297 Materials and Methods Study population Patients aged 18-65 years were recruited from March 2016 until April 2017 at the outpatient IBD clinic of the University Medical Center Groningen (UMCG), Groningen, The Netherlands. All patients were diagnosed with Crohn’s disease (CD), according to clinical, endoscopic and histopathological features. All patients were Caucasians and lived in the Northern part of the Netherlands. Patients were divided into two groups based on fecal calprotectin levels as ameasure of inflammatory disease activity (using 100 μg calprotectin / gram feces as our currently used cutoff value for defining low and increased inflammatory disease activity). Standard demographic characteristics, including age, sex, body-mass index (BMI) and smoking history, were recorded, as well as clinical parameters specific for CD, such as the Harvey-Bradshaw index (HBI), Montreal disease classification (including Age at diagnosis (A), Location of disease (L) and the Behavior (B) of the disease) and current maintenance therapy. Disease location was recorded from the most recently performed endoscopic evaluation, which was completed within 12 months of intestinal permeability testing. CD patients who were treated with antibiotics 3 months prior to screening were not eligible for this study (to rule out an effect on the microbiota composition). Furthermore, CD patients with severe disease activity, as reflected by a HBI score > 12, or patients with an indication for remission-induction therapy, were also excluded from this study. Routine laboratory examinations were performed, including hemoglobin, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), leukocyte count, thrombocyte count and creatinine levels. Fecal calprotectin levels were quantified by enzyme-linked immunosorbent assays (ELISA) (BÜHLMANN Laboratories AG, Switzerland) as a routine measurement in the UMCG. Samples were obtained after patients gave written informed consent. This study has been approved by the Institutional Medical Ethical Review Board (in Dutch: Medisch Ethische Toetsingcommissie, METc) of the University Medical Center Groningen (UMC Groningen) (IRB no. 2014/291) and is in accordance with the principles of the Declaration of Helsinki (2013). 52Cr-EDTA solution The chromium-EDTA oral test solution consists of the active ingredients chromium(III)chloride (CrCl3.6H2O) and disodium edetate (C10H14N2Na2OH.2H2O), 70% sorbitol liquid crystalline and raspberry essence as sweeteners, 0.1 M NaOH for pH adjustment to reach a final pH between 4.5 – 5.5, methylparahydroxybenzoate as preservative and purified water (according to Ph. Eur. requirements) as solvent. The solution contained in total 400 μmol of 52Cr-EDTA (20 mmol/L). Patients were instructed to drink the 52Cr-EDTA solution (20 mL) together with a glass of water after an overnight fast. Subsequently, patients were asked to fast for an additional 2 hours. Urine was collected for 24 hours from the moment of ingestion of the 52Cr-EDTA test solution. Patients were also instructed to refrain from the intake of alcoholic beverages or non-steroidal antiinflammatory drugs (NSAID’s) two days prior to taking the 52Cr-EDTA solution and during the 24-h urine collection period. Full details on the preparation of the 52Cr-EDTA solution were described previously.42 52Cr-EDTA intestinal permeability in Crohn’s disease
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