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Regulatory T cells express the glucocorticoid-induced tumour necrosis factor receptor family related gene (GITR) and antibodies against this receptor block regulatory cell activity. Signaling through this receptor may be involved in diseases where normal immune tolerance is broken, such as inflammatory bowel disease or autoimmune gastritis. The 1970s and early 1980s were the heyday of cellular immunology. Complicated circuits of T cells were constructed in which helper cells were inhibited by suppressor cells which in turn were inhibited by contrasuppressor cells, and so on. By and large the actual experiments involved mixing different populations of T cells and B cells together in vitro and measuring the antibody response to the major human pathogens, sheep erythrocytes, or haptenated synthetic polymers. If the response went down, suppressor cell activity was evident. Suppressor cells were considered to be a lineage of CD8+ T cells, so that CD8 cells were either cytotoxic cells or suppressor cells. In the early to mid 1980s however, suppressor T cell research fell dramatically out of fashion. As molecular immunology took over, cellular immunology was seen as unsophisticated black magic. CD4+ helper T cells were cloned, as were cytotoxic CD8 T cell clones, but no suppressor T cell clones were made. In fact cellular immunology became a term of abuse. For those of us who teach immunology to medical students the existence of suppressor T cells was an embarrassment, dealt with by the phrase “nobody believes in these cells anymore”. Shimizu J, Yamazaki S, Takahashi T, et al . Stimulation of CD25(+)CD4(+) regulatory T cells through GITR breaks immunological self-tolerance. Nat Immunol2002;3:135–42. [OpenUrl][1][CrossRef][2][PubMed][3][Web of Science][4] CD25(+)CD4(+) regulatory T cells in normal animals are engaged in the maintenance of immunological self-tolerance. We show here that glucocorticoid-induced tumor necrosis factor receptor family-related gene (GITR, also known as TNFRSF18)—a … [1]: {openurl}?query=rft.jtitle%253DNature%2Bimmunology%26rft.stitle%253DNat%2BImmunol%26rft.aulast%253DShimizu%26rft.auinit1%253DJ.%26rft.volume%253D3%26rft.issue%253D2%26rft.spage%253D135%26rft.epage%253D142%26rft.atitle%253DStimulation%2Bof%2BCD25%2528%252B%2529CD4%2528%252B%2529%2Bregulatory%2BT%2Bcells%2Bthrough%2BGITR%2Bbreaks%2Bimmunological%2Bself-tolerance.%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fni759%26rft_id%253Dinfo%253Apmid%252F11812990%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx [2]: /lookup/external-ref?access_num=10.1038/ni759&link_type=DOI [3]: /lookup/external-ref?access_num=11812990&link_type=MED&atom=%2Fgutjnl%2F51%2F3%2F311.atom [4]: /lookup/external-ref?access_num=000173562400013&link_type=ISI

Corrigendum: The endogenous cannabinoid system in the gut of patients with inflammatory bowel disease

(WO) in LPS-free culture medium in the presence or absence of a-TOC (1 μg/ml) or NAC (5mM). Muscular morphofunctional properties were evaluated by scanning micrometry and NFkB activation by phosphorylation of the p65 NFkB subunits Ser468 and Ser536. Data are expressed as mean±SE, p

levels of IL-4, IL-17A and IFN-gamma compared to CD samples with no endoscopic recurrence and normal controls. In the same mucosal lesions, TNF-alpha and IL-5 content was higher than that seen in normal controls but unchanged as compared to areas with no endoscopic recurrence. Conclusions: Data indicate that, in contrast to “late” CD, early recurrence is characterized by accumulation of various Th cell subsets, which make distinct profiles of inflammatory cytokines. Notably, even in the absence of endoscopic recurrence, CD mucosa is heavily infiltrated with cytokine-secreting inflammatory cells.

