Cinzia Papadia

Plasma Citrulline Concentration: A Reliable Marker of Small Bowel Absorptive Capacity Independent of Intestinal Inflammation

OBJECTIVES:Postabsorptive plasma citrulline concentration has been proposed as a reliable marker of small bowel absorptive capacity in short bowel patients. The aim of this study was to address the potentially confounding impact of intestinal inflammation.METHODS:Fifty-five patients were selected according to diagnosis, small bowel length, and degree of bowel inflammation. (a) Crohns disease (CD) with massive small bowel resection leaving ≤50 cm (N = 6), (b) CD with 50–150 cm remaining (N = 9), (c) CD with no resection but active inflammation (high C-reactive protein [CRP] and Crohns Disease Activity Index [CDAI] >220) (N = 7), (d) CD without resection or active inflammation (normal CRP and CDAI <150) (N = 9), (e) mesenteric infarction (MI) with resection leaving ≤50 cm (N = 6), (f) MI leaving 50–150 cm (N = 6), (g) active celiac disease (N = 6), (h) healthy volunteers (N = 6). Postabsorptive fasting plasma citrulline was measured using reverse-phase, high performance liquid chromatography. Absorptive capacity and permeability were also measured after oral sugar-mix ingestion (5 g lactulose, 1 g L-rhamnose, 0.5 g D-xylose).RESULTS:The plasma citrulline strongly correlated with small bowel length (P < 0.0001) and xylose absorption (P < 0.001). No correlation was found with CDAI, permeability, CRP, albumin, sedimentation rate, white cell count, or platelet count. Citrulline was significantly higher (P < 0.0004) in CD and MI patients with a remnant small bowel length of 50–150 cm (mean 21.0 μmol/L) than in those with length ≤50 cm (mean 9.2 μmol/L).CONCLUSIONS:Plasma citrulline concentration is a simple and reliable surrogate for small bowel absorptive capacity and is not influenced by intestinal inflammation.

Plasma citrulline as a quantitative biomarker of HIV-associated villous atrophy in a tropical enteropathy population.

BACKGROUND & AIMS Studies have shown that the circulating citrulline concentration is decreased in patients with proximal small bowel villous atrophy from coeliac disease and more so in patients with extensive damage to the intestinal mucosa, but there have been few data on HIV enteritis and tropical enteropathy (TE). Our primary aim was to correlate serum citrulline with the degree of reduction of the enterocyte mass in HIV-infected patients with TE. METHODS Postabsorptive fasting serum citrulline was measured in 150 TE pts, 44 of whom had HIV infection, using reverse phase, high performance liquid chromatography. Absorptive capacity and permeability were measured after intrajejunal instillation of 4 sugars (5 g lactulose, 1 g L-rhamnose, 0.5 g D-xylose, 0.2 g 3-O methyl D glucose) with assay by thin-layer chromatography. Morphometric analysis was carried out on jejunal biopsies. RESULTS In HIV positive patients, the median serum citrulline was significantly lower (median 19, interquartile range (IQR) 17-24 μmol/L) than in HIV negative patients (median 27, IQR 23-33 μmol/L; p < 0.001). There were statistically significant correlations (p < 0.005) between citrulline and: crypt depth; villous height/crypt depth ratio; Shenk-Klipstein score; and xylose absoption, only in the HIV positive. CONCLUSIONS Serum citrulline concentration appears to be a quantitative biomarker of small bowel mass integrity in HIV positive enteropathy and deserves assessment as a surrogate for monitoring anti-retroviral therapy.

Diagnosing small bowel malabsorption: a review

Malabsorption encompasses dysfunctions occurring during the digestion and absorption of nutrients. A small proportion of patients presents with chronic diarrhoea. A clinical history supportive of malabsorption may guide investigations toward either the small bowel or pancreas. Serological testing for coeliac disease will determine most cases without invasive investigations. In the clinical context of persisting weight loss and malnutrition, small bowel enteropathy may be investigated with small intestinal biopsies. Small bowel absorptive capacity and permeability might be measured by oral sugar-mix ingestion. Further, approaches to the investigation of malabsorption might also involve the detection in faeces of a substance that has not been absorbed. A variation of the latter is the use of breath testing which relies on the breakdown of the malabsorbed test substance by colonic flora. Measurement of protein absorption is difficult and unreliable; it is, therefore, rarely advocated in clinical settings. No single biological marker confirming a diagnosis of small bowel malabsorption or small bowel integrity is presently available in clinical practice. Plasma citrulline concentration, an amino acid not incorporated into endogenous or exogenous proteins, has been extensively used in research studies and supportive results are establishing its concentration as a reliable quantitative biomarker of enterocyte absorptive capacity.

