[Chronic diarrhoea: when is it celiac disease and when not?] / Chronische Durchfälle: Wann ist es eine Zöliakie und wann nicht?

Patients who come to clinical consultation for chronic diarrhoea (i.e., diarrhoea lasting for more than four weeks) may suffer from a wide range of clinical conditions. The possible diagnoses range from a misunderstanding of what can be considered normal and what pathological in terms of daily bowel movements, to a severe malabsorption syndrome. Since the list of possible causes of chronic diarrhoea can be puzzling, the physician's approach needs to be systematic and structured in order to allow the correct diagnosis and treatment. This article proposes an algorithm for the diagnosis of chronic diarrhoea and discusses in detail the key clinical aspects of celiac disease, which is considered a paradigmatic disease as regards chronic malabsorptive diarrhoea.

A Look Into the Future: Are We Ready for an Approved Therapy in Celiac Disease?

As it appears that we are currently at the cusp of an era in which drugs that are new, re-purposed, or "supplements" will be introduced to the management of celiac disease, we need to reflect on whether the framework is set for celiac disease to be treated increasingly with pharmaceuticals as well as diet. This refers to reflecting on the rigor of current diagnostic practices; the limitations of the current standard of care, which is a gluten-free diet; and that we lack objective markers of disease severity. Investigating these issues will help us to identify gaps in technology and practices that could be critical for selecting patients with a well-defined need for an improved or alternative treatment. Both aspects, circumscribed limitations of the gluten-free diet and diagnostics helping to define celiac disease target groups, together with the guiding requirements by the responsible regulatory authorities, will contribute to defining the subgroups of patients with confirmed celiac disease eligible for distinct pharmacologic strategies. Because many patients with celiac disease are diagnosed in childhood, these aspects need to be differentially discussed for the pediatric setting. In this perspective, we aimed to describe these contextual issues and then looked ahead to the future. What might be the major challenges in celiac disease clinics in the coming years once drugs are an option alongside diet? And what will be the future objectives for researchers who further decipher the mucosal immunology of celiac disease? Speculating on the answers to these questions is as stimulating as it is fascinating to be part of this turning point.

Myenteric Plexus Immune Cell Infiltrations and Neurotransmitter Expression in Crohn's Disease and Ulcerative Colitis.

BACKGROUND AND AIMS: Pain is a cardinal symptom in inflammatory bowel disease [IBD]. An important structure in the transduction of pain signalling is the myenteric plexus [MP]. Nevertheless, IBD-associated infiltration of the MP by immune cells lacks in-depth characterisation. Herein, we decipher intra- and periganglionic immune cell infiltrations in Crohn´s disease [CD] and ulcerative colitis [UC] and provide a comparison with murine models of colitis. METHODS: Full wall specimens of surgical colon resections served to examine immune cell populations by either conventional immuno-histochemistry or immunofluorescence followed by either bright field or confocal microscopy. Results were compared with equivalent examinations in various murine models of intestinal inflammation. RESULTS: Whereas the MP morphology was not significantly altered in IBD, we identified intraganglionic IBD-specific B cell- and monocyte-dominant cell infiltrations in CD. In contrast, UC-MPs were infiltrated by CD8+ T cells and revealed a higher extent of ganglionic cell apoptosis. With regard to the murine models of intestinal inflammation, the chronic dextran sulphate sodium [DSS]-induced colitis model reflected CD [and to a lesser extent UC] best, as it also showed increased monocytic infiltration as well as a modest B cell and CD8+ T cell infiltration. CONCLUSIONS: In CD, MPs were infiltrated by B cells and monocytes. In UC, mostly CD8+ cytotoxic T cells were found. The chronic DSS-induced colitis in the mouse model reflected best the MP-immune cell infiltrations representative for IBD.

Guidelines for best practices in monitoring established coeliac disease in adult patients.

Coeliac disease (CeD) is an immunological disease triggered by the consumption of gluten contained in food in individuals with a genetic predisposition. Diagnosis is based on the presence of small bowel mucosal atrophy and circulating autoantibodies (anti-type 2 transglutaminase antibodies). After diagnosis, patients follow a strict, life-long gluten-free diet. Although the criteria for diagnosis of this disease are well defined, the monitoring phase has been studied less and there is a lack of specific guidelines for this phase. To develop a set of clinical guidelines for CeD monitoring, we followed the Grading of Recommendations Assessment, Development and Evaluation methodology. Statements and recommendations with the level of evidence were developed and approved by the working group, which comprised gastroenterologists, pathologists, dieticians and biostatisticians. The proposed guidelines, endorsed by the North American and European coeliac disease scientific societies, make recommendations for best practices in monitoring patients with CeD based on the available evidence. The evidence level is low for many topics, suggesting that further research in specific aspects of CeD would be valuable. In conclusion, the present guidelines support clinicians in improving CeD treatment and follow-up and highlight novel issues that should be considered in future studies.

