The Pathogenesis of Extraintestinal Manifestations: Implications for IBD Research, Diagnosis, and Therapy.

This article reports on the sixth scientific workshop of the European Crohn's and Colitis Organisation [ECCO] on the pathogenesis of extraintestinal manifestations [EIMs] in inflammatory bowel disease [IBD]. This paper has been drafted by 15 ECCO members and 6 external experts [in rheumatology, dermatology, ophthalmology, and immunology] from 10 European countries and the USA. Within the workshop, contributors formed subgroups to address specific areas. Following a comprehensive literature search, the supporting text was finalized under the leadership of the heads of the working groups before being integrated by the group consensus leaders.

Regulation of human intestinal T-cell responses by type 1 interferon-STAT1 signaling is disrupted in inflammatory bowel disease.

Type 1 interferon (IFN-1) promotes regulatory T-cell function to suppress inflammation in the mouse intestine, but little is known about IFN-1 in the human gut. We therefore assessed the influence of IFN-1 on CD4+ T-cells isolated from human colon tissue obtained from healthy controls or patients with inflammatory bowel disease (IBD). Immunofluorescent imaging revealed constitutive expression of IFNß in human intestinal tissue, and colonic T-cells were responsive to exogenous IFN-1 as assessed by phosphorylation of signal transduction and activator of transcription 1 (pSTAT1) and induction of interferon stimulated genes (ISGs). Unlike their blood counterparts, intestinal T-cells from non-inflamed regions of IBD colon displayed enhanced responsiveness to IFN-1, increased frequency of pSTAT1+ cells, and greater induction of ISGs upon IFN-1 exposure in vitro. In healthy tissue, antibody neutralization of IFNß selectively reduced T-cell production of the pro-regulatory cytokine interleukin-10 (IL-10) and increased IFNγ synthesis. In contrast, neutralization of IFNß in IBD tissue cultures increased the frequency of T-cells producing inflammatory cytokines but did not alter IL-10 expression. These data support a role for endogenous IFN-1 as a context-dependent modulator of T-cell function that promotes regulatory activity in healthy human intestine, but indicate that the IFN-1/STAT1 pathway is dysregulated in inflammatory bowel disease.

Respiratory tract dendritic cells in paediatric asthma.

BACKGROUND: Airway dendritic cells (DC) are critical mediators of lung inflammation in asthma, but the characteristics of DC in the airways of healthy children, and children with asthma, are currently unknown. OBJECTIVE: We sought to identify changes in DC subset distribution and activation profile in paediatric asthma using flow cytometry to analyse induced sputum samples obtained from healthy and asthmatic children. METHODS: Lung function and atopic status were determined by spirometry and skin prick testing. Induced sputum samples were analysed using 7-colour flow cytometry to identify airway DC populations (lineage(-) HLA-DR(+) sputum cells expressing either CD11c as conventional DC or CD123 as plasmacytoid DC). RESULTS: Sputum samples containing lower airway plugs were obtained from 10 healthy children and 8 children with asthma. Lineage(-) HLA-DR(+) DC were successfully identified in all samples, and DC comprised a significantly higher proportion of sputum cells in children with asthma compared with age-matched healthy controls (1.29% vs. 0.67%, P = 0.02). DC expression of the costimulatory marker CD86 was significantly reduced in asthmatic children (73.4% vs. 59.7%, P = 0.04). Sputum DC also included numerous CD1c(+) cells (mean 57% of the total DC population) and low frequencies of cells expressing the subset markers CD141 or CD123, although the proportions of these did not differ between groups. CONCLUSIONS: Airway DC can be identified and characterized non-invasively using flow cytometry to analyse paediatric sputum samples. Our data reveal that children with steroid-treated asthma exhibit increased frequency of airway DC with reduced expression of the costimulatory marker CD86, suggesting altered trafficking and/or maturation of these cells either due to asthma or steroid therapies.

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.

Relationship between human intestinal dendritic cells, gut microbiota, and disease activity in Crohn's disease.

