• Nenhum resultado encontrado

The Immunological Basis Of Inflammatory Bowel Disease

N/A
N/A
Protected

Academic year: 2021

Share "The Immunological Basis Of Inflammatory Bowel Disease"

Copied!
13
0
0

Texto

(1)REVIEWS THE IMMUNOLOGICAL AND GENETIC BASIS OF INFLAMMATORY BOWEL DISEASE Gerd Bouma*‡ and Warren Strober* The inflammatory bowel diseases (IBDs), Crohn’s disease and ulcerative colitis, are chronic inflammatory disorders of the gastrointestinal tract. Enormous progress has been made recently in understanding the pathogenesis of these diseases. Through the study of patients and mouse models, it has emerged that Crohn’s disease is driven by the production of interleukin-12 (IL-12) and interferon-γ (IFN-γ), whereas ulcerative colitis is probably driven by the production of IL-13. A second area of progress is in the identification of specific genetic abnormalities that are responsible for disease. The most important finding is the identification of mutations in the gene that encodes NOD2 (nucleotide-binding oligomerization domain 2) protein in a subgroup of patients with Crohn’s disease. Here, we discuss these recent findings and the implications for therapy.. HAPTENATING AGENT. Small organic molecules that do not provoke an immune response by themselves, but do so when covalently attached to a carrier protein.. *The Mucosal Immunity Section, National Institutes of Health, Mucosal Immunity Section, Laboratory of Clinical Investigation, National Institutes of Allergy and Infectious Diseases, 10 Center Drive, Room 11N238, Bethesda, Maryland 20892, USA. ‡ The Laboratory of Immunogenetics, Vrije Universiteit Medical Center, Amsterdam, The Netherlands. Correspondence to W.S. e-mail: wstrober@niaid. nih.gov doi:10.1038/nri1132. Crohn’s disease and ulcerative colitis, together referred to as inflammatory bowel disease (IBD), lead to longterm and sometimes irreversible impairment of gastrointestinal structure and function1. Although Crohn’s disease and ulcerative colitis share many clinical and pathological characteristics, they also have some markedly different features, and there is now ample reason to believe that the main pathological processes in these two diseases are distinct (BOX 1). Despite recent advances in our understanding of IBD, important questions that concern the immunopathology and genetic basis of IBD remain unanswered. The present working hypothesis is that IBD results from an inappropriate and exaggerated mucosal immune response to normal constituents of the mucosal microflora that is, in part, genetically determined. Indeed, several recent findings from experimental models of inflammation of the intestine, as well as from patients with IBD, support this hypothesis. Pathogenesis of inflammatory bowel disease. An important advance in the study of IBD has been the discovery and subsequent analysis of a wide variety of mouse models of intestinal inflammation that resemble. NATURE REVIEWS | IMMUNOLOGY. IBD2,3. These models fit broadly into four groups: spontaneous colitis as a result of a naturally occurring genetic abnormality; spontaneous colitis that occurs in mice with particular genetic defects produced by either gene targeting or the introduction of a transgene; colitis induced by exposure to a HAPTENATING AGENT or another type of causative agent; and colitis induced by transferring T-cell populations lacking regulatory cells into a severely lymphopenic host that lacks endogenous regulatory cells (BOX 2). These models give rise to several important principles that are relevant to human IBD. The role of genetic factors. As can be seen in BOX 2, a large number of mice with different genetic defects develop spontaneous mucosal inflammation. So, the first principle is that entirely different genetic abnormalities can lead to similar clinical features of intestinal inflammation. The second principle is that the host genetic background determines the susceptibility to intestinal inflammation, even when this is due to a major identified genetic defect. For example, although some inbred strains of mice with interleukin-10 (IL-10) deficiency are highly susceptible to colitis, others are resistant4. These differences in disease susceptibility and resistance among. VOLUME 3 | JULY 2003 | 5 2 1. © 2003 Nature Publishing Group.

(2) REVIEWS. Box 1 | What is inflammatory bowel disease?. General definition Inflammatory bowel disease (IBD) is a chronic relapsing idiopathic inflammation of the gastrointestinal tract. The two main forms of IBD — Crohn’s disease and ulcerative colitis — have many similarities, but there are also several clinical and pathological differences. In a small minority of cases that involve only the colon, they are indistinguishable and categorized as ‘indeterminate colitis’.. Epidemiology Both Crohn’s disease and ulcerative colitis have a prevalence range of 10–200 cases per 100,000 individuals in North America and Europe. Disease incidence is the highest in developed, urbanized countries. The incidence of Crohn’s disease has increased during the past four decades, whereas no clear trend is identifiable for ulcerative colitis.. Areas of involvement Crohn’s disease. Any part of the gastrointestinal tract can be affected, but most commonly, the terminal ileum, cecum, peri-anal area and colon. It is characterized by the presence of segments of normal bowel between affected regions, known as ‘skip’ lesions. The intersection of linear ulcers with islands of normal or oedematous mucosa might produce a ‘cobblestone’ appearance. Ulcerative colitis. the inflammatory process invariably involves the rectum and extends proximally in a continuous fashion, yet remains restricted to the colon. Sometimes, it is limited to the rectum as ‘ulcerative proctitis’.. Histology Crohn’s disease. A transmural (affecting all layers of the bowel wall), dense infiltration of lymphocytes and macrophages; presence of granulomas in up to 60% of patients; fissuring ulceration and submucosal fibrosis (see left-hand figure). Ulcerative colitis. Inflammation affects superficial (mucosal) layers with infiltration of lymphocytes and granulocytes and loss of goblet cells. Presence of ulcerations and crypt abscesses (see right-hand figure).. Clinical features and complications Crohn’s disease. Diarrhoea, pain, narrowing of the gut lumen leading to strictures and bowel obstruction, abscess formation, and fistulization to skin and internal organs. Ulcerative colitis. Severe diarrhoea, blood loss and progressive loss of peristaltic function leading to rigid colonic tube. In severe cases, this can lead to ‘toxic megacolon’ and perforation. Extra-intestinal inflammatory manifestations in joints, eyes, skin, mouth and liver can occur in both forms of IBD. Increased risk for colon carcinoma in longstanding IBD — in particular, ulcerative colitis.. Current treatments Crohn’s disease. 5-ASA compounds, corticosteroids, azathioprine/6-MP, methotrexate, antibodies specific for tumour-necrosis factor (TNF) and surgical removal of obstructing segments. Ulcerative colitis. 5-ASA compounds, corticosteroids, azathioprine/6-MP, intravenous cyclosporin and colectomy. Crohn's disease. Ulcerative colitis. (Left). Photomicrograph of a histological specimen taken from a patient with Crohn’s disease. Inflammation can be seen to involve the full thickness of the wall from the mucosa to the serosa. Granulomas are seen towards the serosal surface. (Right). Ulcerative colitis is shown microscopically here at low-power magnification to show mucosal inflammation and erosion. High-power magnification might show the presence of acute inflammatory cells in crypts, known as ‘crypt abscesses’. Images courtesy of Edward C. Klatt at Florida State University College of Medicine.. 522. | JULY 2003 | VOLUME 3. mouse strains offer the opportunity to identify mouse genes that regulate the expression of genes with an important role in disease, and might therefore contribute to the identification of genes that are necessary for the occurrence of disease in humans. Importance of the gut flora. In general, experimental colitis does not develop when mice are kept in a germ-free environment5. This leads to the third principle, that the normal mucosal microflora is required to initiate or maintain the inflammatory process, presumably by providing one or more antigens or costimulatory factors that drive the immune response in a genetically susceptible host. However, despite an extensive search, no specific pathogenic microorganisms have been associated with any of the models, so far. In addition, although it is known that antigens from most resident bacteria do not take part in the disease process, there is little evidence that pathogenic antigens come from a single organism, or even a restricted group of organisms6. It is possible to induce disease in various models of mucosal inflammation by the introduction of a single organism into an otherwise germ-free host7. Although this shows that mucosal inflammation can be caused by only a few antigens, it does not imply that only the organism introduced can cause disease in the model used or in any other model. Finally, not all members of the microflora are necessarily pathogenic in IBD. It has recently emerged that a class of microorganisms that are known collectively as ‘probiotics’ ameliorate, rather than induce, inflammation8. At present, they are thought to function through the induction of suppressor cytokines (see further discussion later). Despite the potential of antigens in the microflora to have pro- or anti-inflammatory effects, it is clear that these antigens are present in the body from birth and have some degree of access to the internal milieu. As a result, they might be subject to the intra-thymic processing that allows the immune system to distinguish self from non-self antigens, and, if so, could be considered to be equivalent to self-antigens9. On the basis of this, it could be argued that by reacting to these antigens the organism is mounting an autoimmune response and, by extension, the chronic mucosal inflammation of IBD could be thought of as an autoimmune disease. Effector T cells versus regulatory T cells. A fourth principle, and one that also relates to the role of the microflora in the induction of disease, arises from the fact that experimental inflammation occurs as a result of either excessive effector T-cell function or deficient regulatory T-cell function (FIG. 1). The former possibility is exemplified by models in which essential pro-inflammatory cytokines are overproduced. These include: mice in which tumour-necrosis factor (TNF) is overproduced due to a deletion in the AU-rich regulatory elements (ARE) (TNFARE mice); mice that are transgenic for signal tranducer and activator of transcription 4 (Stat4), which have exaggerated IL-12 signalling that leads to increased production of interferon-γ (IFN-γ). www.nature.com/reviews/immunol. © 2003 Nature Publishing Group.