Selective Uptake Into Inflamed Human Intestinal Tissue and Immune Cell Targeting by Wormlike Polymer Micelles.

Inflammatory bowel disease (IBD) has become a globally prevalent chronic disease with no causal therapeutic options. Targeted drug delivery systems with selectivity for inflamed areas in the gastrointestinal tract promise to reduce severe drug-related side effects. By creating three distinct nanostructures (vesicles, spherical, and wormlike micelles) from the same amphiphilic block copolymer poly(butyl acrylate)-block-poly(ethylene oxide) (PBA-b-PEO), the effect of nanoparticle shape on human mucosal penetration is systematically identified. An Ussing chamber technique is established to perform the ex vivo experiments on human colonic biopsies, demonstrating that the shape of polymeric nanostructures represents a rarely addressed key to tissue selectivity required for efficient IBD treatment. Wormlike micelles specifically enter inflamed mucosa from patients with IBD, but no significant uptake is observed in healthy tissue. Spheres (≈25 nm) and vesicles (≈120 nm) enter either both normal and inflamed tissue types or do not penetrate any tissue. According to quantitative image analysis, the wormlike nanoparticles localize mainly within immune cells, facilitating specific targeting, which is crucial for further increasing the efficacy of IBD treatment. These findings therefore demonstrate the untapped potential of wormlike nanoparticles not only to selectively target the inflamed human mucosa, but also to target key pro-inflammatory cells.

[What is established in the management of celiac disease?] / Was ist gesichert in der Therapie der Zöliakie?

Nowadays, celiac disease is well-established in internal medicine as an autoimmune disease induced by gluten as a trigger. Undoubtedly similarly well-established is the gluten-free diet. It is the only recognized therapy for celiac disease to date. However, this presents some pitfalls in its implementation, which will be discussed in the following review. In addition, current developments that have the potential to significantly change both diagnosis and treatment of celiac disease are discussed. On the one hand, such an outlook was chosen since colleagues want to be "ready" when such developments are integrated into daily clinical routine. On the other hand, the realization that the field of mucosal immunology is moving forward has the potential to lift the spirits of the reader.

A revised concept for deriving reference values for internal exposures to chemical substances and its application to population-representative biomonitoring data in German children and adolescents 2014-2017 (GerES V).

HBM reference values, in contrast to toxicologically derived values, are statistically derived values that provide information on the exposure of the population. The exceedance frequency (if applicable for individual population groups) is often a first assessment standard for the local exposure situation for municipalities. More than 25 years have passed since the German Human Biomonitoring Commission (HBMC) formulated the first recommendations for the derivation of population-based reference values (HBM reference values, RV95) for substance concentrations based on HBM studies. A fundamental revision is timely, for several reasons. There have been considerable advances in relevant statistical methods, which meant that previously time-consuming and inaccessible procedures and calculations are now widely available. Furthermore, not all steps for the derivation of HBM reference values were clearly elaborated in the first recommendations. With this revision we intended to achieve a rigorous standardization of the entire process of deriving HBM reference values, also to realise a higher degree of transparency. In accordance with established international practice, it is recommended to use the 95th percentile of the reference distribution as the HBM reference value. To this end, the empirical 95th percentile of a suitable sample should be rounded, ensuring that the rounded value is within the two-sided 95% confidence interval of the percentile. All estimates should be based on distribution-free methods, and the confidence interval should be estimated using a bootstrap approach, if possible, according to the BCa ("bias-corrected and accelerated bootstrap"). A minimum sample size of 80 observations is considered necessary. The entire procedure ensures that the derived HBM reference value is robust against at least two extreme values and can also be used for underlying mixed distributions. If it is known in advance that certain subgroups (different age groups, smokers, etc.) show differing internal exposures, it is recommended that group-specific HBM reference values should be derived. Especially when the sample sizes for individual subgroups are too small, individual datasets with potential outliers can be excluded in advance to homogenize the reference value population. In the second part, new HBM reference values based on data of the German Environmental Survey for Children and Adolescents (GerES V, 2014-2017) were derived in accordance with the revised recommendations. The GerES V is the most recent population-representative monitoring of human exposure to pollutants in Germany on children and adolescents aged 3-17 years (N = 2294). RV95 for GerES V are reported for four subgroups (males/females and 3-11/12-17 years) for 108 different substances including phthalates and alternative plasticisers, metals, organochlorine pesticides, polychlorinated biphenyls (PCB), per- and polyfluoroalkyl substances (PFAS), parabens, aprotic solvents, chlorophenols, polycyclic aromatic hydrocarbons (PAH) and UV filter, in total 135 biomarkers. Algorithms implemented in R were used for the statistics and the determination of the HBM reference values. To facilitate a quality control of the study data, the corresponding R source code is given, together with graphical representations of results. The HBM reference values listed in this article replace earlier RV95 values derived by the HBMC for children and adolescents from data of precedent GerES studies (e.g. published in Apel et al., 2017).

