VALIDATION OF MORPHOMETRIC ANALYSES OF SMALL-INTESTINAL BIOPSY READOUTS IN CELIAC DISEASE

Keliakian diagnostiikan kulmakivi on edelleen gluteenin aiheuttama limakalvovaurio ohutsuolessa. Vaurion aste arvioidaan yleensä käyttäen luokiteltuja asteikkoja, mutta nämä ovat usein liian epätarkkoja jättäen pieniä merkittäviä muutoksia huomiotta. Tämän vuoksi on tullut tarve kehittää validoituja jatkuvia muuttujia limakalvovaurion arviointiin. Arvioimme standardoidun toimintamenettelymme toimivuutta valikoidussa 93 potilaan ryhmässä. Näytteet sisälsivät eri asteisia limakalvovaurioita ja ne arvioitiin käyttäen jatkuvia muuttujia: nukkalisäke/kuopake-suhdetta ja valkosolutiheyttä. Kaksi akkreditoitua tutkijaa teki mittaukset sokkoutetusti. Tutkijan ja tutkijoiden välinen luotettavuus nukkalisäke/kuopake-suhteessa ja lymfosyyttitiheydessä tutkittiin Bland-Altmanin metodilla ja luokan sisäisellä korrelaatiolla. Molemmat tutkijat tunnistivat yhtenevästi palat, joita ei saanut toimintamenettelymme mukaan mitata. Bland-Altmanin-metodi ja luokan sisäinen korrelaatio osoittivat luotettavia tuloksia ja keskihajonnaksi saatiin tutkijan välisessä analyysissä 0,159 ja tutkijoiden välillä 0,227. Keskihajonnaksi CD3-positiivisessa valkosolutiheydessä saatiin tutkijan sisäisessä analyysissä 16,5% ja tutkijoiden välisessä analyysissä 17,1%. Käytettäessä standardoitua toimintamenettelyämme saavutettiin luotettavia ja toistettavia tuloksia jatkuville limakalvovaurion mittareille. Kliinisesti merkittäviksi muuttoksiksi asetettiin keskihajonnan mukaisesti nukkalisäke/kuopake-suhteessa 0.4 ja CD3-positiivisessa valkosolutiheydessä 30%

Validation of the X-ray microtomography in the assessment of duodenal morphometry and surface area in celiac disease

BackgroundDuodenal histology remains the diagnostic reference standard in celiac disease. However, traditional methods have suboptimal sensitivity and reproducibility for early mucosal changes and research purposes. We validated a recently introduced micro-CT imaging method for an accurate digital evaluation of duodenal histomorphometry and mucosal surface areas. MethodsEndoscopic biopsies from 58 individuals were utilized for the micro-CT imaging, selecting histological changes ranging from normal to severely damaged mucosa. The imaging protocol was optimized for practicability and resolution. The Bland-Altman method was applied to test intra- and interobserver variations in the blinded measurements. ResultsThe 3D micro-CT reconstructions enabled easy and precise digital cutting with optimal orientation and computer-assisted measurement of the surface area. Intraobserver analysis of morphological measurements showed a mean difference of 0.011 with limits of agreement (LA) from -0.397 to 0.375 and a standard deviation (SD) of 0.197. The corresponding figures for interobserver analysis were 0.080, from -0.719 to 0.537 and 0.320, respectively. The intraclass correlation coefficients (ICC) for the intraobserver and interobserver variations were 0.981 and 0.954, respectively. Intraobserver surface area analysis yielded a mean difference of 0.010, LA from -0.764 to 0.785 and an SD of 0.395, and an interobserver analysis mean difference of 0.028, LA from -0.642 to 0.698 and SD of 0.342. The respective ICCs for the intra- and interobserver variations were 0.963 and 0.972. ConclusionsMicro-CT showed excellent accuracy and reproducibility in the evaluation of mucosal morphometry and surface areas. The improved sensitivity for histological changes is a powerful tool for the diagnosis of celiac disease and for clinical and pharmacological studies.Peer reviewe

Prevalence and diagnostic outcomes of children with duodenal lesions and negative celiac serology

Background: Celiac disease diagnostics begin by measuring autoantibodies, which may fail to identify seronegative patients. Duodenal lesion in the absence of antibodies is scarcely studied, especially in children. Aims: To investigate the prevalence and diagnostic outcomes of children with seronegative duodenal lesion in two countries with different disease profiles. Methods: Medical data, including the results of histology and transglutaminase (tTGab) and endomysium (EmA) antibody measurements were collected from 1172 Finnish and 264 Romanian children with systematic duodenal sampling. Database of 509 Finnish children with celiac disease was examined to identify earlier seronegative patients. Results: Celiac disease was diagnosed in 307 Finnish and 83 Romanian children in the endoscopy cohorts. No seronegative patients were found among 899 celiac disease patients, although some were only tTGab or EmA positive. Non-celiac duodenal lesion was detected in eight Finnish and 32 Romanian children, their most common diagnoses being inflammatory bowel disease and infections, respectively. Six children with morphological lesion received no diagnosis. None of them developed celiac disease during a follow-up of 3-11 years. Conclusion: Pediatric seronegative celiac disease is exceptional in the era of modern autoantibodies. Other reasons for duodenal lesion should therefore be sought, bearing in mind possible differences across countries. (C) 2019 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.Peer reviewe

Celiac disease antibody levels reflect duodenal mucosal damage but not clinical symptoms