(3) REVIEWS. Box 2 | Animal models of mucosal inflammation. Spontaneous colitis Cotton-top tamarin111, SAMP1/Yit mice112,113, C3H/HeJ/Bir mice114. Colitis occurring as a consequence of targeted mutations or the introduction of a transgene Barrier dysfunction: Mdr1a-deficient mice115, N-cadherin dominant negative mice28, Intestinal trefoil factor-deficient mice116 Regulatory cell defects: IL-2-deficient mice117, IL-2Rα-deficient mice118, IL-10-deficient mice16, CRFB4-deficient mice119, TGF-β-deficient mice120, TGF-βRII dominant-negative transgenic mice121, Tgε26 transgenic mice18 Increased effector-cell responses: Stat4 transgenic mice11, TNFARE mutant mice10, G-protein subunit αi2-deficient mice122, IL-7 transgenic mice123, TCRα-deficient mice124, NF-κB pathway disruption125−127, transfer of Hsp60-reactive CD8+ T cells136 Other or unknown: MHC class II-deficient mice124, HLA-B27 transgenic rats128, WASPdeficient mice129. cytokine, such as IL-10 knockout mice16 or mice with defective transforming-growth factor-β (TGF-β) signalling17. Finally, colitis occurs in mice in which regulatory T cells fail to develop properly. For example, Tgε26 mice (a line of CD3ε transgenic mice) have an aberrant thymic microenvironment. In these mice, a population of T cells develop that can induce severe colitis; this can be prevented by regulatory T cells that have undergone normal thymic development18. In addition to preventing nascent inflammation, it has recently been shown that regulatory T cells can reverse established inflammation. Administration of CD25+ regulatory T cells to SCID or recombinase-activating gene 1 (Rag1)-deficient mice cures colitis that is previously induced by the administration of CD45RBhi T cells19.. Colitis induced by exogenous agents Enema/intramural injection: TNBS130, oxazolone21, acetic acid131, peptidoglycan polysaccharide132 Oral: dextran sodium sulphate133, indomethacin134, carrageenan135. Colitis due to defective induction of regulatory cells CD4+CD45RBhi transfer into SCID or Rag-deficient mice14, bone-marrow transfer into Tgε26 mice18. ARE, AU-rich regulatory elements; CRF, cytokine receptor family; Hsp, heat-shock protein; IL, interleukin; Mdr; multidrug-resistance gene; NF-κB, nuclear factor-κB; Rag, recombinase-activating gene; SCID, severe combined immunodeficient; Stat, signal tranducer and activator of transcription; TCR, T-cell receptor; Tgε, CD3ε transgenic; TGF-β; transforming growth factor-β; TGF-βR, TGF-β receptor; TNBS, trinitrobenzene sulphonic acid; TNF, tumour-necrosis factor; WASP, Wiskott−Aldrich syndrome protein gene.. and TNF10,11; and trinitrobenzene sulphonic acid (TNBS)-induced colitis, in which excessive production of IL-12 in response to lipopolysaccharide (LPS) stimulation seems to be the crucial underlying genetic abnormality in susceptible mice (see further discussion later)12. The possibility that IBD is caused by defective T-cell-mediated regulation is indicated by the fact that most encounters with mucosal antigens do not induce effector cell responses, but instead induce negative regulatory T-cell responses, or anergy or deletion of antigen-specific T cells — a phenomenon known as ‘oral tolerance’13. As alluded to earlier, such oral tolerance occurs in response to the mucosal microflora and establishes a mucosal homeostasis that ensures that most mucosal responses are self-limited and do not result in inflammation. It follows that failure of this mechanism can be a cause of mucosal inflammation and indeed, a surprising number of mouse models of IBD result from a defect in regulatory T cells. A notable example is mucosal inflammation that is induced in a severe combined immunodeficient (SCID) mouse by the adoptive transfer of naive (CD45RBhi) T cells lacking regulatory cells, which is prevented by the co-transfer of mature (CD45RBlo) cells that contain a regulatory T-cell subpopulation14,15. In addition, intestinal inflammation occurs in models in which there is a clear deficiency in the production or function of a known regulatory. NATURE REVIEWS | IMMUNOLOGY. Two pathways to mucosal inflammation. The fifth principle is that, despite the wide variety of causes, mucosal inflammation is almost always mediated by one of two pathways: either an excessive T helper 1 (TH1)-cell response that is associated with increased secretion of IL-12, IFN-γ and/or TNF, or an excessive TH2-cell response that is associated with increased secretion of IL-4, IL-5 and/or IL-13 (REF. 3). This is shown by the differing cytokine responses in mucosal inflammation that is induced by the administration of haptenating agents TNBS and oxazolone (FIG. 2). TNBS induces an IL-12-mediated TH1-cell response (in SJL/J mice) that is characterized by transmural cellular infiltration that, in some cases, is associated with granulomas, and can be abrogated with antibodies specific for IL-12 (REF. 20). By contrast, oxazolone induces inflammation that is characterized by a more superficial cellular infiltrate that is associated with a greater disruption of the epithelial layer and, in some cases, a greater infiltration of neutrophils21. Recently, it has been shown that oxazolone-induced colitis results from the induction of natural-killer T (NKT) cells that produce IL-13, and that elimination of NKT cells or blockade of IL-13 — by an IL-13 receptor α2 (IL-13Rα2)–immunoglobulin fusion protein — prevents the development of colitis22. Furthermore, the production of IL-4 in oxazolone-induced colitis is rapidly superseded by the production of IL-13, and IL-13 production can be induced by the stimulation of NKT cells that express the invariant Vα14 T-cell receptor (TCR), which is expressed by most NKT cells22. This indicates that tissue damage in oxazolone-induced colitis results from the cytolytic activity of NKT cells towards epithelial cells or, alternatively, from the toxic effects of IL-13 on epithelial cells (FIG. 3). This dichotomy between TH1- and TH2-like mucosal inflammation is seen in human IBD in that the histopathological features of Crohn’s disease resemble those of experimental TH1-cell-mediated colitis, whereas those of ulcerative colitis are most similar to experimental TH2-cell-mediated colitis, and the cytokine patterns that are characteristic of these diseases are in accordance with these distinct mechanisms (see later discussion).. VOLUME 3 | JULY 2003 | 5 2 3. © 2003 Nature Publishing Group.

