Variable alterations of the microbiota, without metabolic or immunological change, following faecal microbiota transplantation in patients with chronic pouchitis.

Faecal microbiota transplantation (FMT) is effective in the treatment of Clostridium difficile infection, where efficacy correlates with changes in microbiota diversity and composition. The effects of FMT on recipient microbiota in inflammatory bowel diseases (IBD) remain unclear. We assessed the effects of FMT on microbiota composition and function, mucosal immune response, and clinical outcome in patients with chronic pouchitis. Eight patients with chronic pouchitis (current PDAI ≥7) were treated with FMT via nasogastric administration. Clinical activity was assessed before and four weeks following FMT. Faecal coliform antibiotic sensitivities were analysed, and changes in pouch faecal and mucosal microbiota assessed by 16S rRNA gene pyrosequencing and (1)H NMR spectroscopy. Lamina propria dendritic cell phenotype and cytokine profiles were assessed by flow cytometric analysis and multiplex assay. Following FMT, there were variable shifts in faecal and mucosal microbiota composition and, in some patients, changes in proportional abundance of species suggestive of a "healthier" pouch microbiota. However, there were no significant FMT-induced metabolic or immunological changes, or beneficial clinical response. Given the lack of clinical response following FMT via a single nasogastric administration our results suggest that FMT/bacteriotherapy for pouchitis patients requires further optimisation.

Is right-sided colon cancer different to left-sided colorectal cancer? - a systematic review.

Colorectal cancer (CRC) exhibits differences in incidence, pathogenesis, molecular pathways and outcome depending on the location of the tumor. This review focuses on the latest developments in epidemiological and scientific studies, which have enhanced our understanding on the underlying genetic and immunological differences between the proximal (right-sided) colon and the distal (left-sided) colorectum. The different ways in which environmental risk factors influence the pathogenesis of CRC depending on its location and the variations in surgical and oncological outcomes are also discussed in this review. In the current era of personalized medicine, we aim to reiterate the importance of tumor location in management of CRC and the implication on future clinical and scientific research.

Seven-year experience of enterocutaneous fistula with univariate and multivariate analysis of factors associated with healing: development of a validated scoring system.

AIM: The management of enterocutaneous fistulae (ECF) is complex and challenging. We examined factors associated with fistula healing at a National Intestinal Failure Centre and devised the first scoring system to predict spontaneous fistula healing prior to surgery. METHOD: A retrospective audit of 177 patients (mean age 48.7 years) treated over 7 years was undertaken. Results were compared with a previously reported series from this unit. Univariate and multivariate analyses wete performed on variables to assess relationship with ECF healing. A scoring system was devised and validated on a prospective cohort. RESULTS: One-hundred and fifty patients underwent surgery between January 2003 and December 2009. The overall healing rate following surgery in the current series was 94.6% (82% in the previous series). Mean delay from previous surgery to the current operation was 1 year (compared with 8 months previously). Thirty-day postfistula resection mortality was 0% (compared with 3.5% previously). Twenty-seven patients underwent medical management alone with overall healing rate of 46.4% (vs 19.9%). Multivariate analysis revealed that comorbidity (P = 0.02), source of referral (P = 0.01) and aetiology (P = 0.006) had associations with healing. Almost all patients with scores of 0 and 1 healed, whereas the highest scores healed least frequently. CONCLUSION: Surgical management of ECF is safe and improving. Fistula healing is affected by aetiology, comorbidity and source of referral. The scoring system has the potential to predict ECF healing and can be a useful clinical decision-making tool.

Lost therapeutic potential of monocyte-derived dendritic cells through lost tissue homing: stable restoration of gut specificity with retinoic acid.

