inflammatory bowel disease: is the intestine a trojan horse?

Science Progress (2002), 85 (1), 33–56

Inflammatory bowel disease:is the intestine a Trojan horse?CHRISTELLE BASSET AND JOHN HOLTON

Inflammatory bowel disease (IBD) is a chronic relapsing inflammatorycondition of the intestines that is clinically heterogenous. The cause(s) ofIBD are currently unknown but the mechanisms of injury are immunologi-cal. Increasingly there is an emphasis on the study of the complex inter-actions at the interface of self and non-self- the gastrointestinal epithelialsurface- in relationship to the pathogenesis of disease. There is mountingevidence that a lack of tolerance to the normal commensal flora of the intes-tine may underly the disease pathogenesis. Several genetic loci that aremarkers of disease susceptibility have been identified. These loci map toareas of the genome that are concerned with antigen presentation orcytokine secretion and suggest a genetic heterogeneity that underlies theclinical differences. Overall a picture is emerging of defects in epithelialbarrier function and, or immunoregulation leading to immune responsesthat are triggered or exaggerated by the antigenic components of the normalflora.

IntroductionInflammatory bowel disease (IBD) is a group of idiopathic condi-tions primarily affecting the gastrointestinal tract, but including

Christelle Basset is a lecturer at the Royal Free & UniversityCollege London Medical School in the Windeyer Institute of MedicalSciences, 46 Cleveland Street, London W1T 4JF, UK. She is amember of the Gastrointestinal Inflammation Group and animmunologist by training. Her main research interests are inmucosal immunology, Helicobacter pylori and inflammatory boweldisease. She is currently funded by the National Association ofCrohns and Colitis and from 2002 will be funded by the EuropeanUnion.

John Holton is a Senior Lecturer at the Royal Free & UniversityCollege London Medical School in the Windeyer Institute of MedicalSciences. He is a microbiologist and a member of theGastrointestinal Inflammation Group and his main researchinterests are in Helicobacter pylori and inflammatory bowel disease.

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some extra-intestinal manifestations. It comprises two principal diseases: Crohn’s disease (CD) that can affect the whole of thegastrointestinal tract: from mouth to anus and ulcerative colitis (UC)that affects the colon and rectum1.

Both conditions are more common in Northern/Western Europeand North America although there is a reduced prevalence in Asia,Africa and South America, which suggests a genetic or environmen-tal factor in the pathogenesis of disease. The incidence of CD hadincreased dramatically from about 1.5 per 100,000 population in the1950s to 3.0–5.0 per 100,000 population in the 1970s and the trendcontinues to increase. On the other hand, the incidence of UC hasremained constant at about 10 per 100,000 population over the sameperiod.

The proximate mechanism of disease pathogenesis in both condi-tions is immunological and the location of initial pathology is theepithelial surface of the gastrointestinal tract, which is at the inter-face with the external environment. The gastrointestinal tract is acomplex ecosystem generated by the alliance of epithelium, immunecells and resident microflora. All of them contribute to the defence ofthe host. Because of the high antigenic load within the gastrointesti-nal tract, from both the normal microbial flora to food antigens, thereis an uneasy balance between the need of the host to respond toinvading pathogenic organisms and the need not to respond to thediversity of antigens present in food or as components of the normalflora. What determines whether the host responds by an inflamma-tory reaction or by maintaining tolerance is currently not understoodin detail. However, immune responses, particularly at the epithelialsurface of the intestine, are going to be important for the under-standing of the pathogenesis of both conditions.

The body’s immune system comprises two major divisions: theinnate and the adaptive (acquired) responses. The former is a firstline of defence that can respond immediately to foreign material,detected by pattern recognition molecules that recognise pathogenassociated molecular patterns (PAMP) culminating in phagocytosisand destruction of the foreign material. These pattern recognitionmolecules are found both in the blood, for example, lipopolysaccha-ride binding protein (LBP)2 and expressed on epithelial surfaces, forexample, Toll-like receptors (TLR)3. Also included in the innateresponse is the production of antibacterial peptides, called defencins,secreted from epithelial surfaces4.

The adaptive response, which is antigen specific, takes longer tobe activated and culminates in the secretion of antibodies, which canenhance the process of phagocytosis and the production of cytotoxic

34 Christelle Basset and John Holton


immune cells that can kill virally infected cells or tumour cells. Theadaptive immune response is regulated by the secretion of cytokines(also called interleukins) from a variety of cell types includingimmune cells, epithelial cells and endothelial cells lining blood vessels5. These cytokines are small molecules that modulate thefunction of cells and the expression of specific molecules on the sur-face of cells. There are two main cell types in the adaptive response-the B cell, which produces antibodies and the T cell. There are twotypes of T cell, each fulfilling different functions in the immuneprocess. One type is effector cells that are cytotoxic (called CD8)killing aberrant host cells. The other type of T cell is called T helpercells (CD4) that provide important signals that help orchestrate theimmune response. There are several types of CD4 cell which arecharacterised by the secretion of different cytokines. Th1 cellssecrete cytokines that are pro-inflammatory and involved in cyto-toxic activity; Th2 cells secrete cytokines involved in antibody pro-duction, Th3 and Tr (regulatory) cells secrete cytokines that are anti-inflammatory and down-regulate the pro-inflammatory cascade setoff by the foreign material. T cells interact with other cells via the Tcell receptor (TCR) which is composed of two protein chains either� and � or less frequently � and �6,7. A major problem the immunesystem faces is to distinguish self antigens from foreign or non-selfantigens. These latter can include micro-organisms, food, the growingfoetus and tumour antigens.