(4) REVIEWS. TOLL-LIKE RECEPTORS. (TLRs). A family of receptors that recognize conserved products that are unique to microorganisms, such as lipopolysaccharide. TLRmediated events signal to the host that a microbial pathogen is present.. Role of the mucosal epithelium. Recent work has established that mucosal epithelial cells can recognize microorganisms and/or substances that are produced by microorganisms in the mucosal lumen, at least in part, through their expression of TOLL-LIKE RECEPTORS (TLRs)23. So, under appropriate conditions, epithelial cells can react to these microorganisms with the production of cytokines, chemokines and other pro- and anti-inflammatory substances24,25. It is probable that this function of epithelial cells has an important role in mucosal host defence, and that they are also involved in the initiation and/or potentiation of mucosal inflammation and IBD26. It is already clear that these active functions of epithelial cells are crucial for the maintenance of their essential passive function — that is, the ability to function as a barrier to lumenal microorganisms that could otherwise stimulate chronic inflammation27. The effect of loss of this barrier function is clearly seen in mice that express a dominant-negative N-cadherin transgene in epithelial cells that disrupts intra-epithelial-cell adhesion28. These mice develop severe inflammation, particularly in areas of the intestine that lie beneath areas of barrier disruption. This model, and others that are attributable to loss of barrier function, give rise to a sixth principle that proposes that the epithelium of the mucosa has an essential role in the physical separation of potentially stimulating microflora and the reactive cells of mucosal immune system. A role for defects in innate immunity. A seventh and final principle of IBD pathogenesis to emerge from the study of mouse models is that mucosal inflammation can also be due to genetic abnormalities of innate immunity that involve the function of antigen-presenting cells (APCs), macrophages or NK cells. One example of this kind of defect is seen in mice with myeloid-specific Stat3 deficiency. These mice have a defect in their response to. Stat3-dependent cytokines, most notably IL-10, and have increased secretion of IL-6, IL-12 and TNF, and mucosal inflammation after challenge with LPS29. It remains to be seen whether mucosal inflammation can also be caused by excessive responses of macrophages mediated by stimulation through TLRs or other sensors of microbial components. This is not an unlikely scenario given the fact that Crohn’s disease in humans can result from the defective function of nucleotide-binding oligomerization domain 2 (NOD2) protein — a protein that senses bacterial peptidoglycans (see further discussion later). Effector T-cell abnormalities in patients. Although the mouse models that were referred to earlier have led to important insights into the pathogenesis of mucosal inflammation and the mechanisms that might underlie IBD, we must ultimately look at the patients themselves to gain an unequivocal understanding of the disease. As noted earlier, the models indicate that mucosal inflammation usually results from a TH1- or TH2-cell-mediated final pathway, which resemble Crohn’s disease and ulcerative colitis, respectively. So, what evidence is there to indicate that Crohn’s disease is due to TH1-mediated inflammation and ulcerative colitis is due to TH2-mediated inflammation? Cytokine responses in Crohn’s disease. The data that support that Crohn’s disease is a TH1-mediated inflammatory disease is quite substantial.Various immunohistological studies indicate that in situ IL-12 is overproduced by macrophages in Crohn’s disease, but not in ulcerative colitis30,31, and macrophages that are isolated from the inflammatory lesions of patients with Crohn’s disease produce increased amounts of IL-12 ex vivo32, whereas macrophages that are isolated from patients with ulcerative colitis produce decreased amounts of IL-12,. Effector cells (TH1 or TH2). Regulatory cells. Mucosal homeostasis. Mucosal inflammation. Mucosal inflammation. Figure 1 | Pathways to mucosal inflammation. Mouse models of mucosal inflammation can be divided into two categories with respect to effector versus regulatory T-cell balance. In the first type, effector-cell responses — either T helper 1 (TH1) or TH2-cell responses — are abnormally robust and the intrinsic homeostasis of the mucosal immune system is biased towards inflammation. Alternatively, effector-cell responses are normal, but regulatory-cell responses are weak; again, inflammation is the result. It is not clear, however, if human inflammatory bowel diease (IBD) is due to regulatory cell defects, because at present it is not possible to measure regulatory-cell responses with any degree of confidence.. 524. | JULY 2003 | VOLUME 3. www.nature.com/reviews/immunol. © 2003 Nature Publishing Group.

