Non-inflamed Pouches Research Articles

The Microbiota Profile in Inflamed and Non-Inflamed Ileal Pouch-Anal Anastomosis.

The objective was to determine the bacterial composition in inflamed and non-inflamed pouches for comparison to the microbiota of healthy individuals. Pouch patients and healthy individuals were included between November 2017 and June 2019 at the Department of Gastrointestinal Surgery, Aalborg University Hospital, Denmark. A faecal sample was collected from all participants for microbiota analysis using 16S rRNA amplicon sequencing. Overall, 38 participants were included in the study. Eleven patients with a normally functioning pouch, 9 patients with chronic pouchitis, 6 patients with familial adenomatous polyposis, and 12 healthy individuals. Patients with chronic pouchitis had overall lower microbial diversity and richness compared to patients with a normal pouch function (p < 0.001 and p = 0.009) and healthy individuals (p < 0.001 and p < 0.001). No significant difference was found between patients with familial adenomatous polyposis and chronic pouchitis (microbial diversity p = 0.39 and richness p = 0.78). Several taxa from the family Enterobacteriaceae, especially genus Escherichia, were associated primarily with patients with chronic pouchitis, while taxa from the genus Bacteroides primarily were associated with healthy individuals and patients with a normally functioning pouch. Finally, a microbial composition gradient could be established from healthy individuals through patients with normal pouch function and familial adenomatous polyposis to patients with chronic pouchitis.

Pouch wall thickness and floppy pouch complex.

Floppy pouch complex (FPC) consists of disease phenotypes in patients with ileal pouches, including pouch prolapse, afferent limb syndrome, enterocele, redundant loop, and pouch folding. Our recent study demonstrated that lower body weight, lower peripouch fat, family history of inflammatory bowel disease (IBD), female gender, and dyschezia are risk factors for FPC patients with IBD. The aims of this study were to assess the relationship between pouch wall thickness and FPC, and to investigate the association between inflamed and non-inflamed pouch wall thickness. This case-control study included all eligible patients with FPC from our prospectively maintained, IRB-approved Pouchitis Registry from 2011 to 2017. We measured pouch wall thickness of fully distended pouches on cross-sectional abdominal and pelvic imaging. Patients with stoma and non-distended pouches were completely excluded. Risk factors for FPC were analyzed. A total of 140 out of 451 patients from our were found to have fully distended pouches on imaging. Of the 140 patients, 36 (25.7%) were diagnosed as having FPC. We analyzed pouch wall thickness for each subcategory of FPC as well as non-FPC conditions. The thickness of pouch wall was follows: pouch prolapse (N = 19): 1.5mm (1.5-2.0), afferent limb syndrome (N = 12): 1.5mm (1.1-2.0), folded pouch (N = 4): 1.5mm (1.1-1.9), and redundant pouch (N = 2): 1.3mm (1.0-1.3). The control group (N = 104) consisting of normal pouch, pouchitis, cuffitis, Crohn's disease of the pouch, and pouch sinus with median pouch wall thickness of 1.5mm, 2.3mm, 2.0mm, 2.0mm, and 1.5mm, respectively. There were significant differences in pouch wall thickness between normal or non-inflamed pouch versus pouchitis versus cuffitis versus Crohn's disease ofthe pouch with p values of 0.01, 0.04, 0.05, and 0.049, respectively. Patients with FPC were shown to have thin pouch wall, which those with inflammatory conditions of the pouch tended to have thick pouch wall. These findings will have implications in both diagnosis and investigation of etiopathogenesis of these disorders.

PWE-077 Altered Epithelial Tight Junction Expression and Elevated Il 6 Levels in Pouchitis

IntroductionIntestinal epithelial barrier function limits the interactions between microbial antigens and the mucosal immune system. In IBD, epithelial barrier function is impaired with altered expression of tight junctions. We aimed...

Antibodies against glycoprotein 2 are novel markers of intestinal inflammation in patients with an ileal pouch

