" (no change in diagnosis) or "variable" (change in diagnosis with gastric reference selection). Variable diagnoses were further divided into "variable normal/dysmotility" (≥1 normal IRP measurement and ≥1 CCv3 diagnosis), or "variable dysmotility" (≥1 CCv3 diagnosis, only elevated IRP measurements). Bland-Altman plots were used to compare IRP measurements within HRM studies. KEY RESULTS: The analysis included 100 HRM studies, among which 18% had variable normal/dysmotility, and 10% had variable dysmotility. The average IRP difference between reference sensors was 6.7 mmHg for variable normal/dysmotility and 5.9 mmHg for variable dysmotility. The average difference between the proximal-most and distal-most sensors was -1.52 mmHg (lower limit of agreement -10.03 mmHg, upper limit of agreement 7.00 mmHg). CONCLUSIONS & INFERENCES: IRP values can vary greatly depending on the reference sensor used, leading to inconsistent diagnoses in 28% of HRM studies. Choosing the correct gastric reference sensor is crucial for accurate test results and avoiding misdiagnosis. Standardization of reference sensor selection or supportive testing for uncertain results should be considered.

The Impact of Intermittent Fasting on Patients With Suspected Gastroesophageal Reflux Disease.

GOAL: The aim was to investigate the short-term impact of time restricted feeding on patients with suspected gastroesophageal reflux disease (GERD). BACKGROUND: Lifestyle modifications are often suggested, but the role of diet in GERD is unclear. Intermittent fasting is popular in the media and has demonstrated potential benefits with weight loss and inflammatory conditions as well as alterations in gastrointestinal hormones. STUDY: Patients who were referred for 96-hour ambulatory wireless pH monitoring off proton pump inhibitor to investigate GERD symptoms were screened for eligibility. Patients were instructed to maintain their baseline diet for the first 2 days of pH monitoring and switch to an intermittent fasting regimen (16 consecutive hour fast and 8 h eating window) for the second 2 days. Objective measures of reflux and GERD symptom severity were collected and analyzed. RESULTS: A total of 25 participants were analyzed. 9/25 (36%) fully adhered to the intermittent fasting regimen, with 21/25 (84%) demonstrating at least partial compliance. Mean acid exposure time on fasting days was 3.5% versus 4.3% on nonfasting days. Intermittent fasting was associated with a 0.64 reduction in acid exposure time (95% CI: -2.32, 1.05). There was a reduction in GERD symptom scores of heartburn and regurgitation during periods of intermittent fasting (14.3 vs. 9.9; difference of -4.46, 95% CI: -7.6,-1.32). CONCLUSIONS: Initial adherence to time restricted eating may be difficult for patients. There is weak statistical evidence to suggest that intermittent fasting mildly reduces acid exposure. Our data show that short-term intermittent fasting improves symptoms of both regurgitation and heartburn.

Quality Indicators for the Diagnosis and Management of Eosinophilic Esophagitis.

INTRODUCTION: Despite best practice recommendations for managing eosinophilic esophagitis (EoE), variation in care exists. METHODS: We used established methodology for quality indicator development to identify metrics to define quality for the treatment of EoE. RESULTS: Among 29 proposed quality indicator statements, 9 (31%) were adopted as highly valid across all categories. Two (22%) of these statements were identified as having existing or suspected quality gaps. DISCUSSION: We identified highly valid EoE quality indicators for adult gastroenterologists, which can be used for quality improvement with resulting benefits for patient outcomes.

KIR+CD8+ T cells suppress pathogenic T cells and are active in autoimmune diseases and COVID-19.

In this work, we find that CD8+ T cells expressing inhibitory killer cell immunoglobulin-like receptors (KIRs) are the human equivalent of Ly49+CD8+ regulatory T cells in mice and are increased in the blood and inflamed tissues of patients with a variety of autoimmune diseases. Moreover, these CD8+ T cells efficiently eliminated pathogenic gliadin-specific CD4+ T cells from the leukocytes of celiac disease patients in vitro. We also find elevated levels of KIR+CD8+ T cells, but not CD4+ regulatory T cells, in COVID-19 patients, correlating with disease severity and vasculitis. Selective ablation of Ly49+CD8+ T cells in virus-infected mice led to autoimmunity after infection. Our results indicate that in both species, these regulatory CD8+ T cells act specifically to suppress pathogenic T cells in autoimmune and infectious diseases.