(5) REVIEWS. TH1-mediated inflammation such as TNBS colitis IL-12 (IL-23?). TH2-mediated inflammation such as oxazolone colitis IL-4 and other cytokines?. CD4+. NKT. IFN-γ TNF. IL-13. Crohn's-like disease. Ulcerative colitis-like disease. Figure 2 | TH1- and TH2-mediated colitis. Trinitrobenzene sulphonic acid (TNBS) and oxazalone, both classic skin-sensitizing agents, give rise to different forms of colitis when administered intrarectally together with ethanol — an agent that temporarily decreases the function of the epithelial barrier. These colitides are strain-specific and genetically determined. TNBS that is administered to SJL/J or C57BL/10 mice elicits a classical T helper 1 (TH1)-cell response owing to interleukin-12 (IL-12)-mediated production of interferon-γ (IFN-γ) by CD4+ T cells. The result is a transmural inflammation that resembles Crohn’s disease. By contrast, oxazalone induces a TH2-cell response in SJL/J and C57BL/10 mice that is mediated by naturalkiller T (NKT) cells that produce IL-13. The result is a superficial inflammation that resembles ulcerative colitis.. compared with those from normal tissues. In addition, nuclear extracts of T cells from the affected tissues of patients with Crohn’s disease contain increased amounts of activated STAT4 and the transcription factor T-bet, which is indicative of IL-12 signalling, and T cells isolated from these tissues express increased amounts of the IL-12Rβ2 chain — a characteristic of TH1 cells33,34. In view of these findings, it is not surprising that T cells isolated from the affected tissues of patients with Crohn’s disease or clones that are derived from such cells produce markedly increased amounts of IFN-γ together with markedly decreased amounts of IL-4, compared with controls31,35. This circumstantial evidence in support of the causal role of TH1-mediated processes in Crohn’s disease has recently obtained further, and perhaps definitive, support from the finding that patients treated with antibody specific for the p40 chain of IL-12 (IL-12p40) leads to a prompt and marked amelioration of inflammation in most patients. The clinical improvement is associated with a reduction in both the production of IL-12 and IFN-γ by mononuclear cells that are isolated from the affected tissues (P. Mannon, I. Fuss and W. S, unpublished observations). Recent mouse studies that show that the TH1inducing cytokine IL-23, which consists of the IL-12p40 chain linked to the IL-23p19 chain, not to the IL-12p35 chain, more effectively stimulates TH1 memory-cell NATURE REVIEWS | IMMUNOLOGY. responses than IL-12 does, at least in mouse systems. So, it is possible that IL-23 could be more important than IL-12 in sustaining the TH1-mediated inflammation in Crohn’s disease36. In support of this possibility, IL-23p19-deficient mice, which lack IL-23 but not IL-12, are highly resistant to the development of experimental autoimmune encephalomyelitis (EAE) — a TH1-mediated autoimmune disease37. Whether IL-23 has a crucial role in sustaining the TH1-mediated inflammation of Crohn’s disease remains to be seen. Cytokine responses in ulcerative colitis. Whereas there is abundant evidence that T cells from lesions in Crohn’s disease produce increased amounts of IFN-γ, the signature TH1 cytokine, there is no evidence that T cells from ulcerative colitis produce increased amounts of IL-4, the definitive TH2 cytokine. Despite this difficulty, there is, in fact, considerable evidence that ulcerative colitis is a TH2-mediated disease. First, ulcerative colitis, to a greater extent than Crohn’s disease, is associated with the production of various autoantibodies, such as antineutrophil cytoplasmic antibody (pANCA) and antitropomyosin38,39. As TH2 cells provide more efficient help in the activation of B cells and the induction of humoral immune responses than TH1 cells do, the presence of autoantibodies might be indicative of a TH2rather than a TH1-mediated immune response. Second, studies on the production of immunoglobulin subclasses show that subclasses that are associated with TH1-cell responses, such as IgG2 antibodies, are increased in Crohn’s disease and TH2-related subclasses, such as IgG1 and IgG4 antibodies, predominate in ulcerative colitis40. Third, although ulcerative colitis has not been associated with increased secretion of IL-4, it has been associated with increased secretion of IL-5, another TH2 cytokine35. In addition, the expression of Epstein–Barr-virus-induced gene 3 (EBI3), which encodes a product that has been tentatively identified as a TH2 cytokine, has been found to be increased in ulcerative colitis, but not in Crohn’s disease41. As mentioned, IL-13-producing NKT cells were shown to have an important role in the inflammation that occurs in oxazolone-induced colitis — the experimental colitis that resembles ulcerative colitis22. This has led to the study of the production of IL-13 in ulcerative colitis and indeed, it has been found that lamina propria cells from patients with ulcerative colitis, but not Crohn’s disease, produce markedly elevated amounts of this cytokine (I. Fuss, F. Heller and W. S., unpublished observations). This observation provides support for the hypothesis that, as in oxazolone-induced colitis, T cells that produce IL-13 mediate an atypical TH2inflammatory response that results in the disease. However, further studies in which the effect of IL-13 blockade on the clinical course of ulcerative colitis is assessed will be necessary to prove this point. A role for other effector cytokines? Other cytokines that do not necessary belong to either the TH1 or TH2 categories, such as IL-15, IL-16 and IL-18, are also produced at higher levels in Crohn’s disease and ulcerative. VOLUME 3 | JULY 2003 | 5 2 5. © 2003 Nature Publishing Group.

(6) REVIEWS colitis42−44. However, on the basis of animal models, in which disease is prevented or cured by antibodies specific for IL-12 or IL-13, it is not probable that any of the aforementioned cytokines have a pivotal role in disease pathogenesis. Additionally, IFN-γ and IL-4 affect downstream effector cells, such as macrophages, to induce the production of pro-inflammatory cytokines, including TNF, IL-1β and IL-6, which are integral to the pathological response. Whereas increases in the levels of the putative initiating cytokines, such as IFN-γ, are relatively modest (~3-fold), increases in the levels of the downstream proinflammatory cytokines are more marked (~10−20fold)45,46. This argues for the occurance of an as yet poorly understood multiplier effect in IBD inflammation, in which even small increases in initiating cytokines give rise to large increases in downstream cytokines. IBD and defective immunoregulation. So far, our discussion of the main immunological characteristics of IBD have focused on the effector-cell responses to mucosal antigens that induce inflammation. However, from the studies of the mouse models discussed earlier, we are now well aware that mucosal inflammation can result from either excessive effectorcell responses or deficient regulatory responses to these antigens. At this point, we consider the data that support the latter mechanism. Is oral tolerance defective in IBD? By referring to a potential regulatory defect in IBD, we are of course referring to a defect in oral tolerance — the immune mechanism that is now recognized to be due to the induction of clonal anergy or deletion of effector T cells, as well as the induction of suppressor cells, known as TH3 cells, which produce TGF-β and/or other suppressive cytokines13,47,48. Whether or not patients with IBD have an underlying defect in some aspect of oral tolerance that leads to disease has not been clearly defined, because functional studies to detect this defect in humans are difficult. In particular, they must be carried out prospectively, before the onset of active inflammation, as inflammation itself might affect the outcome. However, a nonprospective study of regulatory responses in Crohn’s disease has been carried out, in which the ability of patient and control T cells to proliferate and produce cytokines when cultured in vitro with extracts of their own microbial microflora — that is, with antigens to which a normal individual is tolerant — was evaluated. T cells from patients with IBD were shown to proliferate and produce cytokines in response to their own microfloral antigens, whereas cells from controls did not49. This important, but as yet uncorroborated, finding has been interpreted to indicate that a loss of tolerance to antigens in the mucosal microflora occurs in IBD. However, the results could also indicate excessive effector-cell responses to these antigens. So, further work will be required to assess the meaning and validity of this finding.. 526. | JULY 2003 | VOLUME 3. Defects in regulatory T cells. Another approach to identify the presence of a regulatory defect in IBD is to characterize the possible regulatory cells that are present in the lesions and peripheral tissues of patients during various phases of disease. In recent years, several kinds of regulatory cells have been described in both mice and humans, and it is useful at this junction to describe briefly these cells and to discuss how they might be related to IBD. The first regulatory cell to be discussed is a CD4+ T cell, first identified by Sakaguchi et al.50−52 by its constitutive expression of CD25 and, more recently, by its expression of glucocorticoid-induced TNF receptorrelated protein (GITR). The important characteristics of this cell are that it originates in the thymus and has specificity for self-antigens53. In addition, there is now evidence that it belongs to a separate T-cell lineage that expresses unique proteins, such as an intracellular protein known as FOXP3, the absence of which is the basis of severe autoimmune disease in both mice and humans54,55. The mechanism of the suppressor activity of CD4+CD25+ T cells is not yet completely defined. In in vitro assay systems, the suppression requires cell−cell contact and so, it was not thought to be due to a suppressor cytokine56. However, there is now considerable evidence in both mice and humans that after appropriate activation, CD4+CD25+ T cells express cell-surface TGF-β57, and with maximal stimulation, also secrete TGF-β57. An important role for cell-surface expression of TGF-β in the suppressor activity of CD25+ T cells in vitro is supported by the fact that antibodies specific for TGF-β, at least partially, block the suppressor function in this circumstance57,58. Parallel evidence that cell surface or (more probably) secreted TGF-β is involved in the in vivo suppressor effects of CD25+ T cells comes from studies of colitis induced by the transfer of CD45RBhi cells to SCID or Rag2-deficient mice, in which it can be shown that CD4+CD25+ T cells can substitute for CD45RBlow cells for preventing inflammation and the CD25+ T-cell effect can be abrogated by concomitant administration of TGF-β-specific antibody59. Despite these observations, the issue of whether TGF-β has a main role in CD25+ T-cell suppressor function remains unsettled, because it has been shown that CD25+ T cells from TGF-β1-deficient mice can still mediate suppressor activity in vitro60. This would argue that even if secretion of TGF-β is the main suppressor mechanism in normal mice, other mechanisms exist as well. If CD25+ T cells function through the production of TGF-β, this would agree with a possible role in oral tolerance, which, as mentioned, has been associated with the induction of TGF-β-producing TH3 cells. In fact, administration of oral antigen leads to the clonal expansion of CD25+ regulatory cells61,62. However, if these cells are crucial for the prevention of IBD, and if IBD results from the reactivity to antigens in the microbial microflora, how can we explain the fact that these cells are of thymic origin and are self-antigen reactive? One possibility is that antigens in the mucosal microflora gain entry to the internal milieu and are processed and. www.nature.com/reviews/immunol. © 2003 Nature Publishing Group.

