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Introduction: Faecal calprotectin (FC) is commonly used as a diagnostic tool for patients with gastrointestinal (GI) symptoms. However, there is uncertainty in daily clinical practice how to interpret an elevated FC in patients with a normal colonoscopy. We investigated if patients with a normal colonoscopy but with an elevated FC more often were diagnosed with a GI disease in a 3-year follow-up period.

Methods: Patients referred for colonoscopy (n = 1,263) to the Umeå University Hospital endoscopy unit between 2007 and 2013 performed a FC test (CALPRO®) on the day before bowel preparation. A medical chart review was performed on all patients who had normal findings on their colonoscopy (n = 585, median age 64 years).

Results: Thirty-four percent of the patients (n = 202) with normal colonoscopy had elevated FC (>50 μg/g), and these patients were more frequently diagnosed with upper GI disease during the follow-up period than patients with normal FC levels (9.9 vs. 4.7%; p = 0.015). The upper GI diseases were mainly benign (i.e., gastritis). In a binary logistic regression analysis controlling for age, gender, nonsteroid anti-inflammatory drug use, and proton-pump inhibitor use, there was no difference for a new diagnosis of upper GI disease in the follow-up period (multivariate OR 1.70; 95% CI: 0.77–3.74). There was no difference in a new diagnosis of lower GI disease (6.4 vs. 5.2%; p = 0.545) or cardiovascular disease/death (multivariate OR 1.68; 95% CI: 0.83–3.42) in the follow-up period between patients with elevated versus normal FC levels.

Conclusions: In patients with a normal colonoscopy, a simultaneously measured increased FC level was not associated with an increased risk for significant GI disease during a follow-up period of 3 years.

Background: Our objective was to determine if patients who later develop inflammatory bowel disease (IBD) show signs of increased inflammatory activity in plasma measured with high sensitivity C-reactive protein (CRP), calprotectin, and albumin before the clinical onset of IBD.

Methods: We identified 96 subjects who later developed IBD (70 ulcerative colitis [UC] and 26 Crohn's disease [CD]). High sensitivity CRP, calprotectin, and albumin were analyzed in frozen plasma, donated from cases and sex-age matched controls 1-15 years before diagnosis.

Results: We found that subjects who later developed UC had lower albumin levels, and subjects who later developed CD had higher CRP levels than controls. Multivariable conditional logistic regression with albumin, calprotectin, and CRP showed a lower risk for developing IBD and UC with higher albumin levels (odds ratio [OR] 0.79, confidence interval [CI] 0.69-0.90; respective OR 0.77, CI 0.66-0.91). Higher CRP levels were associated with an increased risk of developing CD (OR 1.314, CI 1.060-1.630). When adjusting for body mass index or smoking in the logistic regression model, similar results were found. Plasma calprotectin levels in the preclinical period among patients with IBD did not differ from controls.

Conclusions: In this nested case-control study, subjects who later developed IBD had signs of low-grade systemic inflammation, indicated by significantly higher CRP plasma levels in CD and lower albumin plasma levels in UC, before the onset of clinical disease.

Background

Gastrointestinal (GI) symptoms are commonly reported in a normal population. Mostly, the symptoms are of benign cause but occasionally the symptoms can be signs of a more harmful disease. In general, it is difficult to distinguish whether the reported symptoms are caused by a benign (functional) or organic (i.e., inflammatory) disease. To make this distinction, the tools available in clinical practice are medical history, blood and faecal tests, radiology, endoscopy and histological evaluation. Mucosal inflammation usually separates organic from functional disease and, in patients with inflammatory bowel disease (IBD), mucosal inflammation correlates with disease activity. Faecal calprotectin (FC) corresponds well with mucosal inflammation and is in clinical practice often used as the first line non-invasive test for gut inflammation. Although the sensitivity of the FC test to detect gut inflammation is good, there are uncertainties in how to interpret a modestly elevated FC level (i.e., in the span of 50-200µg/g) and in patients with IBD, there is a disagreement into which degree of inflammatory remission it is sufficient to reach.

Aim

The overall aim of this thesis was to study factors associated with low-grade inflammation based on biochemical markers, and to study the clinical significance of low-grade inflammation in patients with IBD and other patients with elevated FC levels. Is low-grade inflammation associated with reported gastrointestinal symptoms in patients with IBD and could low-grade inflammation be detected in the pre-clinical phase of IBD? How should an elevated FC level in patients with a normal colonoscopy be interpreted and could it be a risk factor for gastrointestinal disease or associated with other factors? Could low-grade inflammation cause IBS-like symptoms in patients with IBD?