Human monocyte-derived dendritic cells (DC) (MoDC) are utilized for immunotherapy. However, in-vitro immunological effects are often not mirrored in vivo. We studied the tissue-homing potential of MoDC. Circulating monocytes and DC expressed different tissue-homing markers and, during in-vitro development of MoDC, homing marker expression was lost resulting in a 'homeless' phenotype. Retinoic acid (RA) induced gut-homing markers (ß7 and CCR9) and a regulatory phenotype and function [decreased human leucocyte antigen D-related (HLA-DR) and increased ILT3 and fluorescein isothiocyanate (FITC-dextran uptake) in MoDC]. RA-MoDC were less stimulatory and primed conditioned T cells with a gut-homing profile (ß7(+)CLA(-)). Unlike the normal intestinal microenvironment, that from inflamed colon of ulcerative colitis (UC) patients did not induce regulatory properties in MoDC. However, RA-MoDC maintained their regulatory gut-specific properties even in the presence of UC microenvironment. Therefore, MoDC may be ineffectual for immunotherapy because they lack tissue-homing and tissue-imprinting specificity. However, MoDC rehabilitation with gut-homing potential by RA could be useful in promoting immunotherapy in pathologies such as UC.

A mechanistic role for leptin in human dendritic cell migration: differences between ileum and colon in health and Crohn's disease.

Dendritic cells (DC) migrate to lymph nodes on expression of C-C motif chemokine receptor 7 (CCR7) and control immune activity. Leptin, an immunomodulatory adipokine, functions via leptin receptors, signaling via the long isoform of receptor, LepRb. Leptin promotes DC maturation and increases CCR7 expression on blood DC. Increased mesenteric fat and leptin occur early in Crohn's disease (CD), suggesting leptin-mediated change in intestinal CCR7 expression on DC as a pro-inflammatory mechanism. We have demonstrated CCR7 expression and capacity to migrate to its ligand macrophage inflammatory protein 3ß in normal human ileal DC but not colonic or blood DC. In CD, functional CCR7 was expressed on DC from all sites. Only DC populations containing CCR7-expressing cells produced LepRb; in vitro exposure to leptin also increased expression of functional CCR7 in intestinal DC in a dose-dependent manner. In conclusion, leptin may regulate DC migration from gut, in homeostatic and inflammatory conditions, providing a link between mesenteric obesity and inflammation.

Skin- and gut-homing molecules on human circulating γδ T cells and their dysregulation in inflammatory bowel disease.

Changes in phenotype and function of γδ T cells have been reported in inflammatory bowel disease (IBD), including Crohn's disease (CD) and ulcerative colitis (UC). Dysregulation of lymphocyte migration plays a key role in IBD pathogenesis; however, data on migratory properties of γδ T cells are scarce. Human circulating γδ T cells from healthy controls (n = 27), patients with active CD (n = 15), active UC (n = 14) or cutaneous manifestations of IBD (n = 2) were characterized by flow cytometry. Circulating γδ T cells in healthy controls were CD3(hi) and expressed CD45RO. They expressed gut-homing molecule ß7 but not gut-homing molecule corresponding chemokine receptors (CCR)9, or skin-homing molecules cutaneous lymphocyte-associated antigen (CLA) and CCR4, despite conventional T cells containing populations expressing these molecules. CCR9 expression was increased on γδ T cells in CD and UC, while skin-homing CLA was expressed aberrantly on γδ T cells in patients with cutaneous manifestations of IBD. Lower levels of CD3 expression were found on γδ T cells in CD but not in UC, and a lower proportion of γδ T cells expressed CD45RO in CD and UC. Enhanced expression of gut-homing molecules on circulating γδ T cells in IBD and skin-homing molecules in cutaneous manifestations of IBD may be of clinical relevance.

Etiology of pouchitis.

Restorative proctocolectomy with ileal-pouch anal anastomosis (RPC) is the operation of choice for ulcerative colitis (UC) patients requiring surgery. It is also used for patients with familial adenomatous polyposis (FAP). Pouchitis accounts for 10% of pouch failures. It is an idiopathic inflammatory condition that may occur in up to 50% of patients after RPC for UC. It is rarely seen in FAP patients after RPC. The etiology of pouchitis remains unclear. An overlap with UC is suggested by the frequency with which pouchitis affects patients with UC compared with FAP patients. There is significant clinical evidence implicating bacteria in the pathogenesis of pouchitis. Studies using culture and molecular methods demonstrate a dysbiosis of the pouch microbiota in pouchitis. Risk factors, genetic associations, and serological markers of pouchitis suggest that the interactions between the host immune responses and the pouch microbiota underlie the etiology of this idiopathic inflammatory condition. Here we present a detailed review of the data focusing on the pouch microbiota and the immune responses that support this hypothesis. We also discuss the contribution of luminal metabolic factors and the epithelial membrane in the etiology of this inflammatory process. The ileoanal pouch offers a unique opportunity to study the inter-relationships between the gut microbiota and host immune responses from before the onset of disease. For this reason the study of pouchitis could serve as a human model that significantly enhances our understanding of inflammatory bowel diseases in general.