FOXE1 and SYNE1 Genes Hypermethylation Panel as Promising Biomarker in Colitis-associated Colorectal Neoplasia

Background:Colitis-associated colorectal cancer affects individuals with inflammatory bowel disease (IBD) more often and earlier than cancer in the general population. Colonoscopy provides the surveillance gold standard. Changes to the surveillance intervals depending on endoscopic activity have been made, given data demonstrating that this is an important predictor of future dysplasia or cancer, but adjuvant, noninvasive clinical tools are still warranted to improve surveillance outcomes and to assist in management and interpretation of dysplasia. Methylation markers may be able to do this. Methods:SYNE1, FOXE1, NDRG4, and PHACTR3 genes were screened using methylation-specific PCR that permit the methylation status of the genes to be determined directly on biopsies. Ninety-three patients with long-standing IBD undergoing a cancer surveillance program, and 30 healthy controls were studied. These included colorectal adenocarcinomas on a background of IBD of various stages (n = 25), IBD-associated dysplastic lesions (n = 29), adenomas arising on a background of ulcerative colitis (n = 8), samples from patients with no evidence of dysplasia or cancer but long-standing IBD (n = 31), and symptomatic patients found to have normal colonoscopy (controls) (n = 30). Results:Gene promotor hypermethylation of SYNE1 and FOXE1 genes varied significantly between the groups and was increasingly likely with increased disease severity. Neither occurred in controls, whereas promotor hypermethylation was detected in biopsies of 60% of patients with colitis-associated colorectal cancer for FOXE1 and 80% for SYNE1. Promotor hypermethylation of either gene was highly significantly different between the groups overall. Conclusions:FOXE1 and SYNE1 hypermethylation markers demonstrated significantly increased expression in neoplastic tissue. Promoter methylation analysis of these genes might be a useful marker of neoplasia in long-standing IBD.

Sensitivity and specificity of magnetic resonance enterography in the clinical management of fistulizing Crohn's disease.

Background:High diagnostic accuracy is reported for magnetic resonance enterography (MRE) in Crohn’s disease (CD), but few studies have evaluated its role in abdominal fistulae. The primary aim of this study was to assess the reliability of MRE in the identification of internal fistulae in CD. Methods:One hundred and eighty-six patients with moderate CD (CD Activity Index : 250–400) were prospectively selected from the inflammatory bowel disease clinic of Parma University Hospital. Eligible patients had already undergone nutritional screening, pancolonoscopy, and computed tomography enterography (CTE) in the month before enrollment. MRE was performed according to the study protocol. Additional fluoroscopic contrast-enhanced studies or surgical evaluation were used for discordance between CTE and MRE results. A consensus committee resolved equivocal findings. Surgical findings and/or fluoroscopic contrast-enhanced studies together with the clinical data were considered the composite “reference standard” to which the results of MRE were compared. Results:MRE identified 22 internal fistulae in 21 patients (11%), of whom 4 (19%) also had perianal fistulae and found 7 abscesses (33%). Forty-one (22%) additional patients with perianal fistulae were identified. Thirteen patients (57%) with internal fistulae required enteral nutrition support. No statistically significant differences were found between MRE and CTE in fistula detection. There was also no significant difference between MRE and the composite diagnosis in those who underwent surgery (n = 8) and/or contrast-enhanced studies (n = 7). Conclusions:CTE and MRE accurately detect internal fistulae in CD. MRE is preferable because it avoids radiation. Reliable identification of internal fistulae by MRE should permit earlier and improved treatment.

Abnormal thymic stromal lymphopoietin expression in the duodenal mucosa of patients with coeliac disease

Objective The short isoform of thymic stromal lymphopoietin (TSLP), a cytokine constitutively expressed by epithelial cells, is crucial in preserving immune tolerance in the gut. TSLP deficiency has been implicated in sustaining intestinal damage in Crohns disease. We explored mucosal TSLP expression and function in refractory and uncomplicated coeliac disease (CD), a T-cell-mediated enteropathy induced by gluten in genetically susceptible individuals. Design TSLP isoforms—long and short—and receptors—TSLPR and interleukin (IL)-7Rα—were assessed by immunofluorescence, immunoblotting and qRT-PCR in the duodenum of untreated, treated, potential and refractory patients with CD. The ability of the serine protease furin or CD biopsy supernatants to cleave TSLP was evaluated by immunoblotting. The production of interferon (IFN)-γ and IL-8 by untreated CD biopsies cultured ex vivo with TSLP isoforms was also assessed. Results Mucosal TSLP, but not TSLPR and IL-7Rα, was reduced in untreated CD and refractory CD in comparison to treated CD, potential CD and controls. Transcripts of both TSLP isoforms were decreased in active CD mucosa. Furin, which was overexpressed in active CD biopsies, was able to cleave TSLP in vitro. Accordingly, refractory and untreated CD supernatants showed higher TSLP-degrading capacity in comparison to treated CD and control supernatants. In our ex vivo model, both TSLP isoforms significantly downregulated IFN-γ and IL-8 production by untreated CD biopsies. Conclusions Reduced mucosal TSLP expression may contribute to intestinal damage in refractory and untreated CD. Further studies are needed to verify whether restoring TSLP might be therapeutically useful especially in refractory patients with CD.

Altered Expression of Type-1 and Type-2 Cannabinoid Receptors in Celiac Disease

Anandamide (AEA) is the prominent member of the endocannabinoid family and its biological action is mediated through the binding to both type-1 (CB1) and type-2 (CB2) cannabinoid receptors (CBR). The presence of AEA and CBR in the gastrointestinal tract highlighted their pathophysiological role in several gut diseases, including celiac disease. Here, we aimed to investigate the expression of CBR at transcriptional and translational levels in the duodenal mucosa of untreated celiac patients, celiac patients on a gluten-free diet for at least 12 months and control subjects. Also biopsies from treated celiac patients cultured ex vivo with peptic-tryptic digest of gliadin were investigated. Our data show higher levels of both CB1 and CB2 receptors during active disease and normal CBR levels in treated celiac patients. In conclusion, we demonstrate an up-regulation of CB1 and CB2 mRNA and protein expression, that points to the therapeutic potential of targeting CBR in patients with celiac disease.

Omega-3 fatty acids in the maintenance of ulcerative colitis

Immunomodulation of the gut associated lynphoid tissue is a key issue in the clinical management of inflammatory bowel disease (IBD). Often toxic drugs are used to obtain clinical remission, sometimes in already immunocompromized patients. The presence of important co-morbidity might also heavily affect the clinical strategy. Polyunsaturated fatty acids (PUFAs) might represent a valid therapeutical option in IBD patients and further controlled clinical studies are warranted.

Abnormal anandamide metabolism in celiac disease

The endocannabinoid system has been extensively investigated in experimental colitis and inflammatory bowel disease, but not in celiac disease, where only a single study showed increased levels of the major endocannabinoid anandamide in the atrophic mucosa. On this basis, we aimed to investigate anandamide metabolism in celiac disease by analyzing transcript levels (through quantitative real-time reverse transcriptase-polymerase chain reaction), protein concentration (through immunoblotting) and activity (through radioassays) of enzymes responsible for anandamide synthesis (N-acylphosphatidyl-ethanolamine specific phospholipase D, NAPE-PLD) and degradation (fatty acid amide hydrolase, FAAH) in the duodenal mucosa of untreated celiac patients, celiac patients on a gluten-free diet for at least 12 months and control subjects. Also, treated celiac biopsies cultured ex vivo with peptic-tryptic digest of gliadin were investigated. Our in vivo experiments showed that mucosal NAPE-PLD expression and activity are higher in untreated celiac patients than treated celiac patients and controls, with no significant difference between the latter two groups. In keeping with the in vivo data, the ex vivo activity of NAPE-PLD was significantly enhanced by incubation of peptic-tryptic digest of gliadin with treated celiac biopsies. On the contrary, in vivo mucosal FAAH expression and activity did not change in the three groups of patients, and accordingly, mucosal FAAH activity was not influenced by treatment with peptic-tryptic digest of gliadin. In conclusion, our findings provide a possible pathophysiological explanation for the increased anandamide concentration previously shown in active celiac mucosa.

Citrulline in health and disease. Review on human studies

The amino acid L-citrulline (CIT) is safely used from the neonatal period onwards in those with urea cycle defects and carbamyl phosphate synthetase or ornithine transcarbamylase deficiencies, but several lines of enquiry indicate that it might have a much wider therapeutic role. When protein intake is low and there is a catabolic state, endogenous arginine (ARG) synthesis cannot fully be met and its supplementation can prove challenging, particularly in patients with critical and multisystem illness. Supplementary CIT could constitute a safer but still focused means of delivering ARG to endothelial and immune cells as CIT is efficiently recycled into these cells and as kidneys can convert CIT into ARG. Unlike ARG, CIT is efficiently transported into enterocytes and bypasses liver uptake. It also appears to prevent excessive and uncontrolled nitric oxide (NO) production. Animal studies and early human data indicate positive effects of CIT on protein synthesis, in which its contribution is thought mediated through the mTOR pathway. It appears that CIT is an anabolic pharmaconutrient that can be safely administered even in critically ill patients. Promising results in cardiovascular diseases and in disease-related malnutrition can now be considered sufficient to justify formal clinical exploration in these areas and in sarcopenia in general.