Methods and results

Three of the manuscripts on which this thesis is based are from the Faecal and Endoscopic Colorectal Study in Umeå Sweden (FECSU) which consists of 1263 patients that underwent colonoscopy during the period of May 2007 to February 2013. The patients that accepted to participate in the FECSU study performed a FC test the day before the bowel preparation for the colonoscopy and simultaneously filled in questionnaires of gastrointestinal symptoms (GSRS), symptoms of anxiety and depression (HADS) and current medications. A thorough medical chart review that focused on endoscopic evaluations, histological judgements and medical history was performed. The included patients with IBD (n=157) in the FECSU study were analysed separately. Patients with ulcerative colitis (UC) in endoscopic remission reported lower total scores on GSRS-irritable bowel syndrome (GSRS-IBS) than controls (6 vs 10.5; p=0.062). However there was a moderate, yet significant association between GSRS-diarrhoea score and FC levels in the span £ 200 µg/g (rho 0.38;p=0.004) in patients with UC. To investigate pre-clinical biomarkers of IBD we identified 96 patients with IBD in the “Västerbotten Intervention Program (VIP)” and the “Mammography screening project” (MA). In the pre-clinical study in patients with IBD we found that patients who later developed UC had lower plasma albumin levels and patients who later developed Crohn’s disease (CD) had higher levels of CRP in plasma, reflecting signs of a low-grade systemic inflammation years before diagnosis. Plasma calprotectin levels were not elevated before IBD-diagnosis. In the FECSU study, all non-IBD patients with a normal colonoscopy were studied for factors associated with an elevated FC level. Patients with a FC > 50 µg/g more often used Proton-pump inhibitors (PPI) (multivariate OR: 3.843; CI: 2.338-6.316), Non-steroidal anti-ivinflammatory drugs (NSAID) (multivariate OR: 2.411; CI: 1.162-5.002) and acetylsalicylic acid (ASA) (multivariate OR: 2.934; CI: 1.085-3.448). One third of the patients with a normal colonoscopy had elevated FC levels (> 50 µg/g) and these patients were observed three years after the colonoscopy. There was no increased risk for developing gastrointestinal disease in the patients with an increased baseline FC level and a normal colonoscopy during the observation period.

Conclusion

Patients with longstanding UC in remission did not experience more IBS-like symptoms than controls. In patients with UC in remission, the FC levels in the lower span were moderately associated with symptoms of diarrhoea. Patients with IBD had elevated inflammatory biochemical markers in blood in the pre-clinical phase. P- CRP and P-albumin were more sensitive to detect a low grade systemic inflammation than P-calprotectin in the pre-clinical phase of IBD. More than one-third of the patients with a normal colonoscopy had a slightly elevated FC. In patients with a normal colonoscopy, the use of PPI, NSAID and ASA was associated with an increased FC level. No significant gastrointestinal disease developed in the patients with an increased FC level together with a normal colonoscopy during the three-year period following colonoscopy. 

Background: The prevalence of irritable bowel syndrome (IBS)-like symptoms is high in untreated patients with microscopic colitis MC), but there is uncertainty of the prevalence of IBS-like symptoms in treated patients. We assessed the degree of IBS-like symptoms in patients with MC in comparison to control subjects, and investigated the association between IBS-like symptoms and faecal calprotectin (FC) in MC patients. Methods: Patients with an established MC diagnosis (n = 57) were compared to sex- and age-matched controls (n = 138) for scores in the GSRS-IBS (Gastrointestinal Symptom Rating Scale for Irritable Bowel Syndrome) and HADS (Hospital Anxiety Depression Scale). In MC patients, an FC level was simultaneously analysed. Results: The median interval from MC diagnoses to the time the subjects participated in the study was 5.5 years (25th-75th percentiles; 4.5-9.5 years). The total GSRS-IBS score, subscores for abdominal pain, bloating, and diarrhoea were significantly higher in MC patients compared to controls (all P< 0.001). There was a significant correlation between FC levels and reported bowel frequency (P = 0.023), but there was no correlation between FC levels and GSRS-IBS scores. Patients with MC had significantly higher scores on anxiety (HADS-A) (P< 0.001) and used more selective serotonin-reuptake-inhibitor drugs (P = 0.016) than the control subjects. However, only the control subjects (not the patients with MC) showed significant correlations between GSRS-IBS scores and HADS scores. Conclusions: Patients with MC reported more IBS-like symptoms and anxiety than control subjects but neither FC levels nor symptoms of affectivity were significantly correlated with IBS-like symptoms.