Clinically, CD and UC are similar, although pathologically theyare quite different1. On the one hand, CD is a granulomatous conditionthat affects the whole thickness of the intestine and usually begins atthe terminal ileum. Skip lesions are characteristic. Ulcers are foundat the mucosal surface but fissuring and fistulae can develop betweenadjacent loops of intestine or other organs. Immunologically, there isan increase in plasma cells, activated T cells of the Th1 phenotypeand macrophages. Typically the principal antibody isotype is IgG2.On the other hand, UC usually begins in the rectum and in a per-centage of individuals can spread proximally to involve the wholelength of the colon. Macroscopically, there is mucosal ulceration andhaemorrhage although the whole thickness of the intestinal wall isnot affected. A characteristic histological lesion is the crypt abscess.Immunologically, there is an increase in lamina propria plasma cells,activated T cells of the Th2 phenotype and macrophages. Typicallythe plasma cells secrete IgG1 and IgG3. In both conditions, it hasbeen suggested that there is a lack of T-regulatory cells. In UC, anti-neutrophil, anti-colon and anti-bacterial antibodies are frequentlyfound but they are found less so in CD. There are some interesting

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differences between both conditions in the relation to the effects ofsmoking and appendectomy. In CD, there is an 4-6 fold increasedrisk of developing disease associated with smoking but a decreased riskof developing UC. A similar inverse correlation exists betweenremoval of the appendix, which may protect against developing colitis.

Although it isn’t known if the primary defect(s) in either conditionis an aberrant immunological response against a self-antigen(s) or aninappropriate immune response to some external non-self antigen(s),it is evident that the three components found at the epithelial surface– epithelium, immune system, microflora – are involved, with bothgenetic and environmental factors playing a part. Increasingly it isbecoming evident that an important factor in the development orprogression of disease is a failure of tolerance to the normal micro-bial flora of the gastrointestinal tract.

General view of defense mechanisms in the gutTo prevent disease, mucosal surfaces of the intestinal tract are pro-tected by both a carefully regulated non-immune (epithelium barrier,commensal microflora) and immune (gut associated lymphoid tissues(GALT)) system.

Epithelium barrier and secretory IgA: first lines ofdefenseThe first lines of defence, the intestinal epithelium and secretoryIgA, are in place to prevent attachment and penetration of pathogens(Figure 1). The intestinal epithelium is arranged in folds consistingof villi and crypts serving to increase the absorptive surface area. Itis a monolayer of polarised columnar epithelial cells with an apicalbrush border containing dense microvilli and is covered by a thickmucus layer and a glycocalyx8. The mucus layer is a competitiveinhibitor of the micro-organisms’ adhesion to epithelial cells. Theintestinal epithelium is constituted with different cell types, includ-ing enterocytes, goblet cells, Paneth cells, neuro-endocrine cells(NEC) and intraepithelial lymphocytes (IEL), all of them contribut-ing to the barrier and transport functions of the epithelium. Gobletcells secrete both mucus that acts as a protective barrier to luminalaggressins and trefoil peptides that are important for cell regenera-tion. Paneth cells secrete antimicrobial peptides called defencins andthe NEC secrete peptides that can alter the barrier function of theepithelium. The IEL, which by their location, must represent one ofthe first components of the immune system to become exposed to



microbial and food antigens, are mainly CD8 cells (suppressor/cytotoxic activity) but also include some CD4 T cells. The in vivoimpact of these cells remains undefined although they may be impor-tant for differentiation of the epithelial cells as well as immuneresponses. A significant proportion (13%) of IEL have the � and �TCR gene but the biological functions of these cells are still underinvestigation9.

The absorptive epithelial cells (enterocytes) are fused at the cellapex by a “tight junction”, which nevertheless can be regulated bythe autonomic nervous system, cytokines and peptides. Beneath theepithelial layer is the basement membrane and the supporting con-nective tissue of the lamina propria. Myofibroblasts are abundant inthe lamina propria and are involved in maintaining the structuralintegrity of the supporting extracellular matrix.

Secretory IgA (sIgA) is the principal mucosal antibody10. DimericIgA produced by plasma cells in the lamina propria are transportedto the lumen, via the polymeric immunoglobulin receptor on thebasolateral surface of epithelial cells. The main function of sIgA is to inhibit attachment of microbes to epithelial cells and neutralizebacterial products. The sIgA seems also able to neutralize viruswithin the epithelial cells and to excrete antigen from the sub-epithelialcompartment across the epithelium into secretions. Moreover, sIgAis anti-inflammatory and this property is important in the intestinaltract because of the abundance of antigens coming from the com-mensal bacteria and the contents of food. If sIgA was efficient at


Fig. 1 Organisation of the gut associated lymphoid tissue.


activating potent secondary phenomena, such as the complementcascade, the body would seem to be at excessive risk for a state ofchronic mucosal inflammation.

In addition to the mechanical barrier function, intestinal epithelialcells play an important role in initiating and regulating the innateimmunity system11. In the case of adhesion or penetration of epithelialcells by pathogens, they are able to secrete pro-inflammatory andchemoattractant mediators (interleukin (IL)-1, IL-6, IL-8, Tumornecrosis factor � (TNF-�) and granulocyte-monocyte colony stimu-lating factor (GM-CSF)), which will recruit polymorphonuclear leuco-cytes, monocytes/macrophages and subpopulations of T cells to thearea. These effects are mediated by up-regulation of NF-�B, anuclear transcription factor that is a key component in the inductionof cytokines12. This transcription factor is activated following infec-tion with invasive enteric bacteria whereas non-virulent enteric bacteria seem to actively suppress this pathway in epithelial cells13.Moreover, during an inflammatory response, the epithelial cell isinduced to express MHC class II molecules, principally by inter-feron � (INF-�), and thus epithelial cells also seem to play a role inthe adaptive immune system by presenting antigens to T cells.