An efficient urine peptidomics workflow identifies chemically defined dietary gluten peptides from patients with celiac disease.

Celiac disease (CeD) is an autoimmune disorder induced by consuming gluten proteins from wheat, barley, and rye. Glutens resist gastrointestinal proteolysis, resulting in peptides that elicit inflammation in patients with CeD. Despite well-established connections between glutens and CeD, chemically defined, bioavailable peptides produced from dietary proteins have never been identified from humans in an unbiased manner. This is largely attributable to technical challenges, impeding our knowledge of potentially diverse peptide species that encounter the immune system. Here, we develop a liquid chromatographic-mass spectrometric workflow for untargeted sequence analysis of the urinary peptidome. We detect over 600 distinct dietary peptides, of which ~35% have a CeD-relevant T cell epitope and ~5% are known to stimulate innate immune responses. Remarkably, gluten peptides from patients with CeD qualitatively and quantitatively differ from controls. Our results provide a new foundation for understanding gluten immunogenicity, improving CeD management, and characterizing the dietary and urinary peptidomes.

Eosinophilic Esophagitis: Incidence, Diagnosis, Management, and Future Directions.

Eosinophilic esophagitis (EoE) is an antigen-mediated esophageal disease defined by the presence of esophageal eosinophilia and symptoms of esophageal dysfunction. The pathophysiology involves an allergen-driven Th2 T cell response that triggers infiltration of eosinophils into the esophagus leading to inflammation, remodeling, and fibrosis. This results in disruption of esophageal function and accompanying symptoms - most notably dysphagia. Effective therapies target inflammation or fibrostenotic complications and include proton pump inhibitors, swallowed topical steroids, dietary exclusion, and dilation. Clinical trials testing promising biologic therapies are ongoing.

Human KIR + CD8 + T cells target pathogenic T cells in Celiac disease and are active in autoimmune diseases and COVID-19.

Previous reports show that Ly49 + CD8 + T cells can suppress autoimmunity in mouse models of autoimmune diseases. Here we find a markedly increased frequency of CD8 + T cells expressing inhibitory Killer cell Immunoglobulin like Receptors (KIR), the human equivalent of the Ly49 family, in the blood and inflamed tissues of various autoimmune diseases. Moreover, KIR + CD8 + T cells can efficiently eliminate pathogenic gliadin-specific CD4 + T cells from Celiac disease (CeD) patients' leukocytes in vitro . Furthermore, we observe elevated levels of KIR + CD8 + T cells, but not CD4 + regulatory T cells, in COVID-19 and influenza-infected patients, and this correlates with disease severity and vasculitis in COVID-19. Expanded KIR + CD8 + T cells from these different diseases display shared phenotypes and similar T cell receptor sequences. These results characterize a regulatory CD8 + T cell subset in humans, broadly active in both autoimmune and infectious diseases, which we hypothesize functions to control self-reactive or otherwise pathogenic T cells. ONE-SENTENCE SUMMARY: Here we identified KIR + CD8 + T cells as a regulatory CD8 + T cell subset in humans that suppresses self-reactive or otherwise pathogenic CD4 + T cells.

Gastrointestinal Eosinophil Responses in a Longitudinal, Randomized Trial of Peanut Oral Immunotherapy.