We performed a retrospective observational study to investigate the diagnostic accuracy of rigid sigmoidoscopy (RS) in patients with rectal cancer (n=279). Fifty-six percent of the patients had performed an RS within three months before diagnosis and mostly by a primary care provider (93%). In 21% of the patients the physician determined that the examination was normal, in 50% a rectal tumor was suspected and in 29% of cases an unspecific pathology (e.g. luminal blood, mucosal abnormalities) was reported. A normal finding on RS was associated with a longer time between the first appointment and subsequent diagnosis (multivariate hazard ratio (HR) 0.50; 95th percentile CI 0.35-0.71) whereas a history of rectal bleeding (multivariate HR 1.49; 95th percentile CI 1.01-2.20) and adherence to new national guidelines (multivariate HR 1.46; 95th percentile CI 1.08-1.99) was associated with a shorter time to diagnosis. We conclude that RS only had modest diagnostic accuracy in the diagnosis of rectal cancer, at least in this mainly primary care-based setting.

Faecal calprotectin (FC) tests and faecal immunological tests (FIT) for haemoglobin have been used to monitor disease activity in patients with ulcerative colitis (UC) but used alone they have some limitation concerning the predictive ability. We aimed to test if an FC test used in combination with FIT could improve the predictive ability. Consecutive out-patients with UC (n = 93) who were admitted for colonoscopy completed a single faecal sample before the start of bowel preparation. A quantitative CALPRO (R) calprotectin ELISA test and a qualitative FIT (cut-off < 40 ng/mL) were analyzed. An estimated Mayo score and a score of histological inflammation was performed blinded to the result of the faecal tests. The sensitivity, specificity, negative predictive value and positive predictive value for endoscopic inflammation (Mayo score > 1) was for FIT 85%, 83%, 96%, 57% and for FC > 186 mu g/g 73%, 87%, 87%, 54%. Corresponding results for FIT*FC > 186 mu g/g (at least one test positive) were 92%, 69%, 97%, 43%. For detecting moderate/severe histological inflammation the results were for FIT 69%, 79%, 92%, 43%, for FC > 75 mu g/g 95%, 62%, 98%, 41%, and for FIT*FC > 75 mu g/g 100%, 60%, 100%, 36%. None of the markers alone or in combination were useful to predict deep remission (Mayo score = 0 and no histological inflammation). We conclude that using the combination of an FC test and FIT shows minor improvement in predictive ability for inflammatory activity and remission in patients with UC.

Background

We aimed to determine whether patients who later develop IBD show signs of inflammatory activity in blood measured with high-sensitivity CRP, calprotectin and albumin before clinical onset of inflammatory bowel disease (IBD).

Methods

We identified 96 subjects who participated in the heath survey ‘Northern Sweden Health and Disease Study’ and who later developed IBD (70 UC and 26 CD). High-sensitivity CRP, calprotectin and albumin was analysed in stored blood donated from cases and sex-age-matched controls 1 to 15 years before diagnosis.

Results

We found that subjects who later developed UC had lower albumin levels and subject who later developed CD had higher levels of CRP compared with the controls. Multi-variate conditional logistic regression with albumin, calprotectin and CRP showed a lower risk for developing IBD and UC with higher albumin levels (OR 0.789; CI 0.691–0.901 respective OR 0.773; CI 0.657–0.909). Higher CRP levels were associated with increased risk of developing CD (OR 1.314; CI 1.060–1.630). Adding BMI or smoking in the logistic regression model similar results was found. Serum calprotectin levels in the prediagnostic period in patients with IBD did not differ from controls.

Conclusions

This nested case–control study show that subjects who later develop IBD have signs of low-grade systemic inflammation years before the diseases become clinical. CRP and albumin was more sensitive to detect low-grade systemic inflammation than calprotectin.

