Fecal Markers Of Inflammation Research Articles

Noninvasive Markers of Inflammation and Protein Loss Augment Diagnosis of Pediatric Celiac Disease.

Circulating tissue transglutaminase immunoglobulin A concentration is a sensitive and specific indicator of celiac disease, but discrepancies between serologic and histologic findings occur. We hypothesized that fecal markers of inflammation and protein loss would be greater in patients with untreated celiac disease than in healthy controls. Our study aims to evaluate multiple fecal and plasma markers in celiac disease and correlate these findings with serologic and histologic findings as noninvasive means of evaluating disease activity. Participants with positive celiac serologies and controls with negative celiac serologies were prospectively enrolled before upper endoscopy. Blood, stool, and duodenal biopsies were collected. Concentrations of fecal lipocalin-2, calprotectin, and alpha-1-antitrypsin and plasma lipocalin-2 were determined. Biopsies underwent modified Marsh scoring. Significance was tested between cases and controls, modified Marsh score and tissue transglutaminase immunoglobulin A concentration. Lipocalin-2 was significantly elevated in the stool ( P = 0.006) but not the plasma of participants with positive celiac serologies. There was no significant difference in fecal calprotectin or alpha-1 antitrypsin between participants with positive celiac serologies and controls. Fecal alpha-1 antitrypsin >100 mg/dL was specific, but not sensitive for biopsy-proven celiac disease. Lipocalin-2 is elevated in the stool but not the plasma of patients with celiac disease suggesting a role of local inflammatory response. Calprotectin was not a useful marker in the diagnosis of celiac disease. While random fecal alpha-1 antitrypsin was not significantly elevated in cases compared with controls, an elevation of greater than 100 mg/dL was 90% specific for biopsy-proven celiac disease.

STANDARDIZATION ISSUES FOR FECAL CALPROTECTIN MEASUREMENT IN INFLAMMATORY BOWEL DISEASE

Fecal calprotectin (FC) is a non-invasive marker of intestinal inflammation that is often used to guide therapeutic decisions in patients with inflammatory bowel disease (IBD). Each step in FC measurement can influence the results, leading to misinterpretation and potentially affecting the management of patients with IBD. To date, FC measurements are highly heterogeneous, and none of the existing methods is generally accepted as a standard. <b>Aim:</b> to provide a clear position on the preanalytical and analytical steps of FC measurement and to minimize the risk of misinterpretation. <b>Materials and methods.</b> A systematic search of the scientific literature was carried out in the PubMed/MEDLINE, EMBASE and Cochrane databases until June 2021 to identify all studies reporting data on FC measurement (for the key phrases "fecal markers of inflammation", "calprotectin", "inflammatory bowel diseases"). <b>Results.</b> Based on available data, quantitative tests should be preferred to measure FC; FC measurements should always be performed using the same method, and factors affecting his levels should be taken into account when interpreting results. <b>Conclusion.</b> FC concentrations determine treatment decisions by physicians. Although FC is playing an increasingly important role in the management of IBD patients, large multicenter studies should be conducted to determine the reproducibility and to confirm the diagnostic accuracy of the available FC tests.

Elexacaftor/tezacaftor/ivacaftor and gastrointestinal outcomes in cystic fibrosis: Report of promise-GI