BACKGROUND & AIMS: Gastrointestinal side effects are common during oral immunotherapy (OIT) and eosinophilic esophagitis (EoE) is a potential complication. We aimed to characterize eosinophilic gastrointestinal responses to peanut OIT, in which peanut protein is given orally, with incremental increases in dose over time. METHODS: Twenty adults with IgE-mediated peanut allergy were randomly assigned to groups given peanut OIT (n = 15) or placebo (n = 5); 1 additional subject withdrew before randomization. Serial gastrointestinal biopsies were collected at baseline (n = 21, 0 weeks), following dose escalation (n = 10, 52 weeks), and during the maintenance phase (n = 11, 104 weeks). Endoscopic findings were characterized using the EoE endoscopic reference score. Biopsies were assessed for eosinophils per high-power field (eos/hpf) and other pathology features using EoE histologic scoring system scores. We performed immunohistochemical analyses of eosinophil peroxidase deposition, quantified using automated image analysis. RESULTS: At baseline, no subjects reported current gastrointestinal symptoms. However, 3 of the 21 subjects (14%) had esophageal peak eosinophil counts ≥15 eos/hpf and all subjects had dilated intercellular spaces (DIS). OIT induced or exacerbated esophageal eosinophilia (EE) at 52 weeks in most subjects (peak eosinophil counts >5 eos/hpf in 6 of 7 patients [86%]; peak eosinophil counts ≥15 eos/hpf in 4 of 7 patients [57%]). One subject met clinicopathologic criteria for EoE and withdrew; no significant changes in esophageal peak eosinophil counts were observed in the placebo group. EE in the OIT group corresponded with significant increases in EoE histologic scoring system scores and deposition of eosinophil peroxidase. In 4 of 6 participants (67%), OIT-induced EE and gastrointestinal eosinophilia resolved by the end of the maintenance phase. Gastrointestinal symptoms were not clearly associated with EE or gastrointestinal eosinophilia. CONCLUSIONS: In this pilot study, we found that peanut OIT-induced EE and gastrointestinal eosinophilia are usually transient and are not always associated with gastrointestinal symptoms. Clinicaltrials.gov no: NCT02103270.

Baseline impedance via manometry and ambulatory reflux testing are not equivalent when utilized in the evaluation of potential extra-esophageal gastroesophageal reflux disease.

BACKGROUND: Esophageal baseline impedance (BI) shows promise for the diagnosis of gastroesophageal reflux disease (GERD), but means of acquisition and relevance to extra-esophageal manifestations of GERD (EE-GERD) remain unclear. In this study we aim to (I) evaluate concordance between BI as measured by 24-hour pH-impedance (pH-MII) and high-resolution impedance manometry (HRIM), and (II) assess relationship to potential EE-GERD symptoms. METHODS: In this prospective open cohort study, patients presenting for outpatient HRIM and pH-MII studies were prospectively enrolled. All patients completed the GERD-HRQL, NOSE, and respiratory symptom index questionnaire (RSI), plus questions regarding wheezing and dental procedures. HRIM and pH-MII were evaluated with calculation of BI. Correlations were assessed using either Pearson's correlation or Spearman's rank coefficients. RESULTS: 70 HRIM patients were enrolled, 35 of whom underwent pH-MII. There was no correlation between BI measurements as assessed by HRIM and pH-MII proximally, but there was moderate-weak correlation distally (r=0.34 to 0.5). Distal acid exposure time correlated with distal BI only for measurements by pH-MII (rho= -0.5 to -0.65), and not by HRIM. There was no relationship between proximal acid exposure time and proximal BI. There were no correlations when comparing proximal or distal BI measurements, acid exposure times, and impedance events to symptoms. CONCLUSIONS: Concordance between BI as measured by HRIM and pH-MII is poor, especially proximally, suggesting that these two methods are not interchangeable. There is no correlation between BI both distally/proximally and symptoms of either GERD/EE-GERD, suggesting that many symptoms are unrelated to acid or that BI is not an adequate marker to assess EE-GERD symptoms.

Achalasia: physiology and diagnosis.

Achalasia is a rare motility disorder with incomplete relaxation of the lower esophageal sphincter and ineffective contractions of the esophageal body. It has been hypothesized that achalasia does not result from only one pathway but rather involves a combination of infectious, autoimmune, and familial etiological components. On the basis of other observations, a novel hypothesis suggests that a muscular form of eosinophilic esophagitis is involved in the pathophysiology of achalasia in some patients. This appears to progressively diminish the myenteric plexus at stage III, gradually destroy it at stage II, and finally eliminate it at stage I, the most advanced and final stage of achalasia. Although high-resolution manometry has identified these three different types of achalasia, another subset of patients with a normal-appearing sphincter relaxation has been proposed. Provocative maneuvers, such as the rapid drinking challenge, have recently been demonstrated to improve diagnosis in certain borderline patients, but have to be studied in more detail. However, whether the different types of achalasia will have a long-term impact on tailored therapies is still a matter of debate. Additionally, novel aspects of the standard timed barium swallow appear to be an important adjunct of diagnosis, as it has been shown to have a diagnostic as well as a predictive value.