Review article: faecal transplantation therapy for gastrointestinal disease.

BACKGROUND: Evidence is emerging regarding the relationship between a dysbiosis of the human gut microbiota and a number of gastrointestinal diseases as well as diseases beyond the gut. Probiotics have been investigated in many gastrointestinal disease states, with variable and often modest outcomes. Faecal transplantation is an alternative approach to manipulate the gut microbiota. AIM: To review the use of faecal transplantation therapy for the management of gastrointestinal disorders. METHODS: Available articles on faecal transplantation in the management of gastrointestinal disorders were identified using a Pubmed search and bibliographies of review articles on the subject were collated. RESULTS: A total of 239 patients who had undergone faecal transplantation were reported. Seventeen of 22 studies of faecal transplantation were in fulminant or refractory Clostridium difficile. Studies of faecal transplantation are heterogeneous regarding the patients, donors, screening, methods of administration and definition of response. Faecal transplantation for C. difficile has been demonstrated to be effective in 145/166 (87%) patients. Small numbers of patients are reported to have undergone successful faecal transplantation for irritable bowel syndrome and inflammatory bowel disease. CONCLUSIONS: Faecal transplantation has been reported with good outcomes for fulminant and refractory C. difficile. No adverse effects of faecal transplantation have been reported. However, there are no level 1 data of faecal transplantation and reports to date may suffer from reporting bias of positive outcomes and under-reporting of adverse effects. This therapy holds great promise, where a dysbiosis of the gut microbiota is responsible for disease and further studies are necessary to explore this potential.

A novel population of human CD56+ human leucocyte antigen D-related (HLA-DR+) colonic lamina propria cells is associated with inflammation in ulcerative colitis.

Ulcerative colitis (UC) involves inappropriate mucosal immune responses to intestinal microbiota. Gut dendritic cells (DC) are central immunoregulators of the response to commensal bacteria, and the subset of CD11c(+) cells within the human leucocyte antigen D-related (HLA-DR(+)) lineage (lin)(-/dim) population are activated in inflammatory bowel disease. We hypothesized that CD11c(-) cells within this population may also be involved in intestinal inflammation. HLA-DR(+) lin(-/dim) cells were identified in freshly isolated lamina propria mononuclear cells by multi-colour flow cytometry in 54 UC patients and 22 controls. Proportion and number of CD11c(+) and CD11c(-) cells, and surface expression of activation markers CD40, CD86, Toll-like receptor (TLR)-2, TLR-4, and CD56(+)[natural killer (NK) marker], were determined. Cytokine production was assessed by intracellular staining. Lamina propria colonic CD11c(-) HLA-DR(+) lin(-/dim) cells were increased significantly in inflamed and 'non-inflamed' UC tissue, compared with control tissue. CD11c(+) HLA-DR(+) lin(-/dim) cells were unchanged. Fewer CD11c(-) cells expressed activation markers and produced intracellular cytokines than their CD11c(+) counterparts, and they were weakly stimulatory in mixed leucocyte reactions. Few CD11c(-) cells expressed blood plasmacytoid DC markers, but a major subset expressed high levels of CD56. CD11c(-) cells decreased after inflammation resolved. Intestinal inflammation in UC is associated with the presence of cells that share phenotypic features of both DC and NK cells. This novel population of human colonic CD56(+) HLA-DR(+) cells may play a role in immune regulation or tissue repair. Their increase in quiescent UC may be a marker of subclinical inflammation.