The intestinal microfloraThe microbial ecology of the gastrointestinal tract differs, dependingupon the region of the intestine although it appears to be stable withtime within an individual in the normal course of events. The numberof cells of the normal bacterial flora exceeds the number of cellswithin the host by a factor of ten and a significant proportion of thesereside within the colon, some 1011 organisms per gram of faeces. Theevidence to date suggests that the anaerobic microflora outnumbersthe aerobic by 1,000–10,000 to one with the principal organismsbeing Gram-negative anaerobes such as Bacteroides species, Gram-positive anaerobes such as Clostridium species and Eubacterium andfacultative anaerobes such as Enterococcus species and Escherichiacoli14. It is additionally recognised that the mucosal surface and themucus are themselves specialised ecological niches with their ownpopulation of, principally, filamentous and spiral shaped bacteriaalthough relatively little is known about this population of bacteriain humans, in either health or disease, because of difficulties in takingsamples compared to faeces. Studies that have been carried out onthe mucosal flora have shown both a high proportion of anaerobes,similar to that found in faeces and that the principle types of organ-isms were Bacteroides species, Gram-positive bacilli and spiralshaped bacteria15.



The normal flora of the intestine is important for the developmentof the gut-associated lymphoid tissue and epithelial function16. Thiscan be seen in animals brought up in a germ-free environment whohave poorly developed mucosal immune systems whilst animalsbrought up in a conventional environment have architecturally moredeveloped immune systems. In gnotobiotic animals, the Peyer’spatches (PP) are small and there are fewer germinal centres andfewer plasma cells in the lamina propria compared to normal animals.A further effect of the microflora on the host is on the developmentof epithelial differentiation. In gnotobiotic animals, the gastrointesti-nal tract wall is thin and less cellular with a reduced total surfacearea compared to conventional animals. There are also morphologi-cal differences in the epithelial cells and the lamina propria has lessstroma and is infiltrated by fewer lymphocytes. Cellular turnover ingnotobiotic animals is very much reduced with the time of migrationof cells from the crypt to the villus tip doubled. Physiological differ-ences are also found in these animals with a decreased motility andincreased absorption of nutrients and increased enzyme activity.After exposure to normal flora, the gastrointestinal tract rapidlytakes on the appearance found in a normal animal. Moreover, arecent study has shown how the commensal microflora can affect thefunctions of epithelial cells17. Finally, the commensal microflora alsoplays a role in defence by providing colonisation resistance andcompeting with pathogens for space and nutrients.

Gut associated lymphoid tissues (GALT)Although epithelial cells can initiate immune responses, the GALT ischaracterized by inductive and effector sites (the lamina propria)(Figure 1). The initiation of immune responses takes place mainly inPP, lymphoid nodules and the appendix which represent the induc-tive arm18,19.

PP and other lymphoid nodules occur along the entire length ofthe gastrointestinal tract with larger numbers present in the colon andthe rectum. These sites are covered by a specialized epithelium,which is constituted with M cells that facilitate antigen samplingfrom the lumen. PP are distinct areas comprised of B cells (B cellfollicle) and which are surrounded by T cells. The area between thefollicles and the mucosal epithelium is enriched with B cells, T cells,macrophages and dendritics cells. M cells are cells that lack a brushborder, contain numerous endocytic vesicles and morphologicallyhave an invagination on the anti-luminal surface, which containslymphocytes and macrophages. After uptake of antigens by M cellsand sampling by the cells on its anti-luminal surface, antigens will be



then presented via the antigen presenting cells (APC: dendritic cells,macrophages and B cells) to T lymphocytes. Dendritic cells, whichare the unique APC capable of priming naïve T cells, are present inlarge numbers in the PP compared to the lamina propria. However, atcertain sites of the gastrointestinal tract they can have a direct accessto luminal antigens through their dendrites. T helper cells produce avariety of inflammatory or immunoregulatory cytokines and havebeen subdivided further into Th0 cells, Th1 cells and Th2 cells(Figure 2). Th1 cells are characterized by production of IL-2, TNF-�, GM-CSF and IFN-� and effect cell-mediated immunity by regu-lation of macrophage functions, delayed-type hypersensitivityresponses and cytotoxic T cell responses. In contrast, Th2 cells aredistinguished by their support of humoral immunity through thesecretion of selected cytokines, such as IL-4, IL-5, IL-6 and IL-10.The induction of a Th1 or Th2 profile depend on many parameters,among them the nature of antigen and the APC play an important


Fig. 2 Differentiation of CD4 T cells.


role. It has been demonstrated that dendritic cells from PP wereshown to promote differentiation of a naïve T helper cell line into aTh2 cell while splenic dendritic cells supported development of aTh1 cell20. However, different phenotypes of dendritic cells are alsopresent in PP. Some can generate Th3 response via secretion of IL-10 and transforming growth factor (TGF-�) or be primed to secreteIL-12 and promote the production of IFN-� by the naïve T cells theyprime.