(7) REVIEWS. Glycolipid antigen Gut Lumen Epithelium. CD1d Glycolipid Vα14 TCR NKT APC. NKT. NKT. IL-13. Figure 3 | NKT cells and IL-13 in oxazalone-induced colitis. During oxazalone-induced colitis, it has been proposed that antigen-presenting cells (APCs) take up glycolipid antigens that come from the mucosal environment and present these antigens to natural killer T (NKT) cells through CD1. The NKT cells are stimulated through invariant Vα14 T-cell receptors (TCRs), leading to the secretion of interleukin-13 (IL-13), which has a toxic effect on epithelial cells. NKT cells might also have direct cytotoxicity against epithelial cells through recognition of CD1associated glycolipid antigens expressed by epithelial cells. This mechanism might explain the targeting of epithelial cells in oxazalone-induced colitis (and perhaps in ulcerative colitis).. presented to developing regulatory T cells in the thymus. These T cells then exit the thymus and traffic to the mucosal tissues, in which they undergo re-stimulation and clonal expansion. So, according to this hypothesis, the regulatory T cells that seem to be induced during oral tolerance might actually be regulatory cells that develop in the thymus and are only re-stimulated in the mucosal tissues. Another possibility is that regulatory cells that develop in the thymus against true endogenous selfantigens, can crossreact with antigens in the mucosal microflora. Models of colitis (such as the Tgε26 model described earlier) show that regulatory cells that prevent colitis do require thymic maturation, but do not differentiate between these two possibilities18. At present, evidence that CD25+ T cells are involved in the pathogenesis of IBD is scarce. One study showed that T cells from patients with Crohn’s disease are unresponsive to TGF-β due to the presence of elevated levels of intracellular SMAD7 — a known repressor of TGF-β signalling63. Whether this is a primary or secondary abnormality is not yet known, but the latter is more probable as expression of SMAD7 is upregulated by cytokines that are produced in a TH1-cell response. A second, and seemingly distinct, population of regulatory cells are known as TR1 regulatory T cells, which suppress responses through the secretion of IL-10 (REFS 64,65). These cells differ from the CD4+CD25+ regulatory T cells that were discussed earlier, in that they do not constitutively express CD25 and do not seem to develop into regulatory cells66−68. Instead, they seem to. NATURE REVIEWS | IMMUNOLOGY. have differentiated from conventional CD4+ T cells under the influence of a particular cytokine environment, namely the presence of IL-10 and (in humans) IFN-α 64,65. These cells have been observed in vivo following experimental infection69,70; they might therefore have an important role in limiting inflammation due to infectious organisms. In the mucosal immune system, TR1 cells might also be part of the oral-tolerance reaction, as it has been shown that the Peyer’s patches are enriched for CD8α–CD11b+ dendritic cells that produce large amounts of IL-10 and small amounts of IL-12, and which induce IL-10-producing T cells71. In addition, it has been shown that an inhaled antigen, ovalbumin (OVA), is taken up by IL-10-producing dendritic cells, which also induce IL-10-producing T cells72. TR1 cells could function together with CD25+ T cells to prevent the excessive response to antigens in the mucosal microflora that might be the basis of IBD. However, the concept described earlier in which antigens in the mucosal microflora are, in effect, self-antigens argues against this possibility, as regulation of responses to self antigens is the responsibility of CD25+ T cells. This is not to say, however, that IL-10 itself (from sources other than TR1 cells) has no role in the regulation of mucosal inflammation. Indeed, there is good evidence that IL-10 is required for the induction of TGF-β secretion and signalling, and that lack of this cytokine is clearly associated with experimental colitis16,73. On this basis, the role of IL-10 in patients, whether derived from TR1 cells or not, is worthy of continued study. A third type of regulatory cell is a CD8+ T cell that is induced in human mucosal tissues as a result of interactions between antigen-presenting epithelial cells and T cells74. Such interactions might be unique in that they require binding of the CD8 molecule with gp180 antigen — a form of carcinoembryonic antigen75 — expressed by the epithelial cell. Although these cells are potentially homeostatic regulators, there is evidence that they are decreased in patients with IBD as a result of dysregulated expression of gp180 by epithelial cells76,77. Further work defining the antigen specificity of these cells and their mechanism of action would help to define their role in IBD. A final word about regulatory cells in IBD concerns the role and nature of TH3 cells in disease pathogenesis. Although it is possible that TH3 cells are a distinct class of regulatory cells, it seems more probable that they are comprised of both CD25+ cells (producing TGF-β) and TR1 cells (producing mostly IL-10). However, further work will be necessary to establish this possibility. Genetic factors in inflammatory bowel disease. Epidemiological and family studies have provided overwhelming evidence that genetic factors have an important role in determining susceptibility to IBD78,79. The most compelling evidence comes from studies that are conducted with twins. The rate of concordance for Crohn’s disease has been reported to be as high as 58% in identical twins, whereas the dizygotic-twin. VOLUME 3 | JULY 2003 | 5 2 7. © 2003 Nature Publishing Group.