BackgroundElexacaftor/tezacaftor/ivacaftor (ETI) improves pulmonary disease in people with cystic fibrosis (PwCF), but its effect on gastrointestinal symptoms, which also affect quality of life, is not clear. MethodsPROMISE is a 56-center prospective, observational study of ETI in PwCF >12 years and at least one F508del allele. Gastrointestinal symptoms, evaluated by validated questionnaires: Patient Assessment of Upper Gastrointestinal Disorders-Symptom (PAGI-SYM), Patient Assessment of Constipation-Symptom (PAC-SYM), Patient Assessment of Constipation-Quality of Life (PAC-QOL)), fecal calprotectin, steatocrit and elastase-1 were measured before and 6 months after ETI initiation. Mean difference and 95% confidence intervals were obtained from linear regression with adjustment for age and sex. Results438 participants fully completed at least 1 questionnaire. Mean (SD) for baseline PAGI-SYM, PAC-SYM, and PAC-QOL total scores were 0.56 (0.59), 0.47 (0.45), and 0.69 (0.53) out of maximum 5, 4, and 5, respectively (higher score indicates greater severity). Corresponding age- and sex-adjusted 6 months mean changes (95% CI) in total scores were -0.15 (-0.21, -0.09) for PAGI-SYM, -0.14 (-0.19, -0.09) for PAC-SYM, and -0.15 (-0.21, -0.10) for PAC-QOL. While statistically significant, changes were small and unlikely to be of clinical importance. Fecal calprotectin showed a change (95% CI) from baseline of -66.2 µg/g (-86.1, -46.2) at 6 months, while fecal elastase and steatocrit did not meaningfully change. ConclusionsAfter 6 months of ETI, fecal markers of inflammation decreased. Gastrointestinal symptoms improved, but the effect size was small. Pancreatic insufficiency did not improve.

Значення дослідження фекального кальпротектину в діагностиці хронічних запальних захворювань кишечника у дітей

Актуальність. Запальні захворювання кишечника є однією з найбільш складних проблем в дитячій гастроентерології, що мають не тільки медичне, але й соціальне значення. Останнім часом велика увага приділяється неінвазивним методам діагностики, серед яких при хронічних запальних захворюваннях кишечника розглядають фекальні маркери запалення. Метою роботи було вивчення діагностичної цінності фекального кальпротектину при хронічних захворюваннях кишечника у дітей. Матеріали та методи. Обстежено 19 пацієнтів від 3 до 17 років з хронічними запальними захворюваннями кишечника в активній фазі (виразковим колітом і хронічним неспецифічним невиразковим колітом). Групу порівняння становили 12 дітей із синдромом подразненого кишечника. Крім стандартних методів обстеження, у хворих вивчали вміст кальпротектину в зразках калу з використанням моноклональних антитіл. Результати. Згідно з даними нашого дослідження, у пацієнтів з хронічними запальними захворюваннями кишечника середній рівень кальпротектину в калі перевищував нормальні показники в 3 рази, причому в усіх обстежених хворих в активній фазі хронічних запальних захворювань кишечника кальпротектин калу був підвищений. Середні значення фекального кальпротектину були вірогідно вище у хворих з виразковим колітом порівняно з пацієнтами з неспецифічним невиразковим колітом. При функціональному захворюванні — синдромі подразненого кишечника — рівні фекального кальпротектину не виходили за межі нормальних значень у жодного з пацієнтів. Висновки. Вивчення фекального кальпротектину при хронічних запальних захворюваннях кишечника у дітей має високу діагностичну цінність як скринінг для виключення органічної природи захворювання и визначення показань для проведення інструментальних методів діагностики. Впровадження неінвазивної методики вивчення кальпротектину в зразках калу дозволить моніторувати перебіг хронічних запальних захворювань кишечника у дітей в динаміці, оцінювати результати терапії, а також скоротити кількість інвазивних методів обстеження.

Examining the Gastrointestinal and Immunomodulatory Effects of the Novel Probiotic Bacillus subtilis DE111.