After the initial exposure to antigen, primed B and T lymphocytesleave PP and migrate to mucosal tissues via the mesenteric lymphnodes and recirculate through the thoracic duct and the blood toreach the spleen and seed back into the intestinal mucosa. Homing oflymphocytes to specific areas is regulated by addressins. Addressinsare molecules expressed on the surface of blood vessels and lymphnodes, which bind lymphocytes expressing its ligand, thus localisingthe lymphocyte to the appropriate histological site. In the intestine,the major addressin is mucosal addressin adhesion molecule 1(MAdCAM-1) which is found in PP and the high endothelial venulesof mesenteric lymph nodes and the vasculature of the lamina propria.The ligands for MAdCAM-1, which are found on the lymphocytes,are L-selectin and �4�7integrin, the latter being important for homingto PP and the lamina propria11,18. The lamina propria contains a fullcomplement of cells including plasma B cells, T cells and mono-cytes/macrophages. In the healthy mouse intestine, in the PP, CD4 Tcells are predominantly Th2-like whereas in human intestine, Th1-like T cells predominate21. This in humans, Th1-like T cells should beinactivated or eliminated in the periphery. Th2-like T cells providehelp for IgE and IgA responses. Of the cytokines secreted by Th2cells, IL-4 is important for activation of IgE responses and IL-5 andIL6 are important for differentiation of B cells into plasma cellssecreting IgA. The most important influence on IgA secretion isTGF-� which promotes class switching in B cells to secrete IgA.Although the micro-environment in the healthy lamina propria isanti-proliferative and suppressive, an ability to respond to invadingpathogens is also required. Intracellular pathogens such asSalmonella that invade the mucosa via the M-cells are processed bymacrophages. These cells secrete IL-12 which promotes differentia-tion to Th1 helper cells, which in turn, by secretion of TNF-� andINF-�, activates macrophages to kill the invading pathogen. As wewill mention later, Th1 and Th2 responses are highly regulated,avoiding overactivity that can be damaging for the body, as observedduring inflammatory bowel disease.



Oral toleranceThe mucosal immune system not only provides resistance to infec-tious agents, but it must also differentiate between harmful agentsand innocuous substances (food antigens and gut flora). Thus, thedevelopment of mucosal immunity is carefully regulated of neces-sity and the mechanism is known as oral tolerance. Oral tolerance isa process by which the immune system establishes and maintainsunresponsiveness to antigens on the mucosal surfaces. This toleranceis important in order to prevent the intestine being in a continuousstate of inflammation. This tolerance acts mainly on Th1 responses,which are responsible for inflammatory reactions. However, Th2cells are susceptible to oral tolerance too because even Th2responses can be pathogenic. Th1 responses are down regulated bythree possible mechanisms, clonal deletion (apoptosis), anergy oralternatively by active suppression22. Active suppression has beenobserved at low doses of antigen whereas clonal deletion and anergyhave been observed with high doses of antigen23. In actual fact,induction of active suppression and clonal deletion/anergy are simul-taneous and competing processes, the balance of which depends onthe frequency, the dose and the nature of administrated oral antigen.

Activation of CD4 T cells is mediated via two signals (Figure 3).The first signal is the recognition of the [MHC (Major histocompati-


Fig. 3 Presentation of antigen to CD4 T cells.


bility complex) molecule-antigen] complex on the APC by the T cellreceptor (TCR) on the T cell. The second signal involves severalbinding and co-stimulatory molecules and permits full activation ofthe T cell. One of the most important co-stimulatory molecules isCD28 on the T cell, which recognises B7 molecules on the APC.This recognition triggers the secretion of IL-2, which acts as a posi-tive feedback and permits the proliferation of the T cell. Expressionof co-stimulatory molecules on APCs are induced by microbes andby cytokines produced during innate immunity. Activation of T cellsis followed by differentiation (Th1 or Th2), and proliferation inorder to eliminate the antigen. After elimination of the antigen,which results in the downregulation of the costimulatory moleculesand cytokines, T cells die by apoptosis. This apoptosis is also called“death by neglect”. Self-antigen or antigen administrated withoutadjuvant does not induce expression of costimulatory molecules andso induces T cell anergy (Figure 4).

Mechanisms exist to control inadequate or excessive activationand/or proliferation. CTLA-4 molecules are induced on T cells afteractivation and bind to B7 molecules. Because CTLA-4 inhibits thetranscription of IL-2 and the progression through the cell cycle, lead-ing to anergy of T cells, CTLA-4 seems to act as an antagonist ofCD28. However, the role of this molecule is not completely under-stood. Activation of T cells also leads to co-expression of the deathreceptor, Fas (CD95), and its ligand, Fas ligand (FasL), resulting indeath by apoptosis of the same and neighbouring cells. This processoccurs to limit the magnitude and duration of the initial Th1response, as occurs during repeated activation by antigen. Finally,activation can be regulated by suppressor factors, TGF-� and IL-10.Cells secreting TGF-� are named Th3 cells and are CD424. Activesuppression exerted by these cells seems to take place initially in PP.The differentiation of T cells into Th3 cells is antigen specific andthese cells produce a bystander suppression of neighbouring cells.Indeed, feeding with one tolerogenic antigen can suppress theresponse against an immunogenic antigen. This phenomenon can berelevant in term of treatment, as demonstrated in animal models ofautoimmune disease23. Development of T cells producing TGF-� inPP is positively correlated with Th2 responses. The other type ofCD4 suppressor cells is T regulatory 1 cells (Tr1 cells) that secreteTGF-� and IL-1025, 26. They are antigen specific and exert a suppres-sive bystander effect similar to Th3 cells. IL-10 is known to inhibitT cell activation and cytokine secretion by APC. The major cell pop-ulation involved in oral tolerance appears to be the CD4 T-helper cellbut CD8 cells have also been described.



In contrast to TGF-� and IL-10, IFN-� secreted by Th1 cells andmacrophages and IL-12 secreted by macrophages or dendritic cellscan prevent oral tolerance. So, it seems that the immune system inthe GALT is a balance between anti-inflammatory and suppressivecytokines as IL-4, IL-10 and TGF-�, and pro-inflammatorycytokines as IL-12 and IFN-�.


Fig. 4 Control mechanisms of activation and proliferation of T cells.