Background and aimsThe Crohn's disease (CD)-specific pancreatic auto-antibodies (PAB), have been recently identified to target glycoprotein 2 (GP2). Pouchitis is an inflammation of the small bowel developing in up to 60% of ulcerative colitis patients undergoing proctocolectomy and ileal pouch anal anastomosis. Occurrence of CD-specific antibodies was reported to be a predictor of pouchitis. We aimed to assess the prevalence of anti-GP2 antibodies (anti-GP2) in the serum and feces of pouch patients and to correlate them with clinical parameters. Furthermore, we examined mucosal expression of the GP2 protein in the pouch. MethodsPouch patients were prospectively recruited and checked for clinical, endoscopic, and laboratory markers of inflammation. IgG and IgA anti-GP2 levels in serum and fecal samples were determined using ELISA. GP2 protein was assessed by immunohistochemistry. ResultsAnti-GP2 was elevated in both serum and fecal samples of patients with inflamed compared to those with non-inflamed pouches and patients with familial-adenomatous polyposis after surgery (p<0.05, respectively). Moreover, patients with CD-like complications exhibited significantly higher anti-GP2 titers than those without CD-like complications (p≤0.01). High levels of anti-GP2 correlated with more frequent bowel movements per day and with the presence of at least one anti-glycan antibody (p≤0.05). GP2 itself was more abundant in the mucosa of patients with chronic pouchitis. ConclusionsAnti-GP2 exists in the serum and feces of pouch patients and correlates with pouch inflammation, and presence of other serological markers. Thus, anti-GP2 may contribute to better stratification of pouchitis, more-so when the inflammation exhibits CD-like complications.

Pouchitis may increase the risk of dysplasia after restorative proctocolectomy in patients with ulcerative colitis.

Dysplasia of the pouch mucosa after restorative proctocolectomy is rare. The aim of this study was to establish whether there is a correlation between pouchitis and dysplasia. A group of 276 patients treated for ulcerative colitis by restorative proctocolectomy between 1984 and 2009 was analysed. The presence or absence of pouchitis and dysplasia within the pouch was evaluated. Inflammation was diagnosed in 66 (23.9%) patients, low-grade dysplasia in five (1.8%), high-grade dysplasia in three (1.1%), and cancer in one patient (0.4%). The prevalence of low-grade dysplasia was significantly higher in patients with inflammation than in those without (P < 0.04). High-grade dysplasia was significantly more frequent in pouchitis than in non-inflamed pouches (P < 0.01). Logistic regression analysis suggested that the occurrence of mucosal inflammation increased the risk of low grade dysplasia. Patients with chronic pouchitis are at risk of dysplasia and require surveillance of the pouch.

A hypothesis for the mode of action of anti-rheumatic drugs in a model of cartilage destruction.

Abstract We have been using as a model rats and mice with air pouches which develop lining cells closely resembling synovium (Edwards et al 1982; Sedgwick et al 1983). These pouches when sufficiently mature, i.e. 6 days, will respond to a variety of stimuli both immune and non-immune with long lasting exudates containing many migrated leucocytes (Sedgwick et al 1983, 1984a). We examined the breakdown of cartilage in both inflamed and non-inflamed pouches and have found that inflammation protected the rate of loss of proteoglycan from cartilage (Sedgwick et al 1984b. 1985; Willoughby et al 1985). Invariably the cartilage in the inflamed air pouch would float free in exudate while cartilage, in the non-inflamed pouch would adhere to the pouch wall in close proximity to the macrophages and fibroblasts of the lining cells. Unpublished findings from this department (de Brito) have shown that the presence of granulation tissue in close proximity to cartilage will speed proteoglycan loss.

Antibiotics and probiotics in chronic pouchitis: a comparative proteomic approach.

To profile protein expression in mucosal biopsies from patients with chronic refractory pouchitis following antibiotic or probiotic treatment, using a comparative proteomic approach. Two-dimensional polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry were used to characterize the changes related to antibiotic therapy in the protein expression profiles of biopsy samples from patients with chronic refractory pouchitis. The same proteomic approach was applied to identify differentially expressed proteins in the non-inflamed pouch before and after probiotic administration. In the first set of 2D gels, 26 different proteins with at least 2-fold changes in their expression levels between the pouchitis condition and antibiotic-induced remission were identified. In the second set of analysis, the comparison between mucosal biopsy proteomes in the normal and probiotic-treated pouch resulted in 17 significantly differently expressed proteins. Of these, 8 exhibited the same pattern of deregulation as in the pouchitis/pouch remission group. For the first time, 2D protein maps of mucosal biopsies from patients with ileal pouch-anal anastomosis were provided, and differentially expressed proteins following antibiotic/probiotic treatment were identified.