The functional lumen imaging probe in gastrointestinal disorders: the past, present, and future.

The functional lumen imaging probe (FLIP) is a diagnostic tool that utilizes impedance planimetry to allow the assessment of luminal diameter and distensibility. It has been used primarily in esophageal diseases, in particular, in the assessment of achalasia, esophagogastric junction outflow obstruction, and eosinophilic esophagitis (EoE). The usage and publications have increased over the past decade and it is now an essential tool in the armamentarium of the esophagologist. Indications are emerging outside of the esophagus, in particular with regard to gastroparesis. Our paper will review the history of FLIP, optimal current usage, data for key esophageal disorders (including achalasia, reflux, and EoE), data for nonesophageal disorders, and our sense as to whether FLIP is ready for prime time, as well as gaps in evidence and suggestions for future research.

Eosinophilic esophagitis: updates on key unanswered questions.

Eosinophilic esophagitis (EoE) is a clinicopathologic disease characterized by symptoms of esophageal dysfunction and esophageal eosinophilia. In the last decade, there has been a dramatic increase in its prevalence for reasons that are not completely understood. The underlying pathophysiology involves an antigen-mediated TH 2 immune response that draws eosinophils to the esophagus, causing mucosal inflammation, esophageal remodeling, and fibrosis. This ultimately leads to esophageal dysfunction that most commonly manifests as dysphagia. In this review, we will discuss updates on key questions regarding the diagnosis and treatment of EoE.

Origins and clonal convergence of gastrointestinal IgE+ B cells in human peanut allergy.

B cells in human food allergy have been studied predominantly in the blood. Little is known about IgE+ B cells or plasma cells in tissues exposed to dietary antigens. We characterized IgE+ clones in blood, stomach, duodenum, and esophagus of 19 peanut-allergic patients, using high-throughput DNA sequencing. IgE+ cells in allergic patients are enriched in stomach and duodenum, and have a plasma cell phenotype. Clonally related IgE+ and non-IgE-expressing cell frequencies in tissues suggest local isotype switching, including transitions between IgA and IgE isotypes. Highly similar antibody sequences specific for peanut allergen Ara h 2 are shared between patients, indicating that common immunoglobulin genetic rearrangements may contribute to pathogenesis. These data define the gastrointestinal tract as a reservoir of IgE+ B lineage cells in food allergy.

Ninety-Six Hour Wireless Esophageal pH Study in Patients with GERD Shows that Restrictive Diet Reduces Esophageal Acid Exposure.

BACKGROUND: Prolonged (96 h) pH monitoring may explore the effect of diet on pH and symptoms in patients with GERD. AIMS: To assess the usefulness of a 96 h esophageal pH study in patients with GER symptoms under different diets (pro- and anti-GER). METHODS: Prospective study of 66 patients with GERD undergoing wireless 96 h pH monitoring. Two-day periods, one on liberal (pro-reflux) and another on restricted (anti-reflux) diet assessed esophageal acid exposure and symptoms. The primary end point was normalization of acid exposure time while on restricted diet. Secondary end point was a > 50% reduction in symptoms with restricted diet. RESULTS: Normal (pH time < 4 of < 6%) was found in 34 patients (51.5%) while on the initial 48 h (liberal) diet [median % time < 4: 3.2 (95% CI, 1.9, 4.0)] and remained normal while on restricted diet [median % time < 4: 2.6 (95% CI, 0.8, 3.4)]. Abnormal acid exposure (% pH time < 4: > 6%) was found in 32 patients (48.5%) while on initial 48 h liberal diet [median % time < 4: 10.5, (95% CI 8.9, 12.6)], and decreased significantly with restricted diet [median % time < 4: 4.5 (95% CI 3.1, 7.3)] (p = 0.001), and normalized with anti-GERD diet in 21 patients (65.6%). Only 11/66 patients were candidates for proton pump inhibitor (PPI) use; 34 had either normal pH studies or normalized them with restricted diet (n = 21). Symptoms did not improve with restricted diet. CONCLUSIONS: The 96-h esophageal pH study tests for GERD under pro- and anti-GER diets and allows minimization of PPI therapy to only 16.6% of patients.

Human Intestinal Enteroids Model MHC-II in the Gut Epithelium.