Inflammatory bowel diseaseAs mentioned before, the GALT is highly regulated in order torespond to pathogens and to tolerate the commensal bacteria andfood antigens. CD and UC are both characterized by chronic inflam-mation in the gut. CD can affect the whole of the gastrointestinaltract but with a predilection for the terminal ileum. UC affects thecolon and the rectum. The inflammation observed in IBD may bedue either to an inadequate or exaggerated immune response againstone/several organism(s) or be initiated by an abnormal permeabilityof the epithelium. Animal models of IBD, bred in germ-free condi-tions, do not develop the disease but do so if either a specific micro-organism or conventional colonic microflora are reintroduced.Moreover, patients with CD respond well to antibiotic therapy,which changes the enteric microflora or to diversion of the faecalstream. Additionally, one study has shown27 that relatives, thatshared the same particular microflora with CD patients, were moresusceptible to develop the disease. Studies in humans have not yetconvincingly determined if one particular pathogen or a globalchange in the microflora, or a combination of both, one being theconsequence of the other, is responsible for IBD. As further evidenceof the importance of the microflora in the disease process,Duchmann et al.28 have demonstrated that lamina propria mono-nuclear cells from inflamed intestine of IBD patients were not tolerantto the autologous intestinal flora of patients. All these observationsunderline the importance of bacterial antigens as triggering the disease. On the other hand, animal models that develop IBD do sohaving a background of a defective immune system or epithelial barrier. The closed inter-relation between the commensal bacteriaand the GALT makes IBD a very complex disease, in which severalgenetic factors play an important role.

Genetic factors in IBDGenome wide scans for susceptibility loci in monozygotic twinswith IBD have shown 46 markers were significantly associated tosome degree29 demonstrating loci on chromosomes 2, 3, 6, 7, 12, 14and 16. Both CD and UC show a large genetic heterogeneity in disease pathogenesis, yet with a clear genetic component. This ismore clearly defined for CD. The proband concordance rate is 50%for monozygotic twins in CD but only 6% in UC. Loci that havebeen identified such as D12S83; D7S669; D3S1573; D14S261 arelinked to either antigen presentation, mucin secretion or cytokinesecretion and are thus biologically plausible. There are clear links to



certain HLA II types with positive association (in UC) with DR1,DR2 and negative association with DR430; and for CD, positiveassociations with DR7 and DRB331. One of the closest associationsis with DRB1*103 and DRB1*0301 in UC but not CD. BothDRB1*0103 and allele 2 of the IL-1receptor antagonist are closelylinked to extensive disease in UC. Severe disease is also associatedwith polymorphism in the inhibitor of kB-like gene32 (similar instructure to the IkBa protein).

Recent interest has also focussed on polymorphism in the NOD233 protein. It is an Apaf1-like protein, found only in monocytes andis an intracellular ligand for LPS. Variations in binding of LPS toNOD 2 may lead to an exaggerated or sustained response by theimmune system due to antigen persistence and hence disease. It isspecifically associated with susceptibility to CD.

Similarly to the genetic factors involved in both conditions, thereis a strong environmental component to disease and much circum-stantial evidence to implicate the involvement of a microbial factor.Current thinking is that there is a breakdown of tolerance to the normalflora of the intestine.

Microbial factorsIt is clear from studies in both animals and in humans that thecolonic microflora is an essential stimulus for the development ofIBD. There are several gene-knockout and transgenic animal modelsthat develop IBD. IL-10, IL-2, IL-12, and TCR–/– knockout micewill develop IBD only if exposed to normal colonic microflora34.Similarly, transgenic mice carrying the human IgE receptor developIBD in response to a chemical stimulus, however, the response isexaggerated by the presence of normal colonic flora35. In these animals, it has been shown that different components of the microbialflora are important for perpetuating the inflammatory response36. Inother models, specific organisms could induce the development ofdisease, such as Helicobacter bilis or Helicobacter hepaticus inSCID mice37,38, or Helicobacter hepaticus in IL-10 knockout mice39.

Several studies have been carried out in order to find a specificpathogen implicated in the pathology of IBD, however, none of themhave been conclusive40,41,42. Some of this attention has focused onHelicobacter pylori and other Helicobacter species for main tworeasons. The first reason is Helicobacter pylori is recognised as thecause of the majority of chronic inflammation of the stomach leadingto ulceration in some43. The second reason is other Helicobacterspecies have been found in the colon of both humans and animalsand in some of them are linked to the development of colitis.



Helicobacter cinaedi and Helicobacter fennelliae have been foundin men with proctitis and colitis44. A novel intestinal Helicobacterspecies has been isolated from Cotton-top tamarins that develop colitis, which resemble those of UC in humans45. Also as mentionedabove, in immune deficient mice, Helicobacter hepaticus andHelicobacter bilis trigger the development of IBD. Two other studieshave reported the presence of Helicobacter species in the normal andpathogenic human colon46,47. Taken together, these studies demon-strated that Helicobacter species can be found in both human andanimal and cause disease. As specifically demonstrated in immunedeficient mice, antigens from Helicobacter species can stimulate the immune system and trigger inflammation. One can imagine that Helicobacter species colonize the normal colon of human andanimal and in some susceptible recipients can trigger inflammation.The inflammation observed in Helicobacter pylori infection resem-bles in some aspects the inflammation observed in IBD. More studies are necessary to confirm the presence of Helicobacter speciesin the colon, because studies are contradictory and their role in IBDuncertain48.

Escherichia coli (E. coli) is the predominant aerobic Gram-nega-tive species of the normal intestinal flora but pathogenic forms arealso well known and cause intestinal disease in humans. An increasein enteroadhesive E. coli has been noted in UC. Further, because CDis a granulomatous condition and this condition is also observed duringinfection with invasive bacteria, a role for E. coli in CD has alsobeen under investigation. Recently, Masseret et al. have isolated E. coli strains from the ileal mucosa of patients with CD. Thesestrains belong to a new potentially pathogenic group called AIEC foradherent invasive E. coli49. These strains adhere and invade culturedintestinal epithelial cells and macrophages. Moreover, these strainsare able to replicate within macrophages but do not trigger theirdeath as observed with others strains of E. coli and macrophages areable to release TNF-�50. Similarly, novel bacterial sequences thathave been identified in the monocytes of patients with CD, have sub-sequently been shown to be superantigens and may thus be related toexcess T cell stimulation and disease pathogenesis.