(8) REVIEWS. Gut lumen. a. Bacteria Epithelium. NOD2 defect. Macrophage Normal path CD4+ Activated macrophage (infection free). b. Effector response and inflammation. Peptidoglycan Defensin production. NOD2 defect. Normal. Barrier function impaired. Epithelial cells activated. Effector response and inflammation. Chemokine secretion. c. Bacteria and/or bacterial products (peptidoglycan). APC. NOD2 defect CD4+. Normal path Activated APC Strong effector response or weak regulatory response and inflammation. Conditioned APC CD4+. Homeostatic effector and regulatory response. Figure 4 | Possible mechanisms of Crohn’s disease caused by NOD2 mutations. a | Defective function of macrophages leads to persistent intracellular infection of macrophages and chronic stimulation of T cells by macrophage-infecting organisms. b | Defective epithelial-cell responses lead to loss of barrier function and increased exposure to the mucosal microflora. c | Defective ‘conditioning’ of antigen-presenting cells (APCs) leads to inappropriate activation of APCs and disruption of the homeostatic balance of effector and regulatory cells. NOD, nucleotide-binding oligomerization domain.. 528. | JULY 2003 | VOLUME 3. concordance is not significantly different from that for all siblings80. Reported concordance rates for monozygotic and dizygotic twins with ulcerative colitis range from 6−17% and 0−5%, respectively80−82. Collectively, these observations strongly support the assumption that susceptibility to IBD, in particular Crohn’s disease, is inherited. However, it also indicates that IBD is not inherited as a Mendelian trait, but rather has a complex genetic basis with many contributing genes. The genetic complexity, including genetic heterogeneity, gene−gene and gene−environment interactions, has for a long time prevented the identification of specific genetic abnormalities. Recently, however, progress has been made in this area of research. We discuss a newly discovered gene that predisposes to Crohn’s disease in a subgroup of patients, as well as recent progress in identifying genetic factors in mouse models of IBD. Identification of NOD2 mutations in Crohn’s disease. Whereas the identification of mutations underlying monogenic disorders is now relatively straightforward, the identification of genes involved in complex genetic disorders is complicated. A recent successful approach to identify genes involved in susceptibility to IBD has been to probe the genomes of families with many affected members using polymorphic MICROSATELLITE MARKERS. These whole-genome screens are based on the principle that a marker that is located close to a disease gene is less likely to be separated during meiosis and will be coinherited with the disease gene. Affected relatives (for example, affected siblings) will share alleles in excess of statistical expectation for markers close to a disease susceptibility gene(s). The analysis of large groups of affected siblings, therefore, allows the identification of broad genomic regions that contain a disease gene(s). Hugot and colleagues83 used this approach to study families with patients with Crohn’s disease. In this study, as well as in subsequent independent studies, a susceptibility locus was detected in the pericentromeric region of chromosome 16, now known as IBD1. Further analysis of this region identified a strong association with a single gene, NOD2 — also known as caspase-recruitment domain protein 15 (CARD15)84 — which encodes an intracellular molecule of the NOD family that is thought to be involved in the recognition of bacteria85,86. Sequencing of this gene in patients with Crohn’s disease indicated a cytosine insertion at position 3020 in exon 11, that gives rise to a stop codon and a truncated NOD2 protein. Several other polymorphisms were identified in patients with Crohn’s disease, including two common missense mutations — 2722G→C [Gly908Arg] and 2104C→T [Arg702Trp]. In parallel studies, Ogura et al.87 applied the candidate-gene approach to search for genes in the IBD1 locus. The authors knew that NOD2 is involved in the recognition of intracellular pathogens and overlaps with the IBD1 locus, and sequencing of this gene in a group of patients with Crohn’s disease and controls indicated the presence of the 3020 frameshift mutation mentioned above.. www.nature.com/reviews/immunol. © 2003 Nature Publishing Group.

(9) REVIEWS NOD2 mutations and risk of IBD. The data from the studies of both Hugot et al. and Ogura et al. indicate that NOD2 mutations occur exclusively in patients with Crohn’s disease and not in patients with ulcerative colitis. From the data obtained by Hugot et al., the RELATIVE RISK for developing Crohn’s disease in simple heterozygotes is 3, whereas for homozygotes it is 38. In addition, compound heterozygotes — that is, individuals that have one mutation on one chromosome and one on the other — function as homozygotes, with a relative risk of 44. In a subsequent population-based study that was carried out by Hampe et al., it was shown that the 3020 insertion in the NOD2 gene is present both in familial and sporadic cases of Crohn’s disease and that homozygosity for this mutation was not found in normal individuals88. These data are consistent with the view that a simple recessive mutation in NOD2 might be sufficient for the development of disease, and it has been established that 8−17% of Caucasian patients with Crohn’s disease have two copies of the main risk alleles of NOD2 (REF. 78). However, normal individuals might occasionally have NOD2 mutations on both chromosomes in the absence of disease89 and none of the three mutations in the NOD2 gene was found in 483 Japanese patients with Crohn’s disease90. The latter provides strong evidence for the presence of genetic heterogeneity among patients of different ethnic groups.. MICROSATELLITE MARKERS. A class of polymorphic markers (di-, tri- or tetra-nucleotide repeats) that are scattered throughout the genome. Individuals differ in the number of repeats, which can be used in determining the position of disease-susceptibility genes in the genome by studying the cosegregation of these markers with disease in families. RELATIVE RISK. The probability of an event in the active group divided by the probability of the event in the control group. In this case, the risk of developing Crohn’s disease in homozygous or heterozygous carriers of the NOD2 mutation divided by the risk in non-carriers of the mutation.. Role of NOD2-gene variants in disease. NOD2 is now recognized as a member of a large family of intracellular proteins — the NOD family — that are widely distributed in nature and includes the resistance (R) proteins in plants that are involved in plant host defence against pathogens. Structurally, they consist of two amino-terminal effector domains, known as caspase-recruitment domains (CARDs), a central nucleotide-binding oligomerization domain (NOD) and multiple carboxyterminal leucine-rich repeat (LRR) domains that can function as an intracellular sensor of bacterial infection91,92. NOD2 protein is expressed by monocytes, granulocytes, dendritic cells and, notably, also by epithelial cells93. In vitro studies have shown that stimulation of NOD2-transfected cells with bacterial proteins results in activation of the nuclear factor-κB (NF-κB) pathway. Such activation occurs through RICK (RIP-like interacting CLARP kinase receptor interacting protein 2) — a serine/threonine kinase that phosphorylates inhibitor of NF-κB kinase (IKK) and, therefore, allows the transport of NF-κB to the nucleus87,94. Initially, LPS was thought to be the bacterial component that is recognized by NOD2 protein, but in recent work it has been shown that it is, in fact, muramyl dipeptide — a component of peptidoglycan that is associated with various bacteria and frequently contaminates preparations of LPS86. These components do not interact with TLR2, the extracellular receptor for peptidoglycan, and so NOD2 functions as an independent intracellular receptor. Interestingly, NOD2 has a high degree of polymorphism among various laboratory and wild mouse populations, many of which affect the sequences that recognize peptidoglycan95. This indicates. NATURE REVIEWS | IMMUNOLOGY. that NOD2 is highly sensitive to the bacterial environment and undergoes frequent mutations to allow organisms in slightly different niches to cope with these environments, and that NOD2 has an important role in the natural immunity to potential bacterial invaders. The findings that concern the activity of NOD2 and its dysfunction in Crohn’s disease have led to several possible explanations as to the pathogenesis of Crohn’s disease due to NOD2 mutations (FIG. 4). The main hypothesis, at present, is that in the absence of NOD2 activity there is defective activation of macrophages that leads to a persistent infection of macrophages owing to a marked NOD2-independent effector-T-cell response. However, persistent intracellular infection of macrophages has not been detected in Crohn’s disease, and other possibilities need to be considered. A second hypothesis is that in the absence of NOD2 expression by epithelial cells, microbial products that normally activate epithelial cells to secrete chemokines and defensins fail to do so, leading to first, the proliferation of bacteria in the crypts and second, loss of barrier function allowing marked stimulation of mucosal cells by mucosal antigens. The recent observation by Nuñez and colleagues that NOD2 is expressed by epithelial cells at the base of the villous crypts, known as the Paneth cells, supports this possibility (G. Nuñez, personal communication). A third hypothesis is that recognition of microbial peptides by NOD2 normally conditions APCs in a way that leads to their induction of regulatory and effector-T-cell responses, and so failure of this mechanism disrupts mucosal homeostasis. Additional susceptibility loci associated with IBD. Although the identification of NOD2 mutations has been an important scientific breakthrough, it should be emphasized that mutations in this gene occur in only a subgroup of patients with Crohn’s disease. Indeed, since the identification of IBD1 as the first susceptibility locus for Crohn’s disease, several additional chromosomal loci have been identified, indicating the genetic complexity of IBD78. Although, in one example, it has been possible to restrict the area of linkage in IBD5 to a common haplotype that spans 250 kb, no single gene mutation in this region could be identified as the disease-causing mutation96. In this situation, it might be necessary to determine if any of these mutations are associated with disease by using knock-in mice that express individual genes with mutations that are found in patients. Susceptibility genes in animal models. As outlined earlier, susceptibility to colitis in most, if not all, models of experimental colitis differs among inbred strains of mice. This allows possible identification of the chromosome regions and genes that mediate differences in susceptibility in these models, and the relevance of their orthologues to human disease. This approach is particularly appealing, as studying inbred strains of rodents in a controlled environment might overcome many of the problems that are encountered in the complex outbred population. The genetic analysis of animal. VOLUME 3 | JULY 2003 | 5 2 9. © 2003 Nature Publishing Group.