Objectives: Faecal Calprotectin (FC) is a sensitive marker for gut inflammation. However, slightly elevated FC levels are also common in subjects without inflammation. We investigated the association between FC and clinical factors including concomitant use of medical therapy in patients with a normal colonoscopy.Material and methods: Out-patients (n=1263) referred for colonoscopy, performed FC test (CALPRO) the day before the start of bowel preparation. All subjects answered questionnaires that included questions on the present and past health history, concomitant medical treatment and gastrointestinal symptoms (GSRS). A medical record chart review was performed to check for concomitant disease, cause of referral and the result of the colonoscopy including biopsies. Inclusion criteria were a normal colonoscopy. Exclusion criteria were inflammatory bowel disease, colon cancer and high-grade dysplasia.Results: Five hundred ninety subjects fulfilled the inclusion criteria and completed the study. Thirty-six per cent of the subjects had a FC >50 mu g/g. In a logistic regression analysis, age (adjusted OR: 1.051; CI: 1.032-1.071), and the use of proton pump inhibitors (adjusted OR: 3.843; CI: 2.338-6.316), non-steroid anti-inflammatory drugs (adjusted OR: 2.411; CI: 1.162-5.002) and acetylsalicylic acid (adjusted OR: 2.934; CI: 1.085-3.448) were significantly associated with an elevated FC (>50 mu g/g).Conclusions: More than one-third of the patients with a normal colonoscopy performed in clinical routine had a slightly elevated FC level. Our results emphasise the need for attention to age, the use of proton pump inhibitors, non-steroid anti-inflammatory drugs and acetylsalicylic acid in the interpretation of FC tests in clinical practice.

Aim: To investigate the diagnostic value of a combined analyses of faecal immunological haemoglobin (FIT) and faecal calprotectin (FC) in detection of colorectal cancer (CRC).

Methods: Out-patients (n=1440) referred to the endoscopy unit were analysed for FIT and FC in stool samples collected before the colonoscopy bowel preparation. The samples were collected from one defecation by the patients at home. Patients with IBD were excluded leaving stool samples from 1133 patients for further analyses. FIT was analysed using the immunological Analyse F.O.B Test and FC was analysed using the CALPRO® Calprotectin Elisa Test. Sensitivity and specificity to detect CRC was calculated for the individual tests, as well as for combined FIT/FC tests.

Results: Out of the included patients, 38 were diagnosed with CRC, 9 with high grade dysplasia (HGD), and 133 with low grade dysplasia (LGD). FIT was analysed in 673 (59.4%), FC in 1021 (90.1%) and both FIT and FC in 561 (49.5%) patients. A ROC curve analysis showed that the most accurate cut-off level for FC in detecting CRC in our study was 105.5 µg/g. The sensitivity for CRC when using FIT, FC (cut-off > 100 µg/g) and the combination of FIT and FC (at least one positive test) was 65.5%, 74.1% and 94.4%, respectively. The corresponding specificity was 84.8%, 74.9% and 68.3%, respectively.

Conclusion: Combined analyses of FIT and FC improved sensitivity for detection of CRC. Further studies in larger cohorts are required to find the optimal cut-off levels for different combinations of tests.

Objectives: To determine if 6-mercaptopurine (MP) is better tolerated than azathioprine (AZA) as the initial thiopurine treatment in patients suffering from inflammatory bowel disease (IBD). Switching patients with IBD from AZA to MP is advocated in patients intolerant to AZA. However, no study has determined if MP is more suited than AZA as a first-line treatment for patients who are naive to thiopurine treatment. Study: The tolerance of AZA and MP treatments in clinical practice was retrospectively evaluated from start to 12 months after initiating treatment in 113 patients with IBD who were all naive to thiopurines (82 patients treated with AZA and 31 patients with MP). Results: 65% of the patients treated with AZA and 61% of the patients treated with MP tolerated their treatment during 12 months (i.e., no group difference, p = 0.742). No difference in reported side effects between the two treatments was observed. The mean equivalent initial dose (0.92 vs. 0.61 mg/kg; p < 0.001) and the mean equivalent dose at 12 months (1.98 vs. 1.65 mg/kg; p = 0.014) was significantly higher in the MP group vs. the AZA group. The proportion of patients with.MCV = 7 at 12 months was numerically higher in the MP group than in the AZA group (53% vs. 31%; p = 0.090). Conclusions: In this retrospective observational study, no differences in tolerance or adherence between AZA and MP were observed in patients naive to thiopurines. However, MP treatment was at a higher equivalent thiopurine dose than AZA treatment, which tended to be associated with better treatment response.