Probiotics make up a large and growing segment of the commercial market of dietary supplements and are touted as offering a variety of human health benefits. Some of the purported positive impacts of probiotics include, but are not limited to, stabilization of the gut microbiota, prevention of gastrointestinal disorders and modulation of the host immune system. Current research suggests that the immunomodulatory effects of probiotics are strain-specific and vary in mode of action. Here, we examined the immunomodulatory properties of Bacillus subtilis strain DE111 in a healthy human population. In a pilot randomized, double blind, placebo-controlled four-week intervention, we examined peripheral blood mononuclear cells (PBMCs) at basal levels pre- and post-intervention, as well as in response to stimulation with bacterial lipopolysaccharide (LPS). We observed an increase in anti-inflammatory immune cell populations in response to ex vivo LPS stimulation of PBMCs in the DE111 intervention group. Overall perceived gastrointestinal health, microbiota, and circulating and fecal markers of inflammation (Il-6, sIgA) and gut barrier function (plasma zonulin) were largely unaffected by DE111 intervention, although the study may have been underpowered to detect these differences. These pilot data provide information and justification to conduct an appropriately powered clinical study to further examine the immunomodulatory potential of B. subtilis DE111 in human populations.

Intervention and Mechanisms of Alanyl-glutamine for Inflammation, Nutrition, and Enteropathy: A Randomized Controlled Trial.

Objective:Determine the minimum dosage of alanyl-glutamine (Ala-Gln) required to improve gut integrity and growth in children at risk of environmental enteropathy (EE).Methods:This was a double-blinded randomized placebo-controlled dose-response trial. We enrolled 140 children residing in a low-income community in Fortaleza, Brazil. Participants were 2 to 60 months old and had weight-for-age (WAZ), height-for-age (HAZ), or weight-for-height (WHZ) z-scores less than −1. We randomized children to 10 days of nutritional supplementation: Ala-Gln at 3 g/day, Ala-Gln at 6 g/day, Ala-Gln at 12 g/day, or an isonitrogenous dose of glycine (Gly) placebo at 12.5 g/day. Our primary outcome was urinary lactulose-mannitol excretion testing. Secondary outcomes were anthropometry, fecal markers of inflammation, urine metabolic profiles, and malabsorption (spot fecal energy).Results:Of 140 children, 103 completed 120 days of follow-up (24% dropout). In the group receiving the highest dose of Ala-Gln, we detected a modest improvement in urinary lactulose excretion from 0.19% on day 1 to 0.17% on day 10 (P=0.05). We observed significant but transient improvements in WHZ at day 10 in 2 Ala-Gln groups, and in WHZ and WAZ in all Ala-Gln groups at day 30. We detected no effects on fecal inflammatory markers, diarrheal morbidity, or urine metabolic profiles; but did observe modest reductions in fecal energy and fecal lactoferrin in participants receiving Ala-Gln.Conclusions:Intermediate dose Ala-Gln promotes short-term improvement in gut integrity and ponderal growth in children at risk of EE. Lower doses produced improvements in ponderal growth in the absence of enhanced gut integrity.

Fecal Markers of Inflammation and Disease Activity in Pediatric Crohn Disease: Results from the ImageKids Study.

Noninvasive and accurate methods to monitor inflammatory bowel disease are required. As a planned ancillary study of the prospective ImageKids cohort, we aimed to assess the performance of fecal calprotectin (FC) with comparison to 3 fecal inflammatory markers; S100A12 (FA12), tumor pyruvate kinase isoenzyme type M2 (FM2PK) and fecal osteoprotegerin (FOPG) as indicators of a number of disease characteristics. The ImageKids study was a multicenter study designed to develop 2 magnetic resonance enterography-based measures for children with Crohn disease (6-18 years old). All patients underwent magnetic resonance enterography, a complete ileocolonoscopic evaluation and provided a fecal sample. Fecal samples were assay for FC, FA12, FM2PK, and FOPG by ELISA. One-hundred fifty-six children provided 190 fecal samples. Median (interquartile range) for fecal makers were FC, 602 (181-1185) μg/g; FA12, 21 (3-109) μg/g; FM2PK, 16 (2-20) U/mL; and FOPG, 125 (125-312) μg/g. All markers correlated with simple endoscopic severity index for Crohn disease and with other constructs of disease activity, but FC had the highest overall correlations. FA12, however, predicted mucosal healing with significantly higher specificity (87% vs 70%, P = 0.004) and equivalent sensitivity (91% vs 90%) compared to FC. This study has confirmed that FC is useful, and overall best, marker to monitor mucosal inflammation in inflammatory bowel disease. FA12, however, appears to be a more suitable maker for prediction of mucosal healing in children.

