Plasma cytokine levels in patients with chronic alcohol overconsumption: Relations to gut microbiota markers and clinical correlates.

BACKGROUND: Alcohol-related morbidity may involve changes in the gut microbiota and immune dysregulation. We have previously demonstrated alterations in gut microbiota composition and functions in patients with alcohol overconsumption, and now aimed to investigate possible associations between cytokine levels, gut microbiota, and clinical symptoms. METHODS: We included hospital inpatients with a history of chronic alcohol overconsumption. For comparison, we included control patients with a low alcohol intake. Cytokine levels (TGF-ß1, TNF-α, IL-10, IL-8, IL-6, IFN-γ, MCP-1, IL-1RA, IL-1ß, and IL-17) were determined using a customized V-plex assay. We then examined associations of cytokine levels with the abundance of Proteobacteria and Faecalibacterium, percentage of the short-chain fatty acid butyrate, psychiatric symptoms (Hospital Anxiety and Depression Scale), and biochemical liver variables. RESULTS: We included 28 patients with alcohol overconsumption (79% men), and 25 control patients (72% men). Patients with alcohol overconsumption had higher levels of IL-6 (p = 0.002), IFN-γ (p = 0.018), and MCP-1 (p = 0.006), and lower levels of TGF-ß1 (p = 0.017) compared with control patients. Inverse correlations were found between Proteobacteria abundance and TNF-α (Rs = -0.55, p = 0.02) and IL-8 (Rs = -0.58, p = 0.014), and between Faecalibacterium and MCP-1 levels (Rs = -0.56, p = 0.02) in the control patients, but not in patients with alcohol overconsumption. Patients with alcohol overconsumption reported more psychiatric symptoms, and these symptoms were inversely correlated with IL-10 levels. There were positive correlations between several of the assessed cytokines and biochemical liver variables, and negative correlations between cytokine levels and albumin. CONCLUSION: Patients with alcohol overconsumption had a cytokine profile suggestive of increased systemic inflammatory activity, with higher levels of pro-inflammatory cytokines (IL-6, IFN-γ, and MCP-1) and lower levels of anti-inflammatory cytokines (TGF-ß1). The findings may represent a link between alcohol use and alcohol-related morbidity.

Fecal Calprotectin: A Reliable Predictor of Mucosal Healing after Treatment for Active Ulcerative Colitis.

OBJECTIVES: Mucosal healing has become the new goal of treatment in ulcerative colitis. Fecal calprotectin has been demonstrated to differentiate between mucosal inflammation and mucosal healing. With this project, we investigated whether a reduction in f-calprotectin to <250 µg/g after medical treatment for active ulcerative colitis could predict mucosal healing. MATERIAL AND METHODS: After a baseline colonoscopy, 20 patients with active ulcerative colitis were followed with consecutive fecal calprotectin monthly until two measurements of fecal calprotectin < 250 µg/g or a maximum follow-up of 12 months. A flexible sigmoidoscopy was then performed and Mayo endoscopic subscore was used to evaluate degree of inflammation. Simple Clinical Colitis Activity Index was used for evaluation of clinical disease activity. RESULTS: A total of 16 patients achieved fecal calprotectin < 250 µg/g during follow-up, and all 16 patients had endoscopic mucosal healing (Mayo endoscopic subscore of ≤1) on the second endoscopy. The remaining four patients had persistently high f-calprotectin levels before the second endoscopy with Mayo endoscopic subscore corresponding to endoscopic mucosal healing in three out of four patients. CONCLUSIONS: Fecal calprotectin <250 µg/g after medical treatment for active ulcerative colitis is a reliable marker of endoscopic mucosal healing.

Clinical importance of faecal calprotectin variability in inflammatory bowel disease: intra-individual variability and standardisation of sampling procedure.

OBJECTIVE: Faecal (f-) calprotectin is a biomarker of intestinal inflammation. Previous studies have described intra-individual day-to-day variability of this biomarker in patients with inflammatory bowel disease (IBD) and morning samples have been suggested for standardisation purposes. With this project, we investigated if day-to-day variability differed from diurnal variability. Additionally, we evaluated a new extraction method for f-calprotectin analysis. METHODS: Fifty patients provided three faeces samples from morning - evening - morning on two consecutive days. Nineteen patients provided two faeces samples from the same bowel movement, one conventional spot sample, and one sample with a device for patient-administered sampling and extraction. RESULTS: The two morning samples differentiated between mucosal inflammation and mucosal healing with same level of agreement as the two samples from the same day (kappa 0.76), using an f-calprotectin cut-off level of 259 µg/g. Although large intra-individual variation in f-calprotectin values, there were no significant day-to-day (p = 0.096) or diurnal variation (p = 0.78). Used by laboratory technicians, the new extraction device correlated significantly with the conventional extraction method (p < 0.001), Spearman's rank correlation coefficient 0.95. Of the 19 patients testing patient administered extraction, two patients provided samples leading to considerably higher f-calprotectin levels than conventional sampling procedure. CONCLUSIONS: The reliability of f-calprotectin morning samples is equal to the reliability of samples from different bowel movements on the same day. The new extraction method is reliable when used by laboratory technicians, but larger studies are recommended to evaluate patient administered extraction.

Prediction of endoscopic disease activity in ulcerative colitis by two different assays for fecal calprotectin.

BACKGROUND AND AIMS: As mucosal healing is the goal of treatment in inflammatory bowel disease, defining a fecal [f-] calprotectin cut-off level for mucosal healing is crucial. Previous studies have presented different cut-off levels. The aim of this study was to investigate the ability of two f-calprotectin assays to differentiate mucosal healing from inflammation in ulcerative colitis. METHODS: Sixty-two patients with ulcerative colitis underwent colonoscopy for classification of mucosal inflammation [Mayo endoscopic subscore]. The patients also submitted a fecal sample for f-calprotectin analysis using two different assays, Calpro ELISA and Buhlmann ELISA. RESULTS: The two assays correlated significantly, with a Spearman rank correlation coefficient of 0.86. Both assays showed significantly different f-calprotectin levels in patients with a Mayo endoscopic subscore of 0 [mucosal healing] and 1­3 [inflamed mucosa] [p <0.001]. Using ROC curve analyses, we selected the best cut-off levels for both assays with responding sensitivity and specificity [presented with 95% confidence intervals]; Calpro ELISA cut-off 61 µg/g, sensitivity 84.1% [75.0­93.2%], specificity 83.3 % [74.0­92.6%], and Buhlmann ELISA cut-off 96 µg/g, sensitivity 90.9 % [83.7­98.1%], specificity 83.3 % [74.0­92.6%]. Defining mucosal healing as a Mayo endoscopic subscore ≤1, cut-off levels increased: Calpro ELISA cut-off 110 µg/g, sensitivity 80.0%[70­90%], specificity 66.6 % [54.9­78.3%]; and Buhlmann ELISA cut-off 259 µg/g, sensitivity 83.3 %[74­92.6%], specificity 71.9 % [60.7­83.1%]. CONCLUSIONS: The study demonstrates the need for assay specific cut-off levels in clinical practice,as the f-calprotectin cut-off level for endoscopic disease activity differed in these two assays.

Small intestinal malabsorption in chronic alcoholism determined by 13C-D-xylose breath test and microscopic examination of the duodenal mucosa.

OBJECTIVE: Diarrhea, weight loss and osteoporosis are prominent symptoms and clinical signs of alcoholism. One of several possible factors causing this clinical picture is small intestinal damage leading to malabsorption. The aim of this study was to prospectively evaluate small intestinal absorption in alcoholics using the (13)C-D-xylose breath test, and to relate the breath test results to morphological findings of the duodenal mucosa. MATERIAL AND METHODS: Sixteen alcoholics without liver failure or serious illness and presenting symptoms of dyspepsia, nausea or diarrhea were included. The (13)C-D-xylose breath test was performed in 14 of the included subjects. The breath tests of the alcoholics were compared to those of untreated coeliac patients and healthy subjects. Duodenal biopsy specimens were taken for assessment of epithelial morphology in 14 of the included subjects, using light- and electron microscopic techniques. RESULTS: Alcoholics had significantly reduced absorption of (13)C-D-xylose compared to healthy subjects. The time curve of (13)C-D-xylose absorption in the group of alcoholics was similar in appearance to that of untreated coeliac patients. Alcoholic patients had few light microscopic changes, but electron microscopic examination exposed morphological pathology in the majority of the patients, with a reduced surface area of microvilli as the main finding. CONCLUSIONS: Alcoholics have a pathological (13)C-D-xylose breath test with a time curve similar to that of untreated coeliac patients. This implies a condition of malabsorption. The morphological pathology found included a reduced absorptive area due to pathology of microvilli. These findings may explain our breath test results.