The role of intestinal epithelial cells (IECs) in mucosal tolerance and immunity remains poorly understood. We present a method for inducing MHC class II (MHC-II) in human enteroids, "mini-guts" derived from small intestinal crypt stem cells, and show that the intracellular MHC-II peptide-pathway is intact and functional in IECs. Our approach enables human enteroids to be used for novel in vitro studies into IEC MHC-II regulation and function during health and disease.

Baseline Gastrointestinal Eosinophilia Is Common in Oral Immunotherapy Subjects With IgE-Mediated Peanut Allergy.

Rationale: Oral immunotherapy (OIT) is an emerging treatment for food allergy. While desensitization is achieved in most subjects, many experience gastrointestinal symptoms and few develop eosinophilic gastrointestinal disease. It is unclear whether these subjects have subclinical gastrointestinal eosinophilia (GE) at baseline. We aimed to evaluate the presence of GE in subjects with food allergy before peanut OIT. Methods: We performed baseline esophagogastroduodenoscopies on 21 adults before undergoing peanut OIT. Subjects completed a detailed gastrointestinal symptom questionnaire. Endoscopic findings were assessed using the Eosinophilic Esophagitis (EoE) Endoscopic Reference Score (EREFS) and biopsies were obtained from the esophagus, gastric antrum, and duodenum. Esophageal biopsies were evaluated using the EoE Histologic Scoring System. Immunohistochemical staining for eosinophil peroxidase (EPX) was also performed. Hematoxylin and eosin and EPX stains of each biopsy were assessed for eosinophil density and EPX/mm2 was quantified using automated image analysis. Results: All subjects were asymptomatic. Pre-existing esophageal eosinophilia (>5 eosinophils per high-power field [eos/hpf]) was present in five participants (24%), three (14%) of whom had >15 eos/hpf associated with mild endoscopic findings (edema, linear furrowing, or rings; median EREFS = 0, IQR 0-0.25). Some subjects also demonstrated basal cell hyperplasia, dilated intercellular spaces, and lamina propria fibrosis. Increased eosinophils were noted in the gastric antrum (>12 eos/hpf) or duodenum (>26 eos/hpf) in 9 subjects (43%). EPX/mm2 correlated strongly with eosinophil counts (r = 0.71, p < 0.0001). Conclusions: Pre-existing GE is common in adults with IgE-mediated peanut allergy. Eosinophilic inflammation (EI) in these subjects may be accompanied by mild endoscopic and histologic findings. Longitudinal data collection during OIT is ongoing.

Use of Esophageal pH Monitoring to Minimize Proton-Pump Inhibitor Utilization in Patients with Gastroesophageal Reflux Symptoms.

BACKGROUND: Due to concerns about long-term PPI use in patients with acid reflux, we aimed at minimizing PPI use, either by avoiding initiating therapy, downscaling to other therapies, or introducing endoscopic or surgical options. AIMS: To examine the role of esophageal ambulatory pHmetry in minimizing PPI use in patients with heartburn and acid regurgitation. METHODS: Retrospective cohort analysis of patients with reflux symptoms, who underwent endoscopy, manometry, and ambulatory pHmetry to define the need for PPI. Patients were classified as: (1) never users; (2) partial responders to PPI; (3) users with complete response to PPI. Patients were then managed as: (1) PPI non-users; (2) PPI-initiated, and (3) PPI-continued. RESULTS: Of 286 patients with heartburn and regurgitation, 103 (36%) were found to have normal and 183 (64%) abnormal esophageal acid exposure (AET). In the normal AET group, 44/103 had not been treated and were not initiated on PPI. Of the 59 who had previously received PPI, 52 stopped and 7 continued PPI. Hence, PPI were avoided in 96/103 patients (93%). In the abnormal AET group, 61/183 had not been treated and 38 were initiated on PPI and 23 on other therapies. In the 122 patients previously treated with PPI, 24 were not treated with PPI, but with H2RAs, prokinetics, endoscopic, or surgical therapy. Hence, PPI therapy was avoided in 47/183 patients (26%). CONCLUSIONS: In patients with GER symptoms, esophageal pHmetry may avert PPI use in 50%. In the era of caution regarding PPIs, early testing may provide assurance and justification.