Inhaled allergen-driven CD1c up-regulation and enhanced antigen uptake by activated human respiratory-tract dendritic cells in atopic asthma.

BACKGROUND: Dendritic cells (DC) mediate inflammation in rodent models of allergic airway disease, but the role played by human respiratory-tract DC (hRTDC) in atopic asthma remains poorly defined. Recent data suggest that CD1 antigen presentation by hRTDC may contribute to asthma pathogenesis. OBJECTIVE: To investigate the influence of hRTDC on the balance between atopy and allergic asthma in human subjects and to determine whether CD1 expression by hRTDC is modulated during asthmatic inflammation. METHODS: Sputum cells were induced from steroid-naïve, allergen-challenged and allergen-naïve subjects (atopic asthmatics, atopic non-asthmatics and non-atopic controls). hRTDC were identified using monoclonal antibody labelling and analysis by flow cytometry. RESULTS: hRTDC stained HLA-DR(+) (negative for markers of other cell lineages) were predominantly myeloid and comprised approximately 0.5% of viable sputum cells. Sputum cells were potent stimulators of allogeneic CD4(+) naïve T cells and enrichment/depletion experiments correlated stimulatory potency with DC numbers. Sputum contained cells that exhibited typical dendritic morphology when analysed by electron microscopy. Myeloid hRTDC were endocytically active, but uptake of FITC-dextran was enhanced in cells from asthmatics (P<0.001). Despite their increased endocytic capacity, asthmatic myeloid hRTDC appeared mature and expressed increased levels of maturation markers (P<0.05-P<0.001), CD1c, CD1d and langerin (P<0.05). CD1c expression by asthmatic myeloid hRTDC was enhanced upon in vivo allergen challenge (three to ninefold within 24 h; P<0.05). CD11c(-)CD123(high) hRTDC were only detected in asthmatic sputum and were increased in number following allergen challenge. CONCLUSION: Despite limited cell numbers, it proved possible to analyse human RTDC in induced sputum, providing evidence that increased antigen uptake and enhanced CD1 presentation by activated hRTDC may contribute to allergic airway disease. CD1 presentation by hRTDC in atopic asthma may therefore constitute a novel target for future intervention strategies.

Clinical, microbiological, and immunological effects of fructo-oligosaccharide in patients with Crohn's disease.

BACKGROUND AND AIMS: The intestinal microbiota play a pivotal role in the inflammation associated with Crohn's disease through their interaction with the mucosal immune system. Some bifidobacteria species are immunoregulatory and induce increased dendritic cell interleukin 10 (IL-10) release in vitro. Fructo-oligosaccharides (FOS) increase faecal and mucosal bifidobacteria in healthy volunteers. The aim of this study was to assess the effect of FOS administration on disease activity, bifidobacteria concentrations, and mucosal dendritic cell function in patients with moderately active Crohn's disease. PATIENTS AND METHODS: Ten patients with active ileocolonic Crohn's disease received 15 g of FOS for three weeks. Disease activity was measured using the Harvey Bradshaw index. Faecal and mucosal bifidobacteria were quantified by fluorescence in situ hybridisation, and mucosal dendritic cell IL-10 and Toll-like receptor (TLR) expression were assessed by flow cytometry of dissociated rectal biopsies. RESULTS: FOS induced a significant reduction in the Harvey Bradshaw index from 9.8 (SD 3.1) to 6.9 (3.4) (p<0.01). There was a significant increase in faecal bifidobacteria concentration from 8.8 (0.9) log(10) to 9.4 (0.9) log(10) cells/g dry faeces (p<0.001). The percentage of IL-10 positive dendritic cells increased from 30 (12)% to 53 (10)% (p=0.06). Finally, the percentage of dendritic cells expressing TLR2 and TLR4 increased from 1.7 (1.7)% to 36.8 (15.9)% (p=0.08) and from 3.6 (3.6)% to 75.4 (3.4)% (p<0.001), respectively. CONCLUSIONS: FOS supplementation increases faecal bifidobacteria concentrations and modifies mucosal dendritic cell function. This novel therapeutic strategy appears to decrease Crohn's disease activity in a small open label trial and therefore warrants further investigation.