Bacteroides fragilis and Bacteroides vulgatus are also creatinginterest. Enterotoxigenic Bacteroides fragilis has been found morefrequently in IBD patients compared to controls51 although this find-ing needs to be confirmed. Additionally, Bacteroides vulgatus hasbeen shown to be important in colitis induced in HLA B27 trans-genic animals, in carrigageenan induced colitis, in guinea pigs and ithas been shown that T cell responses in patients with IBD are



directed against a broad range of enteric bacteria includingBacteroides vulgatus52.

Although any specific pathogen has still not been linked to IBD,the total commensal microflora of the patients seems to play animportant role in the disease. As mentioned before, it has beenreported that relatives that shared the same particular microflora withCD patients were more susceptible to develop the disease27 and inanimal models of IBD, reconstitution of the normal flora exacer-bated the illness. No one has looked in details at these particularmicroflora, however, new techniques have been recently availablewhich might be useful for the study of the complex mixture of organ-isms that represent the colonic microflora53. This particular study27

also underlines the importance of genetic factors, in for exampledetermining the expression of the mucus layer on the epithelial cellsand in the establishment of the commensal microflora and infec-tion54. Further, this particular microflora might affect the functionsof epithelial cells and the immune responses and thereby favourinfection55. For example, the commensal microflora of CD patientsis characterized by a decrease in Bifidobacteria56, which may be pro-tective. Studies have demonstrated that Lactobacillus plays animportant role in protecting the host against invasion with potentialpathogens and can stimulate the immune system to secrete sIgA.One study has demonstrated that IL-10 gene-deficient mice have anincreased level of colonic mucosal adherent and translocated aerobicbacteria, which precedes development of colitis57. Repopulation ofthe colonic lumen with control levels of Lactobacillus preventedcolitis in this model.

Probiotics are under investigations as a potential therapy in thetreatment of IBD58. Probiotics are live micro-organisms (Lactobacilli,Bifidobacteria and E. coli Nissel strain) that alter the enteric micro-flora and have a beneficial effect on health. They may be good for usas demonstrated by many studies although some studies show nobenefit59. However, we do not know how they work and specificallyhow they stimulate the immune system and in the case of IBD howthey can restore a “tolerant environment”. As mentioned before,some antigens are tolerogenic and some are immunogenic and anti-gens from the commensal bacteria play an important role in theestablishment of oral tolerance. Some tolerogenic antigens from thecommensal bacteria may become non-tolerogenic in IBD because ofa failure in a downstream mechanism. Antigens from probioticsmight efficiently compete with immunogenic antigens from patho-genic or commensal bacteria. Indeed, a recent study demonstratesthat probiotics decrease the level of pro-inflammatory cytokines



(INF-�) and increase the level of anti-inflammatory cytokines (IL-10)60.

Factors from the GALT and epithelium barrierOn the one hand, animal models of IBD have demonstrated the roleof the enteric bacteria as stimuli and, on the other hand, they havedemonstrated that a dysfunction of the immune system or the epithe-lial barrier can lead to IBD61.

Most of the animal models, which resemble CD in humans,showed an imbalance between pro-inflammatory and anti-inflamma-tory cytokines with a central role for activated CD4 cells. Indeed, inan animal model of colitis, a synthetic mimetic of the complemen-tarity, determining region of the CD4 molecule decreases the pro-duction of pro-inflammatory cytokines with improvement of symp-toms62 thus suggesting an avenue of therapy. An overproduction ofIL-12 and TNF-�, secreted by APC, and INF-� and TNF-�, secretedby Th1 T cells, is responsible for the perpetuation of the inflamma-tion. In humans, an increase in the number of activated mucosal Tcells secreting IFN-� and an increased mucosal production ofcytokines that activate Th1 T cells (IL-12 and IL-18) have beenreported in CD. In animal models of UC, the inflammation is drivenby IL-4 producing Th2 T cells.

One cytokine that plays an important role in IBD is TNF-�. Themajor source of TNF-� is the monocyte/macrophage population. Aninteresting animal model, that over-expresses TNF-�, develops aninflammation initially in the small intestine (rather than the largeintestine) and a granuloma, resembling CD in humans63. Further evidence that TNF-� is important is the demonstration that in-vitroexposure of lamina propria mononuclear cells to TNF-� can aug-ment the production of Th1 cytokines. Further, an injection of achimeric anti-TNF-� monoclonal antibody to patients with CD,downregulates Th1 cytokines production and is paralleled by clini-cal improvement64. The use of anti-TNF-� is now used successfullyas a treatment for some patients with CD.

As well as Th1 cytokines, TNF-� can stimulate the production ofchemokines such as IL-8, from mucosal epithelium, as part of theinflammatory response. This pathway is mediated by upregulation ofNF-�B brought about by inactivation of the NF-�B inhibitor I�B byI�B kinase (IKK) (Figure 5). Activation of this cell signalling path-way also leads to the production of two anti-apoptotic proteinsGADD and XIAP, which prevent the action of the mitogen associ-ated protein kinase (MAPK) pathway of apoptosis. An alternativepathway may result from Fas-FasL binding which ultimately leads to



activation of caspase 3 and DNA degredation. This pathway is inhibited by IL-12 and IL-6 binding – both Th1 pro-inflammatorycytokines. Thus, in mucosal cells, there is a balance between theinflammatory pathway via the transcription factor NF-�B and apop-tosis of the epithelial cells (or T cells) which is, to a large extentdependent upon the concentration of cytokines and this balance isdisturbed in disease.