Background & Aims The pathogenesis of inflammatory bowel disease (IBD) is believed to involve an altered balance between effector and regulatory T cells. Aryl hydrocarbon receptor (AhR), a ligand-dependent transcription factor that mediates the toxicity of dioxins, controls T-cell responses. We investigated the role of AhR in inflammation and pathogenesis of IBD in humans and mouse models. Methods AhR expression was evaluated in intestinal tissue samples from patients with IBD and controls by real-time polymerase chain reaction (PCR) and flow cytometry. Intestinal lamina propria mononuclear cells (LPMCs) were activated in the presence or absence of the AhR agonist 6-formylindolo(3, 2-b)carbazole (Ficz). Colitis was induced in mice using trinitrobenzene sulfonic acid (TNBS), dextran sulfate sodium (DSS), or T-cell transfer. Mice were given injections of Ficz or the AhR antagonist 2-metyl-2H-pyrazole-3-carboxylic acid; some mice first received injections of a blocking antibody against interleukin (IL)-22. Cytokines were quantified by real-time PCR and flow cytometry. Results Intestine tissue from patients with IBD expressed significantly less AhR than controls. In LPMCs from patients with IBD, incubation with Ficz reduced levels of interferon gamma (IFN)-γ and up-regulated IL-22. Mice injected with Ficz were protected against TNBS-, DSS-, and T-cell transfer–induced colitis; they had marked down-regulation of inflammatory cytokines and induction of IL-22. Mice given AhR antagonist produced more inflammatory cytokines and less IL-22 and developed a severe colitis. Neutralization of endogenous IL-22 disrupted the protective effect of Ficz on TNBS-induced colitis. Conclusions AhR is down-regulated in intestinal tissue of patients with IBD; AhR signaling, via IL-22, inhibits inflammation and colitis in the gastrointestinal tract of mice. AhR-related compounds might be developed to treat patients with IBDs.

In a fetal intestinal organ culture model of intestinal inflammation, activation of T cells by a lectin, superantigen or anti-CD3 antibodies induces E-selectin expression which peaks 6-10 h after activation and disappears by 24 h. We now show that addition of cyclosporin A (Cys A) increases the number of E-selectin-expressing endothelial cells and the intensity of expression, and it prolongs expression up to at least 60 h after stimulation. The dynamics of E-selectin expression on tumor necrosis factor activated human umbilical-vein endothelial cell cultures is not affected by Cys A. Furthermore, other immunosuppressive agents FK 506 and dexamethasone inhibit E-selectin expression, indicating that the enhancement observed in the presence of Cys A is not a consequence of general immunosuppression.

AIMS--To determine the concentrations of interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF alpha) in stools from children. METHODS--Stool samples from 14 healthy children, 32 children with inflammatory bowel disease, and 23 children with acute diarrhoea were emulsified in an equal volume of phosphate buffered saline and then centrifuged to produce a clear supernatant fluid. IL-6 and TNF alpha were measured by enzyme linked immunosorbent assay (ELISA). RESULTS--TNF alpha was detected in the stools of all 14 healthy children (12-130 pg/g stool), but IL-6 was detected only in three. Similar results were seen in children with inactive inflammatory bowel disease. Stool TNF alpha concentrations were raised in samples from children with active inflammatory bowel disease, but in most (11/18) of these samples IL-6 was undetectable. Stool samples contained a heat-labile factor which rapidly destroyed IL-6 immunoreactivity. Most children with diarrhoea had TNF alpha concentrations similar to those of healthy controls and most were also negative for IL-6. Three children with Shigella flexneri infection had extraordinarily high concentrations of both TNF alpha and IL-6 in their stools. CONCLUSIONS--There is constant low grade production of TNF alpha in the intestine of healthy people. Raised values are indicative of mucosal inflammation, but are not specific. Stool IL-6 is of little use in assessing mucosal inflammation because immunoreactivity is rapidly lost in stool samples.

This study uses Helicobacter felis infection of C57BL/6 mice as a model of Helicobacter pylori infection in humans. Infection with H felis produces gastritis and gastric atrophy with mucous hyperplasia, metaplasia, and parietal cell loss. However, in mice preinfected with a nematode parasite Heligmosomoides polygyrus there was reduced parietal cell loss and less mucous cell hypertrophy and metaplasia. Measurement of gastric mucosal cytokines revealed that preinfection with H polygyrus reduced the number of transcripts for Th1 associated cytokines and chemokines but increased transcripts for Th2-type cytokines. The authors suggest that concurrent Th2 immune responses elicited by the nematode downregulate the tissue damaging Th1 response elicited by H felis . In addition, the authors speculate that different rates of helminthic infection in different parts of the world might explain why, despite equally poor socioeconomic conditions and high rates of infection with H pylori , there are major differences in the incidence of gastric cancer. However, before gastroenterologists recommend sending all their patients on a tropical holiday and suggest they eat all the unwashed and uncooked vegetables they can find, a few words of caution are needed. While this is an intriguing study, it is fair to say that the authors highlighted the positive aspects of their results and did not dwell on the negative aspects. The mice were studied at two time points, eight and 16 weeks after H felis infection. …