cCPE Fusion Proteins as Molecular Probes to Detect Claudins and Tight Junction Dysregulation in Gastrointestinal Cell Lines, Tissue Explants and Patient-Derived Organoids.

Claudins regulate paracellular permeability, contribute to epithelial polarization and are dysregulated during inflammation and carcinogenesis. Variants of the claudin-binding domain of Clostridium perfringens enterotoxin (cCPE) are highly sensitive protein ligands for generic detection of a broad spectrum of claudins. Here, we investigated the preferential binding of YFP- or GST-cCPE fusion proteins to non-junctional claudin molecules. Plate reader assays, flow cytometry and microscopy were used to assess the binding of YFP- or GST-cCPE to non-junctional claudins in multiple in vitro and ex vivo models of human and rat gastrointestinal epithelia and to monitor formation of a tight junction barrier. Furthermore, YFP-cCPE was used to probe expression, polar localization and dysregulation of claudins in patient-derived organoids generated from gastric dysplasia and gastric cancer. Live-cell imaging and immunocytochemistry revealed cell polarity and presence of tight junctions in glandular organoids (originating from intestinal-type gastric cancer and gastric dysplasia) and, in contrast, a disrupted diffusion barrier for granular organoids (originating from discohesive tumor areas). In sum, we report the use of cCPE fusion proteins as molecular probes to specifically and efficiently detect claudin expression, localization and tight junction dysregulation in cell lines, tissue explants and patient-derived organoids of the gastrointestinal tract.

Follow-Up of Celiac Disease in Adults: "When, What, Who, and Where".

For patients with celiac disease (CeD), a lifelong gluten-free diet is not a voluntary lifestyle choice-it is a necessity. The key end points in clinical follow-up are symptom resolution, the normalization of weight, prevention of overweight, seroconversion, and negation or minimization of increased long-term morbidity. For the latter, a surrogate endpoint is mucosal healing, which means the normalization of histology to Marsh 0-1. Ideally, celiac follow-up care includes a multidisciplinary approach, effective referral processes, improved access that leverages technological advances, and following guidelines with the identification of measurable quality indicators, ideally informed by evidence-based research. Face-to-face CeD care and telemedicine are considered the standards for this process, although published data are insufficient. Guidelines and statements on diagnosis are readily available. However, data are lacking on optimal clinic visit intervals and outcomes and quality indicators such as improvement of symptoms, function and quality of life, survival and disease control, and how to most effectively use healthcare resources. The results of future research should provide the basis for general recommendations for evidence-based standards of quality of care in CeD.

Analysis of Circulating Food Antigen-Specific T-Cells in Celiac Disease and Inflammatory Bowel Disease.

To demonstrate and analyze the specific T-cell response following barrier disruption and antigen translocation, circulating food antigen-specific effector T-cells isolated from peripheral blood were analyzed in patients suffering from celiac disease (CeD) as well as inflammatory bowel disease (IBD). We applied the antigen-reactive T-cell enrichment (ARTE) technique allowing for phenotypical and functional flow cytometric analyses of rare nutritional antigen-specific T-cells, including the celiac disease-causing gliadin (gluten). For CeD, patient groups, including treatment-refractory cases, differ significantly from healthy controls. Even symptom-free patients on a gluten-free diet were distinguishable from healthy controls, without being previously challenged with gluten. Moreover, frequency and phenotype of nutritional antigen-specific T-cells of IBD patients directly correlated to the presence of small intestinal inflammation. Specifically, the frequency of antigen specific T-cells as well as pro-inflammatory cytokines was increased in patients with active CeD or Crohn's disease, respectively. These results suggest active small intestinal inflammation as key for the development of a peripheral food antigen-specific T-cell response in Crohn's disease and celiac disease.