Bacterial community diversity in cultures derived from healthy and inflamed ileal pouches after restorative proctocolectomy

Pouchitis is believed to occur as a reaction to dysbiosis. In this study we assessed differences between mucosal bacterial communities cultured from noninflamed and inflamed ileal pouches. Thirty-two ileal pouch patients, 22 with ulcerative colitis (UC) and 10 with familial adenomatous polyposis (FAP), underwent symptomatic, endoscopic, and histological assessment. The Objective Pouchitis Score (OPS) and the Pouch Disease Activity Index (PDAI) were used to diagnose pouchitis. Seven UC patients had pouchitis (UC+), 15 had a noninflamed pouch (UC-), 9 had a noninflamed pouch (FAP-), and 1 FAP patient had pouchitis (FAP+). Biopsies taken from the ileal mucosa of the pouch were cultured under aerobic and anaerobic conditions. Following standardized DNA extraction a polymerase chain reaction (PCR) was performed to generate 16S rRNA gene products. A "fingerprint" of the bacterial community within each sample was created using terminal-restriction fragment length polymorphism (T-RFLP) profiling. Species richness and evenness were determined using T-RF band lengths and relative band intensities. From the 64 DNA samples, 834 bands were detected, of which 179 represented different species (operational taxonomic units [OTUs]). The average species richness for the FAP-, FAP+, UC-, and UC+ groups was 26, 35, 23.9, and 29.6 per patient, with the average species diversity within the groups of 10.6, 29, 8.3, and 11.4, respectively. Similar trends were observed when the anaerobic and aerobic-derived bacterial groups were analyzed separately. No significant differences were found between the bacterial cultures derived from any of the clinical groups or between pouchitis and nonpouchitis patients.

Inflamed pouch mucosa possesses altered tight junctions indicating recurrence of inflammatory bowel disease

The etiology of pouchitis after coloproctomucosectomy with ileal pouch-anal anastomosis in patients with ulcerative colitis is still unknown. Beside changes in luminal antigens, the immunological predisposition is assumed to be responsible. In previous electrophysiological studies, we showed that mucosal barrier and transport function in pouchitis is markedly reduced. Thus, the aim of the present study was to analyze barrier function on the molecular level. Pouch biopsies of 36 ulcerative colitis patients were analyzed. Time points were (1) intraoperative immediately prior to ileal pouch-anal anastomosis (n = 13), (2) >1 year after ileostomy closure (pouch, n = 12), and (3) during pouchitis (n = 11). Control terminal ileum biopsies were obtained from eight patients undergoing hemicolectomy due to carcinoma. Expression of tight junction proteins was analyzed by Western blotting and confocal laser-scanning microscopy. To elucidate effects on epithelial barrier properties, impedance spectroscopy was performed in miniaturized Ussing chambers. In pouchitis, epithelial resistance was markedly reduced compared to non-inflamed pouch and control ileum. Expression of tight junction proteins claudin-1, 3, 4, 5, and 7 and occludin revealed differential expression regulation with the tightening tight junction protein claudin-1 being decreased and an increase of the pore-forming claudin-2, whereas other claudins remained constant. Morphometry indicated the mucosal surface to be unchanged. Pouchitis is characterized by a selective change of tight junction proteins in favor of opening the epithelial tight junction and, thus, the paracellular pathway, which contributes to the inflammatory process. This resembles changes in inflammatory bowel disease (IBD) and indicates IBD recurrence in pouchitis.

The Expression Patterns of Toll-Like Receptors in the Ileal Pouch Mucosa of Postoperative Ulcerative Colitis Patients

The aim of this study was to evaluate the expression pattern of Toll-like receptors (TLRs) in the pouch mucosa of ulcerative colitis patients in comparison with that in the ileum mucosa of noninflammatory bowel disease patients. Pouch mucosal biopsy specimens were collected from postoperative patients who had undergone surgery for ulcerative colitis. Normal ileum specimens were collected from colon cancer patients. The specimens were assessed by immunofluorescence histochemistry using TLR2, TLR3, TLR4, and TLR5 polyclonal antibodies. The normal ileal mucosa constitutively expressed TLR3 and TLR5, whereas TLR2 and TLR4 were barely detectable. In the mucosa of active pouchitis, TLR2 and TLR4 was strongly upregulated, and TLR4 was upregulated even in a noninflamed pouch. No TLR3 or TLR5 expression was detectable. These data suggest that pouchitis may be associated with distinctive changes in selective TLR expression in the pouch mucosa, and that TLR4 alterations in the innate response system may contribute to the pathogenesis of these disorders in particular.