Activation-induced T cell apoptosis is an important mechanism todown regulate the inflammatory response. In the mucosa, the rate ofFas mediated apoptosis in T cell is high suggesting the T cells areconstantly activated, presumably by the antigenic stimulus. Laminapropria T cells from patients with IBD show defective apoptosis andare less sensitive to the pro-apoptotic signals. Thus, one main mech-anism for disease pathogenesis may be the prolonged survival ofactivated T cells in the mucosa, which may enhance the inflamma-tory cascade65. An alternative mechanism of mucosal injury may bedue to activated CD8 T cells expressing the ligand FasL, which isupregulated in those cells infiltrating the mucosa of patients withUC. Fas is expressed on epithelial cells from patients with UC andFas-FasL, interaction may lead to epithelial cell apoptosis. This maybe important in mucosal damage in patients with UC66.


Fig. 5 Intracellular signalling controlling inflammation or apoptosis.


An increased permeability of the epithelium has been reported inrelatives of IBD patients67. Further, it is recognised that an increasein epithelial permeability precedes clinical disease68 and relapse. In-vitro experiments show that epithelial monolayers exposed tomononuclear cells from patients with CD that were unactivated byLPS increased the secretory activity and decreased the barrier acti-vity. These mononuclear cells from CD patients spontaniouslysecrete TNF-� and produce changes similar to LPS-activatedmononuclear cells69. A leak in the epithelium barrier might permitaccess of antigens to the immune system, these antigens being nor-mally sequestrated in the lumen. This might be one of the reasonswhy lamina mononuclear cells from inflamed intestine of IBDpatients proliferate in presence of autologous intestinal flora28.Inflammation may exacerbate the intestinal permeability and there issome evidence that it may be a secondary event rather than a primarycause70.

It is also known that the epithelial barrier plays a role in the estab-lishment of oral tolerance. Bacterial products are recognized by aclass of pattern-recognition receptors, called Toll-like receptors(TLR)3. Some of them are expressed on macrophages, dendritic cellsand intestinal epithelial cells. TLR4 on macrophages is known tobind LPS. This binding triggers the activation of the NF-�B cascadeand release of pro-inflammatory cytokines. One study has shownthat intestinal epithelial cells constitutively express TLR3 andTLR5, while TLR2 and TLR4 are only barely detectable. In CD,however, the expression of TLR3 is downregulated, and in CD andUC, the expression of TLR4 is strongly up-regulated71. We still donot know if this phenomenon is primarily causal or due to the effectof macrophages, for example, on intestinal epithelial cells. Anotherstudy has demonstrated that TLR4 is down regulated in intestinalepithelial cell lines72. These two studies underline the possibility thata dys-regulation of TLR in IBD might be in part responsible of thebreakdown of tolerance against components of the intestinalmicroflora.

References1. Jewell, J.P. Crohn’s Disease; Ulcerative Colitis. (1996) In: Wetherall, D.J.,

Ledingham, J.G.G., Warrell, D.A. (eds) Oxford Textbook of Medicine 3rd edn,Vol. 2. pp. 1936–1950. OUP.

2. Le Roy, D., Di Padova, F., Adachi, Y., Glauser, M.P., Calandra, T. & Heumann,D. (2001) Critical role of lipopolysaccharide-binding protein and CD14 inimmune responses against gram-negative bacteria. J. Immunol., 167,2759–2765.



3. Hayashi, F., Smith, K.D., Ozinsky, A., Hawn, T.R., Yi, E.C., Goodlett, D.R.,Eng, J.K., Akira, S., Underhill, D.M. & Aderem, A. (2001) The innate immuneresponse to bacterial flagellin is mediated by Toll-like receptor 5. Nature, 410,1099–1103.

4. Nizet, V., Ohtake, T., Lauth, X., Trowbridge, J., Rudisill, J., Dorschner, R.A.,Pestonjamasp, V., Piraino, J., Huttner, K. & Gallo, R.L. (2001) Innate anti-microbial peptide protects the skin from invasive bacterial infection. Nature,414, 454–457.

5. Curfs, J.H., Meis, J.F. & Hoogkamp-Korstanje, J.A. (1997) A primer on cytokines:sources, receptors, effectors and inducers. Clin. Microbiol. Rev., 10, 742–780.

6. Delves, P.J. & Roitt, I.M. (2000) The Immune system. First of two parts. N. Engl. J. Med., 343, 37–49.

7. Delves, P.J. & Roitt, I.M. (2000) The Immune system. Second of two parts. N.Engl. J. Med., 343, 108–117.

8. Berin, M.C., McKay, D.M. & Perdue, M.H. (1999) Immune-epithelial interac-tions in host defense. Am. J. Trop. Med. Hyg., 60, 16–25.

9. Kronenberg, M. & Cheroutre, H. (2000) Do mucosal T cells prevent intestinalinflammation? Gastroenterology, 118, 974–977.

10. Lamm, M.E. (1997) Interactions of antigens and antibodies at mucosal surfaces.Ann. Rev. Microbiol., 51, 311–340.

11. Kagnoff, M.F. & Eckmann, L. (1997) Epithelial cells as sensors for microbialinfection. J. Clin. Invest., 100, 6–10.

12. Barnes, P.J. & Karin, M. (1997) Nuclear factor-kappaB: a pivotal transcriptionfactor in chronic inflammatory diseases. N. Engl. J. Med., 336, 1066–1071.

13. French, N. & Pettersson, S. (2000) Microbe-host interactions in the alimentarytract: the gateway to understanding inflammatory bowel disease. Gut, 47, 162–163.

14. Holdeman, L.V., Good, I.J. & Moore, W.E.C. (1976) Human fecal flora: variation in bacterial composition within individuals and a possible effect ofemotional stress. Appl. Environ. Microbiol., 31, 359–375.

