Comparison of pre-analytical conditions for quantification of serotonin in platelet-poor plasma

Background: Reported concentrations of serotonin in platelet-poor plasma (PPP) in healthy subjects vary widely due to different pre-analytical procedures. Aim: To examine how different pre-analytical conditions affect the measured concentration of serotonin in PPP. Method: Six pre-analytical protocols were compared for preparation of PPP from EDTA whole blood for quantification of serotonin from nine healthy individuals. Three combinations of centrifugation with a mild centrifugation of gel-free EDTA tubes followed by a stronger centrifugation were compared to single-stage centrifugation of EDTA tubes with separator gel and heat shock treatment of blood prior to centrifugation. All samples were analysed using the same enzyme linked immunosorbent assay (ELISA) method. Results: Findings show that two consecutive centrifugations; first a mild centrifugation at 100 or 200×g followed by centrifugation at 4500 or 14500×g resulted in the lowest serotonin concentration in PPP. Conclusion: Two successive centrifugations to produce PPP for serotonin analysis; first a mild centrifugation to avoid mechanical stress on the platelets, and next a stronger centrifugation to remove platelets, is superior to the use of gel tubes and heat shock treatment.publishedVersio

Intestinal barrier integrity in anorexia nervosa (a pilot study)

Objective There is no conclusive evidence for involvement of intestinal barrier alteration in the etiology of anorexia nervosa (AN). The aims of this pilot study were to identify serum markers of intestinal barrier integrity in patients with AN and to determine the relationships between those markers and body mass index (BMI), eating disorder symptoms, gastrointestinal complaints, and liver synthesis function (international normalized ratio [INR]). Method Twenty-five outpatients with AN prior to starting treatment and 28 healthy controls (HC) were assessed. BMI and serum markers of intestinal barrier integrity were measured, including zonulin family peptides (ZFP), lipopolysaccharide-binding protein (LBP), and intestinal fatty-acid-binding protein (i-FABP). Eating disorder symptoms and gastrointestinal complaints were evaluated via questionnaires. Results The serum ZFP concentration was significantly lower in patients with AN than in HC (44.2 [7.4] vs. 49.2 [5.6] ng/ml, mean [standard deviation], p = .008). LBP and i-FABP did not differ between the two groups. In patients with AN, serum ZFP was significantly predicted by BMI (β = 0.479, p = .009), age (β = 0.411, p = .020), and INR (β = −0.388, p = .028). No such associations were found for either gastrointestinal complaints or eating disorder symptoms. Discussion Abnormal levels of serum ZFP were observed in patients with AN. Further studies with other assessment methods are warranted to examine intestinal barrier function in AN.publishedVersio

The effect of supplementation with low doses of a cod protein hydrolysate on satiety hormones and inflammatory biomarkers in adults with metabolic syndrome: A randomized, double-blind study

Metabolic syndrome (MetS) is characterised by metabolic abnormalities that increase the risk of developing type 2 diabetes mellitus and cardiovascular disease. Altered levels of circulating ghrelin, several adipokines and inflammatory markers secreted from adipose tissue, such as leptin, adiponectin, tumor necrosis factor alpha, are observed in overweight and obese individuals. We assessed the effect of supplementation with low doses of a cod protein hydrolysate (CPH) on fasting and postprandial levels of acylated ghrelin, as well as fasting levels of adiponectin, leptin and inflammatory markers in subjects with MetS. A multicentre, double-blinded, randomized controlled trial with a parallel group design was conducted. Subjects received a daily supplement of CPH (4 g protein, n = 15) or placebo (0 g protein, n = 15). We observed no effect on fasting or postprandial levels of acylated ghrelin, fasting levels of adiponectin (p = 0.089) or leptin (p = 0.967) after supplementation with CPH, compared to placebo. Overall, our study showed that 8 weeks supplementation with a low dose of CPH in subjects with MetS had no effect on satiety hormones or most of the inflammatory markers, but the levels of high-sensitivity C-reactive protein were statistically significantly different in the CPH-group compared to placebo group. The robustness and clinical relevance of these findings should be explored in future studies with a larger sample size.publishedVersio