Increased cytokine transcripts in pouchitis reflect the degree of inflammation but not the underlying entity

After ileopouch anal anastomosis (IPAA), 10-40% of patients with ulcerative colitis (UC) but only 5% of patients with familial adenomatous polyposis (FAP) develop pouchitis. Immunoregulatory abnormalities might be of importance in the pathogenesis of the disease. Therefore, we characterized cytokine and chemokine transcripts in inflamed and non-inflamed pouches in patients with UC compared to those with FAP and Crohn's disease (CD). Mucosal biopsies were taken from 87 patients with IPAA [UC (n=70), CD (n=8) or FAP (n=9)]. Patients with active ileal CD (n=14), active UC (n=17) and non-inflammatory conditions (n=12) served as controls. The expression of 20 gene transcripts was quantified using real-time polymerase chain reaction. Pro-inflammatory cytokines and chemokines are significantly increased in IPAA patients with acute pouchitis. This increase is independent of the underlying disease (UC or CD) and reflects the degree of inflammation. A good correlation between pouchitis activity (using the Pouchitis Disease Activity Index) and the MRP-14, interleukin-8, macrophage inflammatory protein-2alpha and matrix metalloproteinase-1 transcripts was observed. Our data support the view that pouchitis reflects an inflammatory process that is different from that of underlying inflammatory bowel diseases, as the cytokine and chemokine patterns in pouchitis are neither typical of CD nor of UC, but maybe due to bacterial intestinal microflora overgrowth in the pouch lumen. Quantification of transcript levels allows an estimation of the extent of mucosal inflammation and may become helpful in the evaluation of the disease, especially in clinical trials.

Altered Expression of the Lymphocyte Activation Markers CD30 and CD27 in Patients with Pouchitis

Background: The mechanism underlying the development of ileal pouch inflammation in ulcerative colitis patients (pouchitis) after restorative proctocolectomy is unclear. Persistent systemic T cell activation or expansion of specific memory cell populations could predispose certain patients to develop local inflammation within the neo-rectum. Therefore, the aim was to study the expression of the lymphocyte activation markers CD27, CD30, CD25 and CD69 on the CD45RO+ memory cell subset of isolated peripheral blood mononuclear cells (PBMC), soluble CD30 levels and mucosal CD30 expression in patients with pouchitis and in controls. Methods: Flow cytometry was performed on PBMC isolated from patients with pouchitis (n = 9), without pouchitis (n = 10) and normal controls (n = 9). Serum CD30 was measured in patients with pouchitis (n = 25), without pouchitis (n = 26) and normal controls (n = 20) by ELISA. CD30 expression was quantified in pouchitis (n = 15) and normal pouch (n = 15) mucosa using a three-stage immunoperoxidase method. Results: Naïve CD45RO-CD27+ PBMC were significantly decreased in pouchitis (25.6%) compared to normal controls (34.4%), (P = 0.03). CD30, CD25 and CD69 subsets did not differ between the groups. Serum CD30 was increased in pouchitis patients 58 (1-380) U/ml compared to non-pouchitis 16.5 (1-290) U/ml, P = 0.007, and normal controls 11 (2-80) U/ml, P = 0.0005. In the mucosa, the numbers of CD30+ cells were increased in pouchitis compared to noninflamed pouches (P = 0.02). Conclusions: Increased sCD30 in pouchitis is associated with elevated mucosal expression. Of the activation markers studied, only the circulating naïve CD27+ population differed in pouchitis patients compared with controls. The observed decrease in this cell type may reflect antigen priming and subsequent loss of CD27 implying that antigen driven activation of specific T cell subsets may occur in pouchitis.

Pouchitis: Microbiologic findings from biopsy and fecal samples in noninflamed and inflamed pouches

Pouchitis: Microbiologic findings from biopsy and fecal samples in noninflamed and inflamed pouches

Increased state of activation of CD4 positive T cells and elevated interferon γ production in pouchitis

Background—Immunoregulatory abnormalities of T cells might be of importance in the pathogenesis of pouchitis after ileoanal pouch anastomosis (IAP).Aims—To characterise T cell subsets, their state of activation, and production of...

Susceptibility of cartilage to damage by immunological inflammation.

Injection of ovalbumin into subcutaneous air pouches prepared on the backs of rats previously sensitised to the antigen resulted in the induction of a small and transient accumulation of inflammatory fluid with a predominantly polymorph cell infiltrate. Challenge of pouches of appropriately sensitised rats with Bordetella pertussis vaccine, on the other hand, resulted in a larger and more prolonged accumulation of fluid and cells with a predominantly mononuclear presence. When intact homologous femoral head cartilage was implanted in these inflamed pouches proteoglycan loss was found to be not different from similar implants in non-inflamed pouches. Coating the cartilage with human heat-aggregated immunoglobulin G prior to implantation in air pouches was also found to be without effect on subsequent proteoglycan loss.