BACKGROUND: Altered intestinal dendritic cell (DC) function underlies dysregulated T-cell responses to bacteria in Crohn's disease (CD) but it is unclear whether composition of the intestinal microbiota impacts local DC function. We assessed the relationship between DC function with disease activity and intestinal microbiota in patients with CD. METHODS: Surface expression of Toll-like receptor (TLR)-2, TLR-4, and spontaneous intracellular interleukin (IL)-10, IL-12p40, IL-6 production by freshly isolated DC were analyzed by multicolor flow cytometry of cells extracted from rectal tissue of 10 controls and 28 CD patients. Myeloid DC were identified as CD11c(+) HLA-DR(+lin-/dim) cells (lin = anti-CD3, CD14, CD16, CD19, CD34). Intestinal microbiota were analyzed by fluorescent in situ hybridization of fecal samples with oligonucleotide probes targeting 16S rRNA of bifidobacteria, bacteroides-prevotella, C. coccoides-E. rectale, and Faecalibacterium prausnitzii. RESULTS: DC from CD produced higher amounts of IL-12p40 and IL-6 than control DC. IL-6(+) DC were associated with the CD Activity Index (r = 0.425; P = 0.024) and serum C-reactive protein (CRP) (r = 0.643; P = 0.004). DC expression of TLR-4 correlated with disease activity. IL-12p40(+) DC correlated with ratio of bacteroides: bifidobacteria (r = 0.535, P = 0.003). IL-10(+) DC correlated with bifidobacteria, and IL-6(+) DC correlated negatively with F. prausnitzii (r = -0.50; P = 0.008). The amount of TLR-4 on DC correlated negatively with the concentration of F. prausnitzii. CONCLUSIONS: IL-6 production by intestinal DC is increased in CD and correlates with disease activity and CRP. Bacterially driven local IL-6 production by intestinal DC may overcome regulatory activity, resulting in unopposed effector function and tissue damage. Intestinal DC function may be influenced by the composition of the commensal microbiota.

Intestinal dendritic cells: their role in bacterial recognition, lymphocyte homing, and intestinal inflammation.

Dendritic cells (DCs) play a key role in discriminating between commensal microorganisms and potentially harmful pathogens and in maintaining the balance between tolerance and active immunity. The regulatory role of DC is of particular importance in the gut where the immune system lies in intimate contact with the highly antigenic external environment. Intestinal DC constantly survey the luminal microenvironment. They act as sentinels, acquiring antigens in peripheral tissues before migrating to secondary lymphoid organs to activate naive T cells. They are also sensors, responding to a spectrum of environmental cues by extensive differentiation or maturation. Recent studies have begun to elucidate mechanisms for functional specializations of DC in the intestine that may include the involvement of retinoic acid and transforming growth factor-ß. Specialized CD103(+) intestinal DC can promote the differentiation of Foxp3(+) regulatory T cells via a retinoic acid-dependent process. Different DC outcomes are, in part, influenced by their exposure to microbial stimuli. Evidence is also emerging of the close interaction between bacteria, epithelial cells, and DC in the maintenance of intestinal immune homeostasis. Here we review recent advances of functionally specialized intestinal DC and their mechanisms of antigen uptake and recognition. We also discuss the interaction of DC with intestinal microbiota and their ability to orchestrate protective immunity and immune tolerance in the host. Lastly, we describe how DC functions are altered in intestinal inflammation and their emerging potential as a therapeutic target in inflammatory bowel disease.

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.

Mechanisms of action of probiotics: recent advances.

The intestinal microbiota plays a fundamental role in maintaining immune homeostasis. In controlled clinical trials probiotic bacteria have demonstrated a benefit in treating gastrointestinal diseases, including infectious diarrhea in children, recurrent Clostridium difficile-induced infection, and some inflammatory bowel diseases. This evidence has led to the proof of principle that probiotic bacteria can be used as a therapeutic strategy to ameliorate human diseases. The precise mechanisms influencing the crosstalk between the microbe and the host remain unclear but there is growing evidence to suggest that the functioning of the immune system at both a systemic and a mucosal level can be modulated by bacteria in the gut. Recent compelling evidence has demonstrated that manipulating the microbiota can influence the host. Several new mechanisms by which probiotics exert their beneficial effects have been identified and it is now clear that significant differences exist between different probiotic bacterial species and strains; organisms need to be selected in a more rational manner to treat disease. Mechanisms contributing to altered immune function in vivo induced by probiotic bacteria may include modulation of the microbiota itself, improved barrier function with consequent reduction in immune exposure to microbiota, and direct effects of bacteria on different epithelial and immune cell types. These effects are discussed with an emphasis on those organisms that have been used to treat human inflammatory bowel diseases in controlled clinical trials.

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.

Kinetics of the immune response to the (F1+V) vaccine in models of bubonic and pneumonic plague.

Protection against aerosol challenge with > 300 MLD of Yersinia pestis was observed 7 days after a single immunisation of mice with the F1+V vaccine. At day 60, mice were protected against injected challenge (10(7)MLD) in a vaccine dose-related manner. Recall responses to rV in splenocytes ex vivo at day 98 correlated significantly (p<0.001) with the immunising dose-level of V antigen; no memory response or anti-V serum IgG was detected in killed whole cell vaccine (KWCV) recipients. This may explain the susceptibility of KWCV recipients to aerosol challenge and the enhanced protection conferred by the F1+V sub-unit vaccine, particularly since the anti-F1 responses induced by either vaccine were similarly IgG1-polarised.

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.

Production of interleukin (IL)-10 and IL-12 by murine colonic dendritic cells in response to microbial stimuli.

Intestinal dendritic cells (DC) are likely to regulate immunity to gut microflora, but little is known about their responses to bacterial antigens. Therefore, DC from normal murine colon were characterized and their cytokine responses to components of Gram-negative and/or Gram-positive bacteria assessed. Cells were obtained by digestion of colonic tissue and contained DC that were identified by flow cytometry as CD11c(+) major histocompatibility complex (MHC) class II(+) cells. Purified DC were obtained by immunomagnetic separation plus cell sorting. DC had the morphology of immature myeloid cells, were endocytically active, expressed low levels of co-stimulatory molecules and stimulated a weak allogeneic mixed leucocyte reaction. Analysis of flow cytometry data by a sensitive subtraction method allowed measurement of production of interleukin (IL)-12 and IL-10 by small numbers of gut DC by intracellular staining. Fewer than 5% of unstimulated DC produced either IL-10 or IL-12. IL-10 production was significantly up-regulated following stimulation with Bifidobacteria longum, but not after exposure to lipopolysaccharide (LPS) or Streptococcus faecium. In contrast, colonic DC produced IL-12 in response to both LPS and B.longum. Thus, colonic DC can produce both IL-12 and IL-10 following bacterial stimulation. Cell wall components from different bacteria stimulate distinct responses and may direct immune responses differentially in the gut.

Mouse model characterisation for anthrax vaccine development: comparison of one inbred and one outbred mouse strain.

In order to evaluate the immunogenicity and protective efficacy of anthrax vaccine candidates a suitable small animal model is required. The inbred A/J strain of mouse has been selected as a potential model, and its immune response to immunisation with recombinant protective antigen (rPA) vaccine characterised, by assessment of rPA specific antibody production, and protection against injected challenge, with the unencapsulated STI strain of Bacillus anthracis. Studies were conducted to determine the time required post immunisation to develop a protective immune response, to define the minimum protective dose of vaccine required and to assess the long-term immune response to immunisation. From the results of these studies it was possible to establish that the A/J mouse is a consistent and robust small animal model for rPA vaccine testing. A comparison of the immune response to rPA vaccine immunisation in the Turner Outbred (TO) mouse strain was also conducted. Both inbred and outbred mouse strains displayed a predominantly Th2 biased immune response and showed a comparable antibody response to rPA immunisation. An assessment of protection in the TO mouse against aerosol challenge with the fully virulent strain of B. anthracis, Ames, was also made.

Efficacy of the latest fluoroquinolones against experimental Yersinia pestis.

The efficacies of prophylactic and therapeutic gatifloxacin and moxifloxacin were assessed in a BALB/c mouse model of systemic and pneumonic plague and compared with ciprofloxacin. Mice were given 100 mg/kg of the antibiotic by oral administration twice daily for 7 days starting 1h prior to infection or following infection. All antibiotics offered full protection for up to 6h following systemic challenge, and for up to 30 h following an aerosol challenge. The efficacy of each of the antibiotics decreased when antibiotics were started 18 h following systemic challenge and 48 h following aerosol challenge. Fluoroquinolones may therefore be considered useful candidates for the treatment of bubonic and pneumonic plague.

Modulation of human dendritic cell phenotype and function by probiotic bacteria.

BACKGROUND: "Probiotic" bacteria are effective in treating some inflammatory bowel diseases. However which bacteria confer benefit and mechanisms of action remain poorly defined. Dendritic cells, which are pivotal in early bacterial recognition, tolerance induction, and shaping of T cell responses, may be central in mediating the effects of these bacteria. AIMS: To assess effects of different probiotic bacteria on dendritic cell function. METHODS: Human intestinal lamina propria mononuclear cells, whole blood, or an enriched blood dendritic cell population were cultured with cell wall components of the eight bacterial strains in the probiotic preparation VSL#3 (four lactobacilli, three bifidobacteria, and one streptococcal strains). Dendritic cells were identified and changes in dendritic cell maturation/costimulatory markers and cytokine production in response to probiotic bacteria were analysed by multicolour flow cytometry, in addition to subsequent effects on T cell polarisation. RESULTS: VSL#3 was a potent inducer of IL-10 by dendritic cells from blood and intestinal tissue, and inhibited generation of Th1 cells. Individual strains within VSL#3 displayed distinct immunomodulatory effects on dendritic cells; the most marked anti-inflammatory effects were produced by bifidobacteria strains which upregulated IL-10 production by dendritic cells, decreased expression of the costimulatory molecule CD80, and decreased interferon-gamma production by T cells. VSL#3 diminished proinflammatory effects of LPS by decreasing LPS induced production of IL-12 while maintaining IL-10 production. CONCLUSIONS: Probiotic bacteria differ in their immunomodulatory activity and influence polarisation of immune responses at the earliest stage of antigen presentation by dendritic cells.