DIFFERENTIAL DEGRADATION OF ANTI-TUMOR NECROSIS FACTOR-ALPHA AGENTS BYMATRIXMETALLOPROTEINASE-12 IN INFLAMMATORY BOWEL DISEASE P. Biancheri∗ ,1, A. Di Sabatino2 , L. Rovedatti 2 , N. Ahmad1, G. Corazza2 , T. MacDonald1 1Centre for Immunology and Infectious Disease, BICMS, Barts and the London School of Medicine and Dentistry, London, UK; 2First Department of Medicine, S. Matteo Hospital, University of Pavia, Pavia

OBJECTIVE To directly infect human fetal intestine with HIV in vitro. DESIGN Human fetal intestinal explant cultures were exposed to HIV-1 and monitored for evidence of infection by biochemical assay, immunohistochemistry and in situ hybridization. METHODS Human fetal intestinal explants (14-21 weeks) were established in culture and exposed to HIV-1. Tissue culture fluid was assayed for p24 antigen and reverse transcriptase activity over a 14-day period. Explants were removed from culture on days 4, 7, 10 and 14 postinoculation and subjected (1) to immunohistochemistry to detect p24 and gp160/41 antigens, and (2) to in situ hybridization to detect HIV-1 RNA. Explant tissue culture fluid was cocultured with Jurkat T-cells to detect infectious viral particles. RESULTS Reverse transcriptase activity and p24 antigen levels in fetal explant culture fluid rose between 7 and 14 days after viral inoculation. Jurkat T-cell cocultures confirmed the presence of infectious virus. Cells in the lamina propria resembling lymphocytes and macrophages of both small intestine and colon stained positively for the viral proteins p24 and gp41. The same type of cells also stained positively for HIV-1 RNA using in situ hybridization. Dual-label immunohistochemistry, combined immunohistochemistry and in situ hybridization confirmed the presence of viral protein and RNA in cells bearing the CD3, CD4 (lymphocyte) or CD68 (macrophage) surface markers. There was no evidence at any time of HIV-1 infection of epithelial cells. CONCLUSIONS Cells of the lamina propria of the small intestine and colon, bearing lymphocyte or macrophage markers, can be directly infected by and support the replication of HIV-1. Such infection may be implicated in the pathogenesis of HIV enteropathy.

An organ culture model has been used to study the effects of T cell activation in the human colon. Lamina propria T cells in explant cultures of human fetal colon (11 to 23 weeks gestation) were activated in situ using pokeweed mitogen or an anti-CD3 monoclonal antibody, and compared with unstimulated controls. After three days of culture, there was a two to four-fold increase in crypt epithelial cell proliferation in T cell stimulated explants of more than 15 weeks gestation, associated with a fall in crypt goblet cell numbers of up to 20-fold. By three days, the surface epithelium of stimulated explants appeared thin with loss of goblet cells, and by day 7, severe and extensive mucosal damage was observed by light and electron microscopy. These changes did not occur in control cultures and explants deficient in T cells (less than 16 weeks gestation), and were inhibited by cyclosporin A. These experiments indicate that the increase in epithelial cell proliferation and accompanying goblet cell depletion observed in colorectal crypts in chronic inflammatory bowel disease may be mediated by activated T cells.

Series Editor's Note. Preface. 1. The Cells and Tissues of the Gastrointestinal Tract J.L. Viney, T.T. MacDonald. 2. The Mucosal Immune System in Inflammatory Bowel Disease P. Bradtzaeg, T.S. Halstensen, L. Helgeland, K. Kett. 3. Coeliac Disease E. Savilahti. 4. Immunology of Gastrointestinal Food Allergy in Infancy and Early Childhood J.A. Walker-Smith. 5. Immunology of Intractable Diarrhoea B. Cuenod, N. Cerf-Bensussan. 6. Immunology of Intestinal Transplantation in Man and Animals N. Brousse, S. Sarnacki, D. Canioni, N. Cerf-Bensussan. 7. Intestinal Graft-Versus-Host Disease A.McI. Mowat. 8. In Vitro Enteropathy C.P. Braegger, T.T. MacDonald. 9. Molecular Basis of Cell Migration into Normal and Inflamed Gut M. Salmi, S. Jalkanen. 10. Intestinal Permeation of Molecules in Health and Disease I.S. Menzies, M.W. Turner. 11. Immunology of Gastrointestinal Lymphoma J. Spencer, P.G. Isaacson. 12. The Gut in HIV Infection M. Zeitz, R. Ullrich, E.-O. Riecken. 13. Immunity to Enterotoxin-Producing Bacteria A.-M. Svennerholm, J. Holmgren. 14. Host Responses to Helicobacter (Campylobacter) Pylori Infection R.V. Heatley. 15. The Immunology of Giardiasis M.F. Heyworth. 16. Intestinal Nematode Infections E.S. Cooper. Index.

Loss of an immunological tolerance towards non-pathogenic luminal antigens is a key element in the pathogenesis of inflammatory bowel disease. Regulatory T cells (Tregs) are crucially involved in the induction and maintenance of this tolerance. Mice kept under germ-free conditions fail to induce an immunological tolerance in the gut. Intestinal epithelial cells (IECs) stimulate CD4+ Tregs dependent on MHC II. The corresponding MHC II-related processing of luminal antigens was shown to occur in multivesicular bodies (MVB). We hypothesised that the lack of tolerance induction in germfree mice might rely on defects in the MHC II-associated antigen presentation by IECs. To unravel the basic processes of the immunological homeostasis in the gut, we studied the influence of the bacterial flora on trafficking of luminal antigens into MHC II-enriched compartments responsible for antigen processing and presentation. NMRI mice were kept germ-free or conventionally colonised. Bacteria were specified in feces alliquots using 16S rRNA analysis. The results were correlated with the RDP II data base. The endocytic route of luminally applied ovalbumin (OVA) in IECs and associations with the MHC II pathway were visualised using fluorescenceand electron-microscopy. Colonised mice revealed a physiological murine flora without detection of any pathogen. IECs of the jejunum, ileum and colon lacked expression of MHC II and invariant chain (Ii) in germ-free mice. After colonisation both molecules were expressed in IECs of the jejunum and ileum, but not within the colon. While MHC II was seen within the endocytic pathway and at the basolateral membranes, Ii expression was restricted to the endocytic pathway. The majority of both antigens concentrated in MVB. In germ-free mice luminally applied OVA was only faintly detected within compartments of the endocytic tract after periods of up to two hours. In contrast colonised mice showed a significant increased uptake of OVA throughout the endocytic route (p = 0.001). Of note OVA accumulated within MHC II-enriched MVB in these mice. The bacterial colonisation of the gut seems to be essential for the MHC II-dependent antigen presentation by IECs. The physiological flora is responsible for the constitutive expression of MHC II and related molecules in IECs and a sufficient antigen delivery in processing-relevant compartments. Our data indicate that defects in the MHC II-associated antigen handling in IECs might contribute to the failure of germ-free mice in tolerance induction and support a regulatory function of these cells in the intestinal homeostasis. P294 Control of interleukin-17 production in inflammatory bowel disease L. Rovedatti1 *, A. Di Sabatino1, T. Kudo2, M. Sarra3, D. Rampton4, C.H. Knowles5, N. Sengupta5, G. Monteleone3, G.R. Corazza1, T.T. MacDonald2. 1First Department of Medicine, S. Matteo Hospital, University of Pavia, Pavia, Italy, 2Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, London, United Kingdom, 3Department of Internal Medicine, University Tor Vergata, Rome, Italy, 4Centre for Gastroenterology, Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, London, United Kingdom, 5Centre for Academic Surgery, Institute of Cell and Molecular Science, Barts and the London School of Medicine and Dentistry, London, United Kingdom

Caecal biopsy specimens from Jamaican children with the Trichuris dysentery syndrome (TDS) and age matched Jamaican controls were investigated by immunohistochemistry and by light microscopy. Biopsy specimens from all children (with TDS and controls) showed a mild to moderate increase in inflammatory cells. Except in the vicinity of the worm, where the epithelium was flattened, there was no other epithelial abnormality. Compared with controls, children with TDS had increased IgM lamina propria plasma cells and decreased intraepithelial T cells. There was also an increase in crypt epithelial cell proliferation. Lamina propria T cells (both activated and non-activated) were no more common in children with the Trichuris syndrome than controls. Epithelial cell HLA-DR and VLA-1 expression (which are increased in other colitides) were the same in both groups. Despite the presence of large worm burdens and chronic dysentery, therefore, only minor changes were seen in the caecal mucosa of children with TDS.

In the 8-week period from 11 weeks to 19–20 weeks gestation human fetal intestine develops organised Peyer's patches and mucosal T cells in the lamina propria and epithelium (Table 2). T cell numbers are, however, low compared to post-natal intestine, and the Peyer's patches only contain primary B cell follicles. There is little information on changes which occur between 20 weeks gestation and birth. However, there is little further development of human gut-associated lymphoid tissue until the gut is exposed to food and bacterial antigens. Thus, there are no plasma cells in newborn human intestine. It would, therefore, appear that the mucosal immune system is functionally mature by 20 weeks gestation, but remains quiescent until birth.

The rate of crypt epithelial cell proliferation and mucosal morphology have been studied in vitro in explants of fetal human small intestine in organ culture in which a cell-mediated immune response has been elicited by stimulating lamina propria T cells with pokeweed mitogen or monoclonal anti-CD3 antibodies. Twelve hours after the addition of anti-CD3 or pokeweed mitogen, most lamina propria T cells expressed CD25. By 18 and 24 h after the addition of anti-CD3, there were significantly more crypt cells in the cell cycle than in controls, although villus height and crypt depth are the same in both groups. After 3 days, by dissecting microscopy, the villi appeared shorter in cultures in which the T cells were stimulated than in control cultures, and the mucosal surface was obscured by a layer of extruded enterocytes. Villus atrophy was confirmed by direct measurement of Feulgen-stained, microdissected villi. Crypts were longer in T-cell-stimulated cultures, and the rate of crypt epithelial cell proliferation measured by metaphase arrest was increased 10-fold. By electron microscopy, the microvilli and cellular morphology of the surface enterocytes were normal in T-cell-stimulated cultures. These experiments clearly show that a profound immune-mediated crypt epithelial cell hyperplasia can occur in the absence of damage to surface enterocytes.

Immunohistochemical techniques were used to investigate the epithelial expression of VLA-1 in inflammatory bowel disease in six patients with Crohn's disease, in four patients with ulcerative colitis, and in one patient with indeterminate colitis, and compared with that in the small intestine and colons of 10 normal controls. In normal small bowel VLA-1 was expressed on crypt epithelial cells and only weakly or not at all on surface epithelium. VLA-1 was again expressed weakly in normal colon, except in one case, a 1 year old child with diarrhoea but no histological abnormalities. In small and large intestine affected with Crohn's disease, ulcerative colitis, or indeterminate colitis, there was increased expression of VLA-1 on the basolateral aspects of crypt cells and de novo expression on surface epithelium. It is suggested that this is an adaptive response to prevent epithelial cell loss as a result of inflammation in the underlying lamina propria.

IL-2 receptor (CD25) bearing cells can be visualised by immunohistochemistry in the intestinal lamina propria of children with Crohn’s disease or ulcerative colitis, but not in control bowel. The CD25+ cells occur as aggregates in the lamina propria below the epithelium, and scattered throughout the lamina propria. Double staining revealed that the CD25+ cells in Crohn’s disease are 60–88% CD3+,CD4+,CD8-, indicating that they are T cells whereas in ulcerative colitis the CD25+ cells are mostly CD3-,CD4+,HLA-DR+, indicating that they are macrophages. Differential expression of CD25 on T cells and macrophages serves to distinguish the immunologic lesions in ulcerative colitis and Crohn’s disease.