(10) REVIEWS. Commensal bacteria. a. NF-κB inhibitors IL-12-specific antibody. ?. c. Non-follicular induction. TNF-specific antibody. Epithelium. IL-12 Lamina propria DC. MHC. TCR. IFN-γ. Effector T cell. IL-6 IL-1β TNF. TNF. TReg cell. Macrophage. IL-10. TGF-β, IL-10 Peyer's patch. d b Regulatory T-cell induction/function enhanced. Integrin or chemokine receptor inhibitor. Mesenteric lymph node or spleen. Figure 5 | Anatomy of mucosal inflammation and points of therapeutic attack. Depicted here is a generalized pathway of the mucosal inflammation underlying inflammatory bowel disease (IBD) and potential points of therapeutic intervention. a | Secretion of inductive cytokines, such as interleukin-12 (IL-12) or IL-13, can be inhibited with nuclear factor-κB (NF-κB) inhibitors or, more specifically, with various cytokine-specific monoclonal antibodies. This can occur in the mucosal follicle, as shown, or at more downstream sites. b | The traffic of effector cells into the lamina propria can be blocked by inhibitors or monoclonal antibodies specific for integrins or chemokine receptors involved in homing, such as α4β7. c | Tumour-necrosis factor (TNF) expressed on the surface of effector cells can be crosslinked by antibodies specific for TNF leading to apoptosis of effector cells. d | Regulatory T-cell induction or function could be enhanced so as to counteract effector T-cell function. This can be achieved by the delivery of vectors encoding regulatory cytokines.. models of colitis, however, has only recently been instigated. The mouse model that has, so far, provided the most information that is relevant to genetic factors in IBD, is the TNBS-induced colitis model12 (BOX 2). In a genetic study using this model, two susceptibility loci, one on chromosome 9 and one on chromosome 11, were identified in F2 progeny of susceptible SJL/J mice and resistant C57BL/6 mice12. The susceptibility locus on chromosome 11 is of interest as it contains, among others, the gene encoding the IL-12p40 chain. In addition, this locus is associated with susceptibility to other TH1-mediated disease models, such as EAE (Eae6 locus137) and insulin-dependent diabetes in humans (Iddm18 locus138). This indicates that this locus might contain a disease gene that is important to the TH1-cell response. Indeed, other evidence obtained in the same study supports this notion. In particular, it was shown that colitis-susceptible SJL/J mice develop high IL-12p70 responses following systemic administration of LPS, whereas colitis-resistant C57BL/6 mice develop low IL-12p70 responses. In addition, it was shown that the. 530. | JULY 2003 | VOLUME 3. ability to induce a high levels of IL-12 was under the control of a gene that is present in the same region of chromosome 11. So, these data provided indirect evidence for the concept that mucosal inflammation occurs as the consequence of a genetically determined aberrant IL-12 response to constituents of the microbial flora, which in turn induces the pathological TH1-cell response. Additional support for this hypothesis comes from myeloid-specific Stat3-deficient mice with colitis. Using this model, it was shown that in the absence of myeloid-specific Stat3, IL-10 does not effectively suppress the production of IL-12 or IL-23 by myeloid cells, and a TH1 bias of CD4+ T cells occurs that leads to enterocolitis. In addition, it was shown that interbreeding these mice with IL-12p40-deficient mice or TLR4deficient mice — which are incapable of LPS signalling — prevents the development of or ameliorates disease, respectively. So, these findings indicate that the overproduction of IL-12p40 that occurs through LPSinduced TLR4 signalling is essential for the development of a chronic enterocolitis97.. www.nature.com/reviews/immunol. © 2003 Nature Publishing Group.