Proteolytic Activity of the Paracaspase MALT1 Is Involved in Epithelial Restitution and Mucosal Healing.

The paracaspase MALT1 is a crucial regulator of immune responses in various cellular contexts. Recently, there is increasing evidence suggesting that MALT1 might represent a novel key player in mucosal inflammation. However, the molecular mechanisms underlying this process and the targeted cell population remain unclear. In this study, we investigate the role of MALT1 proteolytic activity in the context of mucosal inflammation. We demonstrate a significant enrichment of MALT1 gene and protein expression in colonic epithelial cells of UC patients, as well as in the context of experimental colitis. Mechanistically we demonstrate that MALT1 protease function inhibits ferroptosis, a form of iron-dependent cell death, upstream of NF-κB signaling, which can promote inflammation and tissue damage in IBD. We further show that MALT1 activity contributes to STAT3 signaling, which is essential for the regeneration of the intestinal epithelium after injury. In summary, our data strongly suggests that the protease function of MALT1 plays a critical role in the regulation of immune and inflammatory responses, as well as mucosal healing. Understanding the mechanisms by which MALT1 protease function regulates these processes may offer novel therapeutic targets for the treatment of IBD and other inflammatory diseases.

Uptake of Tropheryma whipplei by Intestinal Epithelia.

BACKGROUND: Tropheryma whipplei (TW) can cause different pathologies, e.g., Whipple's disease and transient gastroenteritis. The mechanism by which the bacteria pass the intestinal epithelial barrier, and the mechanism of TW-induced gastroenteritis are currently unknown. METHODS: Using ex vivo disease models comprising human duodenal mucosa exposed to TW in Ussing chambers, various intestinal epithelial cell (IEC) cultures exposed to TW and a macrophage/IEC coculture model served to characterize endocytic uptake mechanisms and barrier function. RESULTS: TW exposed ex vivo to human small intestinal mucosae is capable of autonomously entering IECs, thereby invading the mucosa. Using dominant-negative mutants, TW uptake was shown to be dynamin- and caveolin-dependent but independent of clathrin-mediated endocytosis. Complementary inhibitor experiments suggested a role for the activation of the Ras/Rac1 pathway and actin polymerization. TW-invaded IECs underwent apoptosis, thereby causing an epithelial barrier defect, and were subsequently subject to phagocytosis by macrophages. CONCLUSIONS: TW enters epithelia via an actin-, dynamin-, caveolin-, and Ras-Rac1-dependent endocytosis mechanism and consecutively causes IEC apoptosis primarily in IECs invaded by multiple TW bacteria. This results in a barrier leak. Moreover, we propose that TW-packed IECs can be subject to phagocytic uptake by macrophages, thereby opening a potential entry point of TW into intestinal macrophages.

Portal hypertension in common variable immunodeficiency disorders - a single center analysis on clinical and immunological parameter in 196 patients.

Background: Liver manifestations and in particular portal hypertension (PH) contribute significantly to morbidity and mortality of patients with common variable immunodeficiency disorders (CVID). Screening strategies and early detection are limited due to the lack of specific diagnostic tools. Methods: We evaluated clinical, immunological, histological, and imaging parameters in CVID patients with clinical manifestation of portal hypertension (CVID+PH). Results: Portal hypertension was present in 5.6% of CVID patients and was associated with high clinical burden and increased mortality (18%). Longitudinal data on clinical and immunological parameters in patients before and during clinically manifest portal hypertension revealed a growing splenomegaly and increasing gamma-glutamyl transferase (GGT) and soluble interleukin 2 receptor (SIL-2R) levels with decreasing platelets over time. While ultrasound of the liver failed to detect signs of portal hypertension in most affected patients, transient elastography was elevated in all patients. All CVID+PH patients had reduced naïve CD45RA+CD4+ T-cells (mean of 6,2%). The frequency of severe B-lymphocytopenia (Euroclass B-) was higher in CVID+PH patients. The main histological findings included lymphocytic infiltration, nodular regenerative hyperplasia-like changes (NRH-LC), and porto(-septal) fibrosis. Conclusion: CVID patients with lower naïve CD45RA+CD4+ T-cells or severely reduced B-cells might be at higher risk for portal hypertension. The combination of biochemical (increasing sIL-2R, GGT, and decreasing platelets) and imaging parameters (increasing splenomegaly) should raise suspicion of the beginning of portal hypertension.

S2k Guideline for the diagnosis and treatment of mucous membrane pemphigoid.

Mucous membrane pemphigoid (MMP) is a pemphigoid disease with predominant mucous membrane involvement. It mainly affects the mucous membranes of the mouth, eyes, nose and pharynx, but also the larynx, trachea, esophagus, genital and perianal regions. The manifestation of the disease covers a wide spectrum from gingival erythema and single oral lesions to severe tracheal strictures that obstruct breathing and conjunctival scarring with marked visual impairment and, not infrequently, blindness. In addition to a clinical picture of predominant mucosal involvement, diagnosis is based on direct immunofluorescence of a peri-lesional biopsy and serology. The main target antigen is BP180 (collagen XVII), and reactivity with laminin 332 is associated with malignancy in approximately 25 % of MMP patients. The treatment of MMP is challenging. On the one hand, due to the involvement of different mucous membranes, good interdisciplinary cooperation is required; on the other hand, due to the rarity of the disease, no randomized controlled clinical trials are available. The aim of this guideline is to present the clinical picture, including severity and scoring systems, and to give guidance for diagnosing and treating this complex disease. In MMP, interdisciplinary cooperation plays an essential role as well as the prompt diagnosis and initiation of adequate therapy in order to avoid irreversible damage to the mucous membranes with serious complications.

Successful treatment of severe COVID-19 pneumonia, a case series with simultaneous interleukin-1 and interleukin-6 blockade with 1-month follow-up.

Interleukin (IL)-6 and IL-1 blockade showed beneficial results in patients with severe COVID-19 pneumonia and evidence of cytokine release at the early disease stage. Here, we report outcomes of open-label therapy with a combination of blocking IL-6 with tocilizumab 8 mg/kg up to 800 mg and IL-1 receptor antagonist anakinra 100-300 mg over 3-5 days. Thirty-one adult patients with severe COVID-19 pneumonia and signs of cytokine release, mean age 54 (30-79) years, 5 female, 26 male, and mean symptom duration 6 (3-10) days were treated. Patients with more than 10 days of symptoms, evidence of bacterial infection/elevated procalcitonin and other severe lung diseases were excluded. Computed tomography (CT) scans of the lung were performed initially and after 1 month; inflammatory activity was assessed on a scale 0-25. Twenty-five patients survived without intubation and mechanical lung ventilation, two patients died. C-reactive protein decreased in 19/31 patients to normal ranges. The mean activity CT score decreased from 14 (8-20) to 6 (0-16, n = 16). In conclusion, most of our patients recovered fast and sustained, indicating that early interruption of cytokine release might be very effective in preventing patients from mechanical ventilation, death, and long-term damage.

The combination of clinical parameters and immunophenotyping of intraepithelial lymphocytes allows to assess disease severity in refractory celiac disease.

BACKGROUND: Flow cytometry of intestinal lymphocytes is discussed to be a stronger predictor of enteropathy-associated T-cell lymphoma development in refractory celiac disease than T-cell clonality analysis. AIMS: To investigate possible associations between clinical characteristics of refractory celiac disease patients and aberrant intraepithelial lymphocytes and to evaluate the accuracy of immunophenotyping for the identification of high-risk refractory celiac disease. METHODS: Flow cytometry of isolated lymphocytes from duodenal biopsies of controls and celiac disease patients was performed and results were compared to clinical data. RESULTS: Flow cytometry analysis was performed on 42 controls, 37 non-complicated celiac disease and 30 refractory celiac disease cases with or without T-cell receptor clonality. Elevated aberrant intraepithelial lymphocyte counts were significantly associated with severe malabsorption. A 15% cut-off (aberrant lymphocytes among all lymphocytes) had the best discriminatory ability to identify high-risk patients. However, this technique failed to identify some high-risk cases (sensitivity 63%, specificity 100%). The severity of malabsorption was added to the criteria for high-risk refractory celiac disease, improving the correct patients' allocation (sensitivity 100%, specificity 96%). CONCLUSION: Immunophenotyping of aberrant intraepithelial lymphocytes is a good predictor for high-risk refractory celiac disease. Furthermore, adding the evaluation of malabsorption to the diagnostic assessment of refractory celiac disease optimizes accuracy.