Monitoring of Vedolizumab Infusion Therapy (MOVE-IT) Response With Fecal Inflammation Markers, Ultrasound, and Trough Serum Level in Patients With Ulcerative Colitis: Protocol for a Multicentric, Prospective, Noninterventional Study.

BackgroundVedolizumab has been shown to induce clinical remission in patients with active ulcerative colitis. Treatment with anti-integrin vedolizumab leads to clinical remission in 16.9% and clinical response in 47.1% of cases after 6 weeks. However, in clinical practice, no decision to discontinue or continue vedolizumab therapy is made until 14 weeks at the earliest.ObjectiveThe aim of this study is to develop an algorithm for optimizing vedolizumab administration in patients with moderate-to-severe ulcerative colitis by calculating the probability of clinical response at week 14, on the basis of the data from week 6.MethodsThis is a prospective, single-arm, multicentric, noninterventional, observational study with no interim analyses and a sample size of 35 evaluable patients.ResultsThe enrollment started in August 2018 and was still open at the date of submission. The study is expected to complete in September 2020.ConclusionsThe early identification of patients who are responding to an integrin antibody is therapeutically beneficial. At the same time, patients who are not responding can be identified earlier. The development of a therapeutic algorithm for identifying patients as responders or nonresponders can thus help prescribing physicians avoid ineffective treatments and stop these very early.

2235. Fecal Biomarkers for Clostridioides difficile Infection in Cancer Patients

BackgroundThe diagnosis of C. difficile infection (CDI) relies on using a nucleic acid amplification test (NAAT) followed by confirmatory toxin enzyme immunoassay (EIA). This study examined the utility of fecal biomarkers and C. difficile bacterial quantity (BQ) in differentiating patients with true infection (NAAT+/EIA+) from patients with colonization (NAAT+/EIA-) in the context of C. difficile ribotypes.MethodsWe studied 136 patients with diarrhea and CDI identified by NAAT in stools for which a confirmatory toxin A, B, EIA was performed. Fecal IL-8, IL-1β, calprotectin, and lactoferrin were studied by ELISA. C. difficile BQ was determined by 16S rRNA qPCR. Data were stratified according to cancer diagnosis into three groups [hematologic (H) n = 43, solid tumor (ST) n = 62, or stem cell transplant (SCT) n = 31].ResultsStools were EIA+ in 36/136 (26%) of the cohort. Although ST patients had a higher Charlson co-morbidity index when compared with the other two groups (P < 0.002), demographic characteristics and symptoms at the time of presentation were similar between groups regardless of EIA status. Most common ribotypes identified included F106 and F014-020. Ribotype distribution differed according to oncologic diagnosis as determined by the Shannon diversity index. There were fewer distinct C. difficile ribotypes in the SCT (n = 8) vs. ST (n = 15) and H (n = 15) groups (P < 0.001 and P < 0.002, respectively). BQ were higher in EIA+ than EIA- across all strata (log of BQ/mg 2.38 ± 1.49 vs. 0.92 ± 1.28, P > 0.001). Similarly, higher levels of fecal IL-8 (1.72 ± 1.9 vs. 0.83 ± 1.6 ng/mL), IL-1β (3.74 ± 13.7 vs.. 1.21 ± 4.6) and calprotectin (14.9 ± 27 vs. 6 ±1.8 ug/mL) levels were seen in EIA+ patients. While IL-8, IL-1β, and calprotectin were increased in EIA+ ST and H, no differences were seen in the SCT group. A sensitivity analysis using ROC curves, revealed that BQ resulted in a greater area under the curve than fecal markers of inflammation (A = 0.77, P < 0.001, 95% CI [0.67–0.86]).ConclusionIn this study in cancer and immunocompromised patients, C. difficile bacterial burden regardless of infecting ribotype and fecal cytokines showed to be a helpful assay in distinguishing true CDI from colonization. Disclosures All authors: No reported disclosures.

2380. Fecal Collinsella Abundance is Negatively Associated with Toxin A/B Production in Cancer Patients with Clostridioides difficile

BackgroundThe detection of C. difficile (CDI) by nucleic acid amplification test (NAAT) with negative toxin enzyme immunoassay (EIA-) is difficult to interpret in cancer patients. Markers that differentiate true infection from colonization, and are associated with clinical outcomes are needed. We hypothesized that the microbiome composition and inflammatory fecal markers in EIA- patients differed from those who are EIA+ and were associated with disease severity and recurrence.MethodsWe studied the fecal microbiome composition (16s rRNA, V3) of 147 cancer patients with CDI diagnosed by a two-step testing algorithm. Clinical data, CDI bacterial quantity (BQ) by qPCR and markers of intestinal inflammation (calprotectin, lactoferrin, IL-1β and IL-8) were analyzed. Data were stratified according to cancer type [hematologic (H) n = 49, solid tumor (ST) n = 66, or stem cell transplant (SCT) n = 32].ResultsDemographic characteristics and symptoms were similar between the three groups. At baseline, species diversity by Shannon index was similar in all three groups regardless of EIA detection and did not correlate with clinical presentation, response to therapy or recurrence. Microbiome composition did not correlate with inflammatory response except in H in whom a higher diversity correlated with increased IL-8 (P = 0.021) and calprotectin (P = 0.01) levels. At the genus level across all strata and when compared with EIA- cases, EIA+ cases presented with a higher abundance of Peptoclostridium (P = 0.0008) which correlated with CDI BQ qPCR (log of BQ/mg 2.38 ± 1.49 vs 0.92 ± 1.28, P < 0.001). In contrast, EIA- cases had a higher abundance of Collinsella (P = 0.001). SCT patients carried fewer Peptoclostridium when compared with other groups, whereas all three patient groups carried similar amounts of Collinsella. The relative abundance of Peptoclostridium and Collinsella was not associated with response to therapy, or fecal markers of inflammation. Principal component analysis did not demonstrate differences between the three groups studied.ConclusionIn this study, the presence of Collinsella, a known butyrate and bile salt hydrolase producer, was associated with the lack of CDI toxin A/B production. Loss of Collinsella may represent a novel risk factor for active CDI. Disclosures All authors: No reported disclosures.

Enriched Environmental Conditions Modify the Gut Microbiome Composition and Fecal Markers of Inflammation in Parkinson’s Disease

Recent findings suggest an implication of the gut microbiome in Parkinson’s disease (PD) patients. PD onset and progression has also been linked with various environmental factors such as physical activity, exposure to pesticides, head injury, nicotine, and dietary factors. In this study, we used a mouse model, overexpressing the complete human SNCA gene (SNCA-TG mice) modeling familial and sporadic forms of PD to study whether environmental conditions such as standard vs. enriched environment changes the gut microbiome and influences disease progression. We performed 16S rRNA DNA sequencing on fecal samples for microbiome analysis and studied fecal inflammatory calprotectin from the colon of control and SNCA-TG mice kept under standard environment (SE) and enriched environment (EE) conditions. The overall composition of the gut microbiota was not changed in SNCA-TG mice compared with WT in EE with respect to SE. However, individual gut bacteria at genus level such as Lactobacillus sp. was a significant changed in the SNCA-TG mice. EE significantly reduced colon fecal inflammatory calprotectin protein in WT and SNCA-TG EE compared to SE. Moreover, EE reduces the pro-inflammatory cytokines in the feces and inflammation inducing genes in the colon. Our data suggest that an enriched social environment has a positive effect on the induction of SNCA mediated inflammation in the intestine and by modulating anti-inflammatory gut bacteria.

41-Year-Old Man With Fever and Bloody Diarrhea

41-Year-Old Man With Fever and Bloody Diarrhea

Differentiation of Sensitivity Diagnosis of C. difficile Infection Using Fecal Inflammation Markers Combined With C. difficile Toxins Test

Differentiation of Sensitivity Diagnosis of C. difficile Infection Using Fecal Inflammation Markers Combined With C. difficile Toxins Test

Fecal markers of intestinal inflammation and intestinal permeability are elevated in Parkinson's disease

Background/ObjectiveIntestinal inflammation and increased intestinal permeability (both possibly fueled by dysbiosis) have been suggested to be implicated in the multifactorial pathogenesis of Parkinson's disease (PD). The objective of the current study was to investigate whether fecal markers of inflammation and impaired intestinal barrier function corroborate this pathogenic aspect of PD. MethodsIn a case-control study, we quantitatively analyzed established fecal markers of intestinal inflammation (calprotectin and lactoferrin) and fecal markers of intestinal permeability (alpha-1-antitrypsin and zonulin) in PD patients (n = 34) and controls (n = 28, group-matched for age) by enzyme-linked immunosorbent assay. The study design controlled for potential confounding factors. ResultsCalprotectin, a fecal marker of intestinal inflammation, and two fecal markers of increased intestinal permeability (alpha-1-antitrypsin and zonulin) were significantly elevated in PD patients compared to age-matched controls. Lactoferrin, as a second fecal marker of intestinal inflammation, showed a non-significant trend towards elevated concentrations in PD patients. None of the four fecal markers correlated with disease severity, PD subtype, dopaminergic therapy, or presence of constipation. ConclusionsFecal markers reflecting intestinal inflammation and increased intestinal permeability have been primarily investigated in inflammatory bowel disease so far. Our data indicate that calprotectin, alpha-1-antitrypsin and zonulin could be useful non-invasive markers in PD as well. Even though these markers are not disease-specific, they corroborate the hypothesis of an intestinal inflammation as contributing factor in the pathogenesis of PD. Further investigations are needed to determine whether calprotectin, alpha-1-antitrypsin and zonulin can be used to define PD subgroups and to monitor the effect of interventions in PD.

Fecal markers of inflammation, protein loss, and microbial changes in dogs with the acute hemorrhagic diarrhea syndrome (AHDS).

Idiopathic acute hemorrhagic diarrhea syndrome (AHDS) is characterized by acute onset of bloody diarrhea, severe dehydration, and increased vascular and intestinal mucosal permeability. Markers of gastrointestinal inflammation, protein loss, and changes in the intestinal microbiota have not been studied extensively in dogs with AHDS. For 3 consecutive days, feces were collected from dogs with AHDS, and assayed for calprotectin and S100A12 (both markers of inflammation), α1 -proteinase inhibitor (a marker of gastrointestinal protein loss), and the presence of selected species of bacteria. Concentrations of all assayed markers were significantly greater than the institutional reference intervals at the time of presentation, and all decreased significantly by Day 3 of treatment. Abundances of selected bacterial groups (Ruminococcaceae, Faecalibacterium spp., Bifidobacterium spp., and Proteobacteria) at the time of diagnosis were consistent with an intestinal bacterial dysbiosis. No differences in the abundance of bacterial groups over time was seen, except for a mild but significant decrease in Ruminococcaceae at Day 3. These results suggest that canine AHDS is associated with a significant but transient gastrointestinal loss of protein and intestinal inflammation. The intestinal bacterial dysbiosis appears to outlast the protein loss and inflammation.

Evolving Definitions of Remission in Crohn’s Disease

Using clinical symptoms alone to inform treatment decisions in Crohn's disease (CD) may increase the risk of disease progression and complications. Treatment beyond symptoms may offer improved outcomes. We explore alternative definitions of remission, beyond traditional clinical remission, incorporating more objective parameters of inflammation control, which may support prevention or delay the disease progression. These definitions could serve as a platform for future clinical research, evaluating whether treating beyond symptoms alters the natural history of CD. Proposed definitions may include endoscopic remission (mucosal healing), normalization of serologic or fecal markers of inflammation, and even radiographic remission, in addition to clinical remission (symptom control). Endoscopic remission is the leading candidate for inclusion because it is the best studied. The definition should include considerations for both early and late disease given that in late disease, which may be associated with operation-related symptoms or irreversible bowel damage, symptomatic remission may not achievable. Desired outcomes in early disease are complete absence of symptoms, no disease progression, no complications or disability, and normal quality of life. In late disease, there are stabilization of noninflammatory symptoms, no progression of damage or disability, and improved quality of life. Over time, we anticipate that a working definition of remission that includes both biological remission and clinical remission will evolve and be evaluated in clinical trials. Our proposed definition is a possible starting point for that evolution. Ultimately, the goal in evolving the definition of remission is to improve the outcomes in patients with CD.

Update of fecal markers of inflammation in inflammatory bowel disease

The diagnosis, prognosis, and assessment of disease activity of inflammatory bowel disease (IBD) require investigating clinical, radiological, and histological criteria, as well as serum inflammatory markers. However, a range of fecal inflammatory markers now appears to have the potential to greatly assist in these processes. Calprotectin, a prominent neutrophil protein, was identified two decades ago as a potentially revolutionary marker for IBD. Following this discovery, numerous additional markers, including S100A12, lactoferrin, and M2-pyruvate kinase, have also been suggested as novel markers of IBD. In the present study, we provide an up-to-date review of fecal markers of IBD, and further, provide a novel analysis of each of these fecal markers in severe ulcerative colitis and compare their expression pattern in contrast to calprotectin.

Discrete changes in circulating regulatory T cells during infliximab treatment of Crohn's disease

Deficiency of CD4+CD25+ regulatory T cells (Tregs) may be involved in Crohn's disease (CD) pathogenesis. In rheumatoid arthritis (RA), the anti-TNF-α antibody infliximab increases circulating Treg numbers. We aimed to evaluate circulating Tregs in CD before and after infliximab therapy. In 20 patients with active CD, blood samples were obtained before infusion of infliximab 5 mg/kg and 1, 7, and 42 days after therapy. Clinical, biochemical, and fecal markers of inflammation were obtained. Nine healthy volunteers served as controls. We applied a novel Treg marker, the absence of CD127 expression, to identify Tregs by whole-blood flow cytometry. Treg percentages were similar among CD patients [median 7.7%, interquartile range (IQR) 5.3–10.1%] and healthy volunteers (median 7.6% IQR 6.3–8.9%) with discrete changes (median 7.3%, IQR 4.5–10.1%) throughout the study period, irrespective of the significant clinical effect of infliximab. Unlike in RA, we found no arising population of CD62L − Tregs; however, we observed a rapid recruitment of lymphocytes and upregulation of the intestinal homing marker α4β7 integrin on CD4+T cells. In conclusion, our results do not support the hypothesis that the clinical effect of infliximab is mediated by a reinforcement of defective, circulating Tregs in CD.

Elevated Human β-Defensin-2 Levels Indicate an Activation of the Innate Immune System in Patients With Irritable Bowel Syndrome

Irritable bowel syndrome (IBS) is a highly prevalent functional disorder. According to the Rome criteria, macroscopic and histological inflammation is a crucial exclusion criterion for IBS. Human defensins appear to be part of the innate immune system in the gastrointestinal tract. Human beta-defensin-2 (HBD-2) was the first inducible human antimicrobial protein discovered. The expression is induced by probiotic microorganisms and proinflammatory cytokines. Recent results imply that HBD-2 is expressed in active intestinal inflammation, especially in ulcerative colitis (UC). Our aim was to evaluate fecal measurements of HBD-2 in patients with active UC and IBS, and in healthy controls (HCs). Fecal specimens were collected from a total of 100 participants (30 with active UC, 46 IBS, and 24 HCs). Exclusion criteria were the current use of probiotics and antibiotics. Furthermore, IBS patients with elevated C-reactive protein or leukocytes, a history of bacterial overgrowth or infectious gastrointestinal disease over the last 6 month were excluded. Disease status was addressed in all participating subjects by medical history and current symptoms. In addition, each IBS and UC patient underwent ileocolonoscopy with histopathology. Fecal inflammation markers lactoferrin (Lf) and calprotectin (Cal) were measured by enzyme-linked immunosorbent assay (ELISA) and reported as microg/g. Fecal HBD-2 was measured by ELISA and reported as ng/g feces. In addition, immunoblots were performed for fecal HBD-2. Paraffin-embedded tissue from colonic biopsies was tested for HBD-2 peptides by immunohistochemistry. Lf as well as Cal was elevated in active UC (mean: 152.1+/-s.d. 374.7 microg/g; 103.5+/-87.1 microg/g), compared with IBS (8.3+/-19.4 microg/g; 18.6+/-23.3 microg/g), and HCs (0.4+/-0.5 microg/g; 7.1+/-7.9 microg/g). Scheffe post hoc tests revealed significant differences (P=0.006; P<0.001) between active UC vs. IBS and HC. In contrast, HBD-2 levels were highest in active UC (mean: 106.9+/-s.d. 91.5 ng/g), almost as high in IBS (pts 76.0+/-67.9 ng/g), and lowest for HCs (29.9+/-16.1 ng/g). Scheffe post hoc tests revealed significant differences (P<0.001) between the groups of patients (UC and IBS) vs. HCs. Immunohistochemical investigation was consistent with fecal secretion data and demonstrated the presence of beta-defensin 2 peptides in colonic epithelial enterocytes in UC as well as IBS patients with elevated fecal HBD-2. The results indicate significantly elevated levels of HBD-2 in patients with IBS compared with HCs and similar to those with active UC. The results support an activation of the mucosal innate defense system toward a proinflammatory response in IBS patients in the absence of macroscopic signs of inflammation.

Fecal markers of inflammation used as surrogate markers for treatment outcome in relapsing inflammatory bowel disease

To evaluate fecal calprotectin (FC) as a surrogate marker of treatment outcome of relapse of inflammatory bowel disease (IBD) and, to compare FC with fecal myeloperoxidase (MPO) and fecal eosinophil protein X (EPX). Thirty eight patients with IBD, comprising of 27 with ulcerative colitis (UC) and 11 with Crohn's disease (CD) were investigated before treatment (inclusion), and after 4 and 8 wk of treatment. Treatment outcomes were evaluated by clinical features of disease activity and endoscopy in UC patients, and disease activity in CD patients. In addition, fecal samples were analyzed for FC by enzyme-linked immunosorbent assay (ELISA), and for MPO and EPX with radioimmunoassay (RIA). At inclusion 37 of 38 (97%) patients had elevated FC levels (> 94.7 microg/g). At the end of the study, 31 of 38 (82%) patients fulfilled predefined criteria of a complete response [UC 21/27 (78%); CD 10/11 (91%)]. Overall, a normalised FC level at the end of the study predicted a complete response in 100% patients, whereas elevated FC level predicted incomplete response in 30%. Normalised MPO or EPX levels predicted a complete response in 100% and 90% of the patients, respectively. However, elevated MPO or EPX levels predicted incomplete response in 23% and 22%, respectively. A normalised FC level has the potential to be used as a surrogate marker for successful treatment outcome in IBD patients. However, patients with persistent elevation of FC levels need further evaluation. FC and MPO provide superior discrimination than EPX in IBD treatment outcome.