(11) REVIEWS Other experimental models that have been examined, so far, include the dextran sulphate sodium (DSS)-colitis model, colitis in IL-10-deficient mice and spontaneous colitis in the Samp1/Yit mouse strain98−101. These studies have led to the identification of several chromosome loci that are involved in disease susceptibility, however, although these studies might ultimately lead to the identification of new important pathways that result in mucosal inflammation, identification of the underlying genetic defects is work in progress. Implications for the treatment of IBD. The immunological and genetic insights that concern the pathogenesis of IBD reviewed here provide a roadmap for new therapeutic approaches to these diseases. The knowledge that Crohn’s disease results from TH1-mediated inflammation immediately focuses attention on methods of controlling the TH1-cell pathway by the inhibition of IL-12 and/or IL-23 function and possibly by the inhibition of more downstream cytokines. In the latter regard, the use of antibody specific for TNF has already proven to be a highly effective treatment in Crohn’s disease, achieving positive effects in 60% of patients102. The success rate of TNF-specific antibody is probably due, not to the fact that it blocks TNF activity, but rather to the fact that it induces apoptosis of TH1 effector cells — which presumably express cell-surface TNF. Induction of apoptosis has, in fact, emerged as a main therapeutic mechanism in IBD and one that is applicable to various different therapeutic agents103. Accordingly, IL-12-specific antibody seems to function through this mechanism104, as does IL-6R-specific antibody — an agent that has recently been shown to effectively treat experimental colitis because it blocks the anti-apoptotic signal that is normally delivered by. 1. 2.. 3.. 4.. 5.. 6.. 7. 8.. 9.. Podolsky, D. K. Inflammatory bowel disease. N. Engl. J. Med. 347, 417–429 (2002). Blumberg, R. S., Saubermann, L. J. & Strober, W. Animal models of mucosal inflammation and their relation to human inflammatory bowel disease. Curr. Opin. Immunol. 11, 648–656 (1999). Strober, W., Fuss, I. J. & Blumberg, R. S. The immunology of mucosal models of inflammation. Annu. Rev. Immunol. 20, 495–549 (2002). Mahler, M. & Leiter, E. H. Genetic and environmental context determines the course of colitis developing in IL-10 deficient mice. Inflammatory Bowel Diseases 8, 347–355 (2002). Sartor, R. B. The influence of normal bacterial flora in the development of chronic mucosal inflammation. Res. Immunol. 148, 467–476 (1997). Cong, Y. et al. CD4+ T cells reactive to enteric bacterial antigens in spontaneously colitic C3H/HeJBir mice: increased T helper cell type 1 response and ability to transfer disease. J. Exp. Med. 187, 855–864 (1998). This study shows that T cells that are reactive with conventional antigens of the enteric bacterial flora can mediate chronic inflammatory bowel disease (IBD). Sartor, R. B. in Inflammatory Bowel Diseases (ed. Kirshner, J. B.) 153–178 (Saunders, Philadelphia/London, 1999). Madsen, K. L., Doyle, J. S., Jewell, L. D., Tavernini, M. M. & Fedorak, R. N. Lactobacillus species prevents colitis in interleukin 10 gene-deficient mice. Gastroenterology 116, 1107–14 (1999). Karlsson, M. R., Kahu, H., Hanson, L. A., Telemo, E. & Dahlgren, U. I. Neonatal colonization of rats induces immunological tolerance to bacterial antigens. Eur. J. Immunol. 29, 109–118 (1999).. IL-6 (REF. 105). In addition, azathioprine — a commonly used ‘conventional’ agent in the treatment of Crohn’s disease — has recently been shown to function by subverting a co-stimulatory signal into an apoptotic signal106. Finally, the knowledge that ulcerative colitis results from TH2-mediated inflammation leads to the possibility that inhibitors of the TH2 cytokines, such as IL-13-specific antibody or other cytokine-specific antibodies that are relevant to the TH2 pathway, might be useful in this form of IBD. Another approach to interrupting the TH1 or TH2 pathway takes advantage of the fact that mucosal effector T cells need to traffic from inductive sites, such as mucosal follicles, to effector sites, such as the diffuse lamina propria, the latter being the sites of inflammation107. In fact, treatment with antibodies specific for molecules that are involved in mucosalspecific migration, such as the integrin α4β7, has already been shown to have some therapeutic value108. An alternative approach could be to augment the activity of regulatory cells. This could take the form of the delivery of a regulatory cytokine, such as IL-10 or TGF-β, directly to the inflamed mucosa by genetically engineered microorganisms or cells109,110. Both of these approaches have proven effective in mouse models of inflammation and await the development of delivery systems for use in humans. Although these cytokine-based therapies hold great promise, none can be considered as a permanent cure for IBD. The ideal approach to the cure of IBD is a genetically based approach in which defective genes are replaced. The feasibility of this possibility is likely with respect to the NOD2 mutation as, in this case, the primary lesion might involve an haematopoietic cell that can be replaced with normal genes.. 10. Kontoyiannis, D., Pasparakis, M., Pizarro, T. T., Cominelli, F. & Kollias, G. Impaired on/off regulation of TNF biosynthesis in mice lacking TNF AU-rich elements: implications for joint and gut-associated immunopathologies. Immunity 10, 387–398 (1999). 11. Wirtz, S. et al. Cutting edge: chronic intestinal inflammation in STAT-4 transgenic mice: characterization of disease and adoptive transfer by TNF- plus IFN-γ-producing CD4+ T cells that respond to bacterial antigens. J. Immunol. 162, 1884–1888 (1999). 12. Bouma, G., Kaushiva, A. & Strober, W. Experimental murine colitis is regulated by two genetic loci, including one on chromosome 11 that regulates IL-12 responses. Gastroenterol. 123, 554–565 (2002). This study shows that the intrinsic ability to mount a high interleukin-12 (IL-12) response to constituents of the enteric flora is genetically linked to colitis susceptibility. 13. Strober, W., Kelsall, B. & Marth, T. Oral tolerance. J. Clin. Immunol. 18, 1–30 (1998). 14. Powrie, F., Leach, M. W., Mauze, S., Caddle, L. B. & Coffman, R. L. Phenotypically distinct subsets of CD4+ T cells induce or protect from chronic intestinal inflammation in C. B-17 scid mice. Int. Immunol. 5, 1461–1471 (1993). This study, together with reference 15, shows that the CD45RBlow population of T cells can suppress colitisinducing T helper 1 (TH1)-cell responses, and that colitis can occur as the consequence of deficient or absent T-cell regulatory function. 15. Powrie, F., Correa-Oliveira, R., Mauze, S. & Coffman, R. L. Regulatory interactions between CD45RBhi and CD45RBlow CD4+ T cells are important for the balance between protective and pathogenic cell-mediated immunity. J. Exp. Med. 179, 589–600 (1994).. NATURE REVIEWS | IMMUNOLOGY. 16. Kuhn, R., Lohler, J., Rennick, D., Rajewsky, K. & Muller, W. Interleukin-10-deficient mice develop chronic enterocolitis. Cell 75, 263–274 (1993). This study was published together with references 117 and 124 in the same issue of Cell, and showed that inactivation of entirely different immune genes can lead to spontaneous intestinal inflammation, providing strong evidence for the presence of genetic heterogeneity in IBD. 17. Gorelik, L. & Flavell, R. A. Abrogation of TGF-β signalling in T cells leads to spontaneous T cell differentiation of autoimmune disease. Immunity 12, 171–181 (2000). 18. Hollander, G. A. et al. Severe colitis in mice with aberrant thymic selection. Immunity 3, 27–38 (1995). This study showed that T cells selected in an aberrant thymic microenvironment contain a population of cells that can induce severe colitis that can be prevented by T cells that have undergone normal thymic development, and so emphasizes the role of the thymus in the development of T regulatory cells. 19. Mottet, C., Uhlig, H. H. & Powrie, F. Cure of colitis by CD4+CD25+ regulatory T cells. J. Immunol. 170, 3939–3943 (2003). 20. Neurath, M. F., Fuss, I., Kelsall, B. L., Stuber, E. & Strober, W. Antibodies to interleukin-12 abrogate established experimental colitis in mice. J. Exp. Med. 182, 1281–1290 (1995). This study describes the importance of IL-12 in chronic granulomatous colitis, and the effectiveness of antibodies specific for IL-12 in these diseases. 21. Boirivant, M., Fuss, I. J., Chu, A. & Strober, W. Oxazalone colitis: a murine model of T helper cell type 2 colitis treatable with antibodies to interleukin 4. J. Exp. Med. 188, 129–139 (1998).. VOLUME 3 | JULY 2003 | 5 3 1. © 2003 Nature Publishing Group.

Referências

Documentos relacionados

didático e resolva as ​listas de exercícios (disponíveis no ​Classroom​) referentes às obras de Carlos Drummond de Andrade, João Guimarães Rosa, Machado de Assis,

The fourth generation of sinkholes is connected with the older Đulin ponor-Medvedica cave system and collects the water which appears deeper in the cave as permanent

Despercebido: não visto, não notado, não observado, ignorado.. Não me passou despercebido

Extinction with social support is blocked by the protein synthesis inhibitors anisomycin and rapamycin and by the inhibitor of gene expression 5,6-dichloro-1- β-

i) A condutividade da matriz vítrea diminui com o aumento do tempo de tratamento térmico (Fig.. 241 pequena quantidade de cristais existentes na amostra já provoca um efeito

H„ autores que preferem excluir dos estudos de prevalˆncia lesŽes associadas a dentes restaurados para evitar confus‚o de diagn€stico com lesŽes de

Ousasse apontar algumas hipóteses para a solução desse problema público a partir do exposto dos autores usados como base para fundamentação teórica, da análise dos dados

Os fatores que apresentam maior percentual de casos de não concordância de gênero entre o sujeito e o predicativo/particípio passivo no falar rural do Paraná Tradicional são: