Mutations in RAD21 disrupt regulation of apob in patients with chronic intestinal pseudo-obstruction

Background Aims Chronic intestinal pseudo-obstruction (CIPO) is characterized by severe intestinal dysmotility that mimics a mechanical subocclusion with no evidence of gut obstruction. We searched for genetic variants associated with CIPO to increase our understanding of its pathogenesis and to identify potential biomarkers. Methods We performed whole-exome sequencing of genomic DNA from patients with familial CIPO syndrome. Blood and lymphoblastoid cells were collected from patients and controls (individuals without CIPO); levels of messenger RNA (mRNA) and proteins were analyzed by quantitative reverse-transcription polymerase chain reaction, immunoblot, and mobility shift assays. Complementary DNAs were transfected into HEK293 cells. Expression of rad21 was suppressed in zebrafish embryos using a splice-blocking morpholino (rad21a). Gut tissues were collected and analyzed. Results We identified a homozygous mutation (p.622, encodes Ala>Thr) in RAD21 in patients from a consanguineous family with CIPO. Expression of RUNX1, a target of RAD21, was reduced in cells from patients with CIPO compared with controls. In zebrafish, suppression of rad21a reduced expression of runx1; this phenotype was corrected by injection of human RAD21 mRNA, but not with the mRNA from the mutated p.622 allele. rad21a Morpholino zebrafish had delayed intestinal transit and greatly reduced numbers of enteric neurons, similar to patients with CIPO. This defect was greater in zebrafish with suppressed expression of ret and rad21, indicating their interaction in the regulation of gut neurogenesis. The promoter region of APOB bound RAD21 but not RAD21 p.622 Ala>Thr; expression of wild-type RAD21 in HEK293 cells repressed expression of APOB, compared with control vector. The gut-specific isoform of APOB (APOB48) is overexpressed in sera from patients with CIPO who carry the RAD21 mutation. APOB48 also is overexpressed in sporadic CIPO in sera and gut biopsy specimens. Conclusions Some patients with CIPO carry mutations in RAD21 that disrupt the ability of its product to regulate genes such as RUNX1 and APOB. Reduced expression of rad21 in zebrafish, and dysregulation of these target genes, disrupts intestinal transit and the development of enteric neurons. © 2015 by the AGA Institute

Is gastroparesis a gastric disease?

Background: Gastroparesis is a digestive syndrome characterized by delayed gastric emptying (GE) and by symptoms that are suggestive of gastroduodenal motor disorders. There are three grades of gastroparesis of increasing severity: (a) mild gastroparesis; (b) compensated gastroparesis; and (c) gastric failure. GE abnormalities are partially related to symptom type and severity, and other mechanisms may be involved. Aim: To investigate enteric dysmotility (ED) in patients with suspected gastroparesis. Methods: Patients with symptoms suggestive of gastroparesis were consecutively included in the study and underwent a 13 C-octanoic acid GE breath test and small bowel manometry (SBM). Clinical features were recorded using predefined, validated questionnaires at entry. Key Results: The study enrolled 88 patients (71 women; mean age: 37.8 \ub1 14.3 years). Gastric emptying was delayed in 25 patients (28.4%), and 70 patients (79.5%) presented small bowel motor abnormalities including bursts, abnormal activity fronts, inability to respond to meal ingestion, and hypocontractility. Gastric emptying was delayed in 24 of the 70 patients with ED (34.3% vs 5.5% of patients with normal SBM). Enteric dysmotility was detected in 24 of 25 patients (96%) with delayed GE. Patients with and without delayed GE showed similar moderate/severe gastroparesis manifestations, but patients with ED significantly more often had moderate/severe gastroparesis manifestations than patients with normal SBM (grade 1:14% vs 39%, grade 2:62% vs 56%, grade 3:24% vs 5%, respectively). Conclusions and Inferences: Enteric dysmotility was more frequent than delayed GE in patients with symptoms suggestive of gastroparesis. Gastroparesis severity was associated with small bowel motor abnormalities but not with delayed GE

Enteric neuron density correlates with clinical features of severe gut dysmotility

Gastrointestinal (GI) symptoms can originate from severe dysmotility due to enteric neuropathies. Current methods used to demonstrate enteric neuropathies are based mainly on classic qualitative histopathological/immunohistochemical evaluation. This study was designed to identify an objective morphometric method for paraffin-embedded tissue samples to quantify the interganglionic distance between neighboring myenteric ganglia immunoreactive for neuron-specific enolase, as well as the number of myenteric and submucosal neuronal cell bodies/ganglion in jejunal specimens of patients with severe GI dysmotility. Jejunal full-thickness biopsies were collected from 32 patients (22 females; 16 –77 yr) with well-characterized severe dysmotility and 8 controls (4 females; 47–73 yr). A symptom questionnaire was filled before surgery. Mann-Whitney U test, Kruskal-Wallis coupled with Dunn’s posttest and nonparametric linear regression tests were used for analyzing morphometric data and clinical correlations, respectively. Compared with controls, patients with severe dysmotility exhibited a significant increase in myenteric interganglionic distance (P = 0.0005) along with a decrease in the number of myenteric (P < 0.00001) and submucosal (P < 0.0004) neurons. A 50% reduction in the number of submucosal and myenteric neurons correlated with an increased interganglionic distance and severity of dysmotility. Our study proposes a relatively simple tool that can be applied for quantitative evaluation of paraffin sections from patients with severe dysmotility. The finding of an increased interganglionic distance may aid diagnosis and limit the direct quantitative analysis of neurons per ganglion in patients with an interganglionic distance within the control range

Randomised clinical trial: the analgesic properties of dietary supplementation with palmitoylethanolamide and polydatin in irritable bowel syndrome

Background: Intestinal immune activation is involved in irritable bowel syndrome (IBS) pathophysiology. While most dietary approaches in IBS involve food avoidance, there are fewer indications on food supplementation. Palmithoylethanolamide, structurally related to the endocannabinoid anandamide, and polydatin are dietary compounds which act synergistically to reduce mast cell activation. Aim: To assess the effect on mast cell count and the efficacy of palmithoylethanolamide/polydatin in patients with IBS. Methods: We conducted a pilot, 12-week, randomised, double-blind, placebo-controlled, multicentre study assessing the effect of palmithoylethanolamide/polydatin 200\ua0mg/20\ua0mg or placebo b.d. on low-grade immune activation, endocannabinoid system and symptoms in IBS patients. Biopsy samples, obtained at screening visit and at the end of the study, were analysed by immunohistochemistry, enzyme-linked immunoassay, liquid chromatography and Western blot. Results: A total of 54 patients with IBS and 12 healthy controls were enrolled from five European centres. Compared with controls, IBS patients showed higher mucosal mast cell counts (3.2\ua0\ub1\ua01.3 vs. 5.3\ua0\ub1\ua02.7%, P\ua0=\ua00.013), reduced fatty acid amide oleoylethanolamide (12.7\ua0\ub1\ua09.8 vs. 45.8\ua0\ub1\ua055.6 pmol/mg, P\ua0=\ua00.002) and increased expression of cannabinoid receptor 2 (0.7\ua0\ub1\ua00.1 vs. 1.0\ua0\ub1\ua00.8, P\ua0=\ua00.012). The treatment did not significantly modify IBS biological profile, including mast cell count. Compared with placebo, palmithoylethanolamide/polydatin markedly improved abdominal pain severity (P\ua0<\ua00.05). Conclusions: The marked effect of the dietary supplement palmithoylethanolamide/polydatin on abdominal pain in patients with IBS suggests that this is a promising natural approach for pain management in this condition. Further studies are now required to elucidate the mechanism of action of palmithoylethanolamide/polydatin in IBS. ClinicalTrials.gov number, NCT01370720