Plasma levels of guanylins are reduced in patients with Crohn’s disease

Background: Guanylin (GN) and uroguanylin (UGN) are endogenous ligands for the intestinal receptor guanylate cyclase C (GC-C), an important regulator of intestinal fluid homeostasis. Gene expression and protein levels of GN are suppressed in inflamed intestinal tissue from patients with inflammatory bowel disease (IBD), but knowledge about plasma levels of guanylins in these conditions is sparse. We aimed to investigate the fasting plasma levels of the prohormones proGN and proUGN in patients with Crohn’s Disease (CD) and relate these to levels found in persons with other diarrheal conditions, as well as persons with normal bowel habits. Methods: Plasma from patients with CD, patients with Familial GUCY2C Diarrheal Disease (FGDS), diarrhea-predominant irritable bowel syndrome (IBS-D) and healthy controls (HC) was analyzed using ELISA assays. Results: Significantly lower fasting plasma levels of proguanylins were found in CD and FGDS patients, compared to HC. In CD patients, plasma proGN levels correlated negatively with Harvey Bradshaw Index and with number of stools/24 h. Conclusion: Our data indicate that diarrhea may be a determinant for levels of proGN in plasma, and should be further explored in studies of different diarrheal disorders.acceptedVersio

Study protocol of the Bergen brain-gut-microbiota-axis study: A prospective case-report characterization and dietary intervention study to evaluate the effects of microbiota alterations on cognition and anatomical and functional brain connectivity in patients with irritable bowel syndrome

Introduction: Irritable bowel syndrome (IBS) is a common clinical label for medically unexplained gastrointestinal (GI) symptoms, recently described as a disturbance of the brain-gut-microbiota (BGM) axis. To gain a better understanding of the mechanisms underlying the poorly understood etiology of IBS, we have designed a multifaceted study that aim to stratify the complex interaction and dysfunction between the brain, the gut, and the microbiota in patients with IBS. Methods: Deep phenotyping data from patients with IBS (n = 100) and healthy age- (between 18 and 65) and gender-matched controls (n = 40) will be collected between May 2019 and December 2021. Psychometric tests, questionnaires, human biological tissue/samples (blood, faeces, saliva, and GI biopsies from antrum, duodenum, and sigmoid colon), assessment of gastric accommodation and emptying using transabdominal ultrasound, vagal activity, and functional and structural magnetic resonance imaging (MRI) of the brain, are included in the investigation of each participant. A subgroup of 60 patients with IBS-D will be further included in a 12-week low FODMAP dietary intervention-study to determine short and long-term effects of diet on GI symptoms, microbiota composition and functions, molecular GI signatures, cognitive, emotional and social functions, and structural and functional brain signatures. Deep machine learning, prediction tools, and big data analyses will be used for multivariate analyses allowing disease stratification and diagnostic biomarker detection. Discussion: To our knowledge, this is the first study to employ unsupervised machine learning techniques and incorporate systems-based interactions between the central and the peripheral components of the brain-gut-microbiota axis at the levels of the multiomics, microbiota profiles, and brain connectome of a cohort of 100 patients with IBS and matched controls; study long-term safety and efficacy of the low-FODMAP diet on changes in nutritional status, gut microbiota composition, and metabolites; and to investigate changes in the brain and gut connectome after 12 weeks strict low-FODMAP-diet in patients with IBS. However, there are also limitations to the study. As a restrictive diet, the low-FODMAP diet carries risks of nutritional inadequacy and may foster disordered eating patterns. Strict FODMAP restriction induces a potentially unfavourable gut microbiota, although the health effects are unknown.publishedVersio

Genome-wide copy number variation (CNV) in patients with autoimmune Addison's disease

<p>Abstract</p> <p>Background</p> <p>Addison's disease (AD) is caused by an autoimmune destruction of the adrenal cortex. The pathogenesis is multi-factorial, involving genetic components and hitherto unknown environmental factors. The aim of the present study was to investigate if gene dosage in the form of copy number variation (CNV) could add to the repertoire of genetic susceptibility to autoimmune AD.</p> <p>Methods</p> <p>A genome-wide study using the Affymetrix GeneChip^®Genome-Wide Human SNP Array 6.0 was conducted in 26 patients with AD. CNVs in selected genes were further investigated in a larger material of patients with autoimmune AD (n = 352) and healthy controls (n = 353) by duplex Taqman real-time polymerase chain reaction assays.</p> <p>Results</p> <p>We found that low copy number of <it>UGT2B28 </it>was significantly more frequent in AD patients compared to controls; conversely high copy number of <it>ADAM3A </it>was associated with AD.</p> <p>Conclusions</p> <p>We have identified two novel CNV associations to <it>ADAM3A </it>and <it>UGT2B28 </it>in AD. The mechanism by which this susceptibility is conferred is at present unclear, but may involve steroid inactivation (<it>UGT2B28</it>) and T cell maturation (<it>ADAM3A</it>). Characterization of these proteins may unravel novel information on the pathogenesis of autoimmunity.</p

Assessment of markers of gut integrity and inflammation in non-celiac gluten sensitivity after a gluten free-diet

Purpose: Markers for gut integrity and inflammation have received increasing interest as intestinal permeability and innate immune system activation are suggested as possible pathophysiological mechanisms in non-celiac gluten sensitivity (NCGS). We aimed to assess relevant biomarkers in NCGS by analyzing serum levels of gut integrity and permeability markers, pro-inflammatory cytokines and antigliadin IgG in patients with suspected NCGS on a gluten-free diet (GFD), and compare them to serum levels in patients with irritable bowel syndrome (IBS) and healthy controls (HC). Patients and Methods: Serum samples collected from patients with suspected NCGS on a GFD (n=20, 14 women, 21– 62 years), IBS (n=20, 16 women, 24– 67 years) and HC (n=20, 14 women, 21– 54 years) were analyzed. IBS severity scoring system (IBS-SSS) was applied to evaluate gastrointestinal symptoms. Results: The IBS-SSS score was higher in subjects with suspected NCGS and IBS patients compared to HC (p< 0.0001). No significant differences were found in the serum levels of any of the gut integrity and permeability markers, cytokines or antigliadin IgG antibodies between the three groups. However, positive correlations were observed between claudin-1 and i-FABP, and between claudin-1 and antigliadin IgG antibodies. Conclusion: No differences in serum levels of gut integrity and permeability markers, pro-inflammatory cytokines or antigliadin IgG antibodies were found among patients with suspected NCGS on a GFD, IBS and HC.publishedVersio

Comparison of pre-analytical conditions for quantification of serotonin in platelet-poor plasma

Background: Reported concentrations of serotonin in platelet-poor plasma (PPP) in healthy subjects vary widely due to different pre-analytical procedures. Aim: To examine how different pre-analytical conditions affect the measured concentration of serotonin in PPP. Method: Six pre-analytical protocols were compared for preparation of PPP from EDTA whole blood for quantification of serotonin from nine healthy individuals. Three combinations of centrifugation with a mild centrifugation of gel-free EDTA tubes followed by a stronger centrifugation were compared to single-stage centrifugation of EDTA tubes with separator gel and heat shock treatment of blood prior to centrifugation. All samples were analysed using the same enzyme linked immunosorbent assay (ELISA) method. Results: Findings show that two consecutive centrifugations; first a mild centrifugation at 100 or 200×g followed by centrifugation at 4500 or 14500×g resulted in the lowest serotonin concentration in PPP. Conclusion: Two successive centrifugations to produce PPP for serotonin analysis; first a mild centrifugation to avoid mechanical stress on the platelets, and next a stronger centrifugation to remove platelets, is superior to the use of gel tubes and heat shock treatment