Prospective evaluation of intestinal homing memory T cells in ulcerative colitis.

BACKGROUND: Intestinal homing (beta7+) memory T cells reflect the mucosal environment in which they were primed. We hypothesized that prospective assessment of cytokine production by intestinal homing (beta7+) memory T cells in ulcerative colitis patients followed from remission to early relapse may elucidate shifts in cytokine production relevant to the mucosal environment associated with the early phase of inflammation. METHODS: Twelve patients with frequently relapsing ulcerative colitis (> or = 2 relapses in the previous 12 months) were recruited in remission and followed prospectively until relapse. Antibody labeling of whole blood and flow cytometry were used to identify beta7+ cells and beta7- populations within CD3+CD45RA- leukocytes. Production of cytokines (IFN-gamma, TNF-alpha, IL-2, IL-10, TGF-beta, and IL-4) was determined by intracellular labeling. RESULTS: Early relapse of ulcerative colitis was associated with a shift of T cells from the naive to the memory T cell pool, and further the ratio of beta7+:beta7- memory T cells was significantly reduced at relapse (p < 0.01). A greater proportion of intestinal homing beta7+ memory T cells produced IL-4 (p < 0.02) and TNF-alpha (p < 0.05) at disease relapse compared with remission. Non-intestinal homing beta7- memory T cells also showed a tendency toward an increased production of TH1 and TH2 cytokines. CONCLUSIONS: The earliest phase of intestinal inflammation in ulcerative colitis patients is associated with an increase in both TH1 (TNF-alpha and TH2 (IL-4) cytokines by intestinal homing beta7+ memory T cells. These data support the principles of targeting lymphocyte trafficking as therapies in ulcerative colitis.

Quantitative and functional characteristics of intestinal-homing memory T cells: analysis of Crohn's disease patients and healthy controls.

Circulating memory T cells can be subdivided on the basis of beta7 integrin expression. The beta7+ population contains cells primed in the intestine capable of homing back to the gut. We hypothesized that cytokine production by beta7+ memory T cells reflects the specialized mucosal compartment in which they were primed. Flow cytometry of whole blood was used to assess numbers of beta7+ (beta7hi and beta7int) and beta7- memory T cells and their production of Th1 and regulatory cytokines in healthy controls and Crohn's disease patients. In controls, beta7+ and beta7- memory T cells displayed a similar qualitative profile of cytokine production but the beta7+ population was enriched for cytokine-producing effector cells. In addition, the beta7hi population contained more cytokine-producing cells than the beta7int population, suggesting a gradient of cytokine production based on beta7 integrin expression. In active Crohn's disease, there was altered expression of beta7 integrin with a decrease in intestinal-homing memory T cells and an increase in systemic memory T cells. Furthermore, there was a selective loss of IL-10 and increase in TGF-beta in both beta7+ and beta7- memory T cell subsets which may contribute to the pathogenesis of the inflammatory process in Crohn's disease.

The dendritic cell: its role in intestinal inflammation and relationship with gut bacteria.

Dendritic cells are antigen presenting cells that are likely to be pivotal in the balance between tolerance and active immunity to commensal microorganisms that is fundamental to inflammatory conditions, including Crohn's disease and ulcerative colitis. Interactions between dendritic cells and microbial products are discussed and how they contribute to regulation of immune responses. The concept that interactions between dendritic cells and commensal organisms may be responsible for maintaining intestinal immune homeostasis is also explored.

The role of the gut flora in health and disease, and its modification as therapy.

The gut flora is a vast interior ecosystem whose nature is only beginning to be unravelled, due to the emergence of sophisticated molecular tools. Techniques such as 16S ribosomal RNA analysis, polymerase chain reaction amplification and the use of DNA microarrays now facilitate rapid identification and characterization of species resistant to conventional culture and possibly unknown species. Life-long cross-talk between the host and the gut flora determines whether health is maintained or disease intervenes. An understanding of these bacteria-bacteria and bacteria-host immune and epithelial cell interactions is likely to lead to a greater insight into disease pathogenesis. Studies of single organism-epithelial interactions have revealed the large range of metabolic processes that gut bacteria may influence. In inflammatory bowel diseases, bacteria drive the inflammatory process, and genetic predisposition to disease identified to date, such as the recently described NOD2/CARD15 gene variants, may relate to altered bacterial recognition. Extra-intestinal disorders, such as atopy and arthritis, may also have an altered gut milieu as their basis. Clinical evidence is emerging that the modification of this internal environment, using either antibiotics or probiotic bacteria, is beneficial in preventing and treating disease. This natural and apparently safe approach holds great appeal.