Objectives: This study aimed to investigate, in a real-world population, whether the histological and clinical phenotype differ at baseline and during follow-up in patients with high and low CD (celiac disease) antibody titers. Materials and methods: The study cohort consisted of 96 consecutive patients diagnosed to have CD during the years 2010–2018. The clinical parameters, symptoms and laboratory results were registered and histomorphometry was analyzed from the available duodenal biopsies taken during the primary and follow-up esophageal-gastricduodenoscopies. Patients having immunoglobulin A transglutaminase antibody (tTG-ab) levels above 70 U/mL were classified as high titer patients. Results: Measured by the villous-crypt ratio, the duodenal mucosa was more severely damaged in the high tTG-ab group than in the low tTG-group at baseline (n = 70, 0.61 ± 0.63 vs. 1.02 ± 0.87, p =.003) and during the follow-up when the patients were on gluten-free diet (n = 27, 1.80 ± 0.72 vs. 2.35 ± 0.64, p =.041). Interestingly, the high tTG-ab group members had fewer gastrointestinal symptoms at baseline than those in the low tTG-ab group (43% vs. 68%, p =.013) but lower vitamin D levels (68 ± 34 nmol/L vs. 88 ± 29 nmol/L, p =.034) and more often microcytosis (28% vs. 10%, p =.040). During the follow-up, these differences were no longer detected. Conclusions: At baseline, CD patients with high tTG-ab have more severe duodenum injury and signs of malabsorption but fewer symptoms. After gluten-free diet has been initiated, the mucosal healing in the high tTG-ab group is prolonged, but symptoms and signs of malabsorption recover equally in both groups.acceptedVersionPeer reviewe

X-ray microtomography is a novel method for accurate evaluation of small-bowel mucosal morphology and surface area

The often poorly orientated small-bowel mucosal biopsies taken for the diagnostics of celiac disease and other intestinal disorders are prone to misinterpretation. Furthermore, conventional histopathology has suboptimal sensitivity for early histopathological changes observed in short-term challenge studies. X-ray microtomography (micro-CT) is a promising new method for accurate imaging of human-derived biological samples. Here, we report that micro-CT could be utilized to create virtual reconstructions of endoscopically obtained intestinal biopsies. The formed digital 3D images enabled selection of always optimal cutting angles for accurate measurement of the mucosal damage and revealed diagnostic lesions in cases interpreted as normal with conventional histomorphometry. We also demonstrate that computer-assisted point cloud analysis can be used to calculate biologically meaningful surface areas of the biopsies in different stages of mucosal damage with excellent replicability and correlation with other disease parameters. We expect the improved diagnostic accuracy and capability to measure the surface areas to provide a powerful tool for the diagnostics of intestinal diseases and for future clinical and pharmaceutical trials.Peer reviewe

Gluten Challenge Induces Skin and Small Bowel Relapse in Long-Term Gluten-Free Diet-Treated Dermatitis Herpetiformis

Dermatitis herpetiformis (DH) is an extraintestinal manifestation of celiac disease causing an itchy, blistering rash. Granular IgA deposits in the skin are pathognomonic for DH, and the treatment of choice is a lifelong gluten-free diet (GFD). Preliminary evidence suggests that there are patients with DH who redevelop gluten tolerance after adherence to a GFD treatment. To evaluate this, we performed a 12-month gluten challenge with skin and small-bowel mucosal biopsy samples in 19 patients with DH who had adhered to a GFD for a mean of 23 years. Prechallenge biopsy was negative for skin IgA and transglutaminase 3 deposits in 16 patients (84%) and indicated normal villous height-to-crypt depth ratios in the small bowel mucosa in all 19 patients. The gluten challenge caused a relapse of the rash in 15 patients (79%) in a mean of 5.6 months; of these 15 patients, 13 had skin IgA and transglutaminase 3 deposits, and 12 had small-bowel villous atrophy. In addition, three patients without rash or immune deposits in the skin developed villous atrophy, whereas one patient persisted without any signs of relapse. In conclusion, 95% of the patients with DH were unable to tolerate gluten even after long-term adherence to a GFD. Therefore, lifelong GFD treatment remains justified in all patients with DH.Peer reviewe

Healthy and pro-inflammatory gut ecology plays a crucial role in the digestion and tolerance of a novel Gluten Friendly™ bread in celiac subjects : Randomized, double blind, placebo control in vivo study

Gluten Friendly™ (GF) is a new gluten achieved through a physicochemical process applied to wheat kernels. The goal of this research was to assess the in vivo effects of Gluten Friendly™ bread on celiac gut mucosa and microbiota. In a double-blind placebo-controlled intervention study, 48 celiac disease (CD) patients were randomized into 3 groups to eat 100 g of bread daily, containing different doses (0; 3 g; 6 g) of GF for 12 weeks. The small-bowel morphology (VH/CrD), intraepithelial densities of CD3+, celiac serology, MUC2, CB1, gut permeability, proinflammatory cytokines, gluten in stools, symptoms, and gut microbial composition were assessed. All 48 CD subjects experienced no symptoms. K-means analysis evidenced celiac subjects clustering around unknown parameters independent of GF dosage: K1 35%; K2 30%; K3 35%. VH/CrD significantly decreased in K1 and K2. VH/CrD did not correlate with IEL increase in K2. 33-mer was not detected in 47% and 73% of patients in both K1 and K2, respectively. VH/CrD and IEL did not change significantly and strongly correlated with the absence of 33-mer in K3. Inflammation and VH/CrD decrease are strongly related with the presence of proinflammatory species at the baseline. A boost in probiotic, butyrate-producing genera, is strongly related with GF tolerance at the end of the trial. Our research suggests that a healthy and proinflammatory ecology could play a crucial role in the digestion and tolerance of the new gluten molecule in celiac subjects. However, GF can be completely digested by gut microbiota of CD subjects and shapes it toward gut homeostasis by boosting healthy butyrate-producing populations. The clinical trial registry number is NCT03137862 (https://clinicaltrials.gov).publishedVersionPeer reviewe