15. Savage, D.C. (1983) Morphological diversity among members of the gastro-intestinal microflora. Int. Rev. Cytol., 82, 305–334.

16. Steege, J.C., Buurman, W.A. & Forge, P.P. (1997) The neonatal development ofintraepithelial and lamina propria lymphocytes in the murine small intestine.Dev. Immunol., 5, 121–128.

17. Hooper, L.V., Wong, M.H., Thelin, A., Hansson, L., Falk, P.G. & Gordon, J.I.(2001) Molecular analysis of commensal host-microbial relationships in theintestine. Science, 291, 881–884.

18. Simecka, J.W. (1998) Mucosal immunity of the gastrointestinal tract and oraltolerance. Adv. Drug. Deliv. Rev., 34, 235–259.

19. Ernst, P.B., Song, F., Klimpel, G.R., Haeberle, H., Bamford, K.B., Crowe, S.E.,Ye, G. & Reyes, V.E. (1999) Regulation of the mucosal immune response. Am.J. Trop. Med. Hyg., 60, 2–9.

20. Iwasaki, A. & Kelsall, B.L. (1999) Mucosal immunity and inflammation. IMucosal dendritic cells: their specialized role in initiating T cell responses. Am.J. Physiol. Gastrointest. Liver Physiol., 276, G1074–G1078.

21. Mac Donald, T.T. & Monteleone, G. (2001) IL-12 and Th1 immune responsesin human Peyer’s patches. Trends Immunol., 22, 244–247.

22. van Parijs, L. & Abbas, A.K. (1998) Homeostasis and self-tolerance in theimmune system: turning lymphocytes off. Science, 280, 243–248.



23. Fowler, E. & Weiner, H.L. (1997) Oral tolerance: Elucidation of mechanismsand application to treatment of autoimmune diseases. Biopoly, 43, 323–335.

24. Strober, W., Kelsall, B. & Marth, T. (1998) Oral tolerance. J. Clin. Immunol.,18, 1–30.

25. Groux, H., O’Garra, A., Bigler, M., Rouleau, M., Antonenko, S., de Vries, J.E.& Roncarolo, M.G. (1997) A CD4+ T-cell subset inhibits antigen-specific T-cell responses and prevents colitis. Nature, 389, 737–742.

26. Groux, H. & Powrie, F. (1999) Regulatory T cells and inflammatory bowel disease. Immunol. Today., 20, 442–445.

27. van de Merwe, J.P., Schroder, A.M., Wensinck, F. & Hazenberg, M.P. (1988)The obligate anaerobic faecal flora of patients with Crohn’s disease and theirfirst-degree relatives. Scand. J. Gastroenterol., 23, 1125–1131.

28. Duchmann, R., Kaiser, I., Hermann, E., Mayet, W., Ewe, K. & Meyer zumBuschenfelde, K.H. (1995) Tolerance exists towards resident intestinal flora butis broken in active inflammatory bowel disease (IBD). Clin. Exp. Immunol.,102, 448–455.

29. Satsangi, J., Parkes, M., Louis, E., Hashimoto, L., Kato, N., Welsh, K.,Terwilliger, J.D., Lathrop, G.M., Bell, J.I. & Jewell, D.P. (1996) Two stagegenome wide search in inflammatory bowel disease provides evidence for susceptibility loci on chromosomes 3, 7 and 12. Nature (Genet), 14,199–202.

30. Toyoda, H., Wang, S.J., Yang, H.Y., Redford, A., Magalong, D., Tyan, D.,McElree, C.K., Pressman, S.R., Shanahan, F. & Targan, S.R. (1993) Distinctassociations of HLA class II genes with inflammatory bowel disease.Gastroenterology, 104, 741–748.

31. Stokkers, P.C., Reitsma, P.H., Tytgat, G.N. & van Deventer, S.J. (1999) HLA-DR and DQ phenotypes in IBD- a meta analysis. Gut, 45, 395–401.

32. de la Concha, E.G., Fernandez-Arquero, M., Lopez-Nava, G., Martin, E.,Allcock, R.J., Conejero, L., Paredes, J.G. & Diaz-Rubio, M. (2000)Susceptibility to severe ulcerative colitis is associated with polymorphism in thecentral MHC gene IKBL. Gastroenterology. 119, 1491–1495.

33. Ogura, Y., Bonen, D.K, Inohara, N, Nicolae, D.L, Chen, F.F, Ramos, R,Britton, H, Moran, T., Karaliuskas, R., Duerr, R.H., Achkar, J.P., Brant, S.R.,Bayless, T.M., Kirschner, B.S., Hanauer, S.B., Nunez, G. & Cho, J.H. (2001) A frameshift mutation in NOD 2 associated with susceptibility to Crohn’s dis-ease. Nature, 411, 603–606.

34. Elson, C.O., Sartor, R.B., Tennyson, G.S. & Riddell, R.H (1995) Experimentalmodels of inflammatory bowel disease. Gastroenterology, 109, 1344–1367.

35. Dombrowicz, D., Nutten, S., Desreumaux, P., Neut, C., Torpier, G., Peeters, M.,Colombel, J.F. & Capron, M. (2001) Role of the high affinity immunoglobulinE receptor in bacterial translocation and intestinal inflammation. J. Exp. Med.,193, 25–34.

36. Rath, H.C., Schultz, M., Freitag, R., Dieman, L.A., Li, F., Linde, H.J.,Scholmerich, J. & Sartor, R.B (2001) Different subsets of enteric bacteriainduce and perpetuate experimental colitis in rats and mice. Infect. Immun., 69,2277–2285.

37. Cahill, R.J., Foltz, C.J., Fox, J.G., Dangler, C.A., Powrie, F. & Schauer, D.B.(1997) Inflammatory bowel disease: an immunity-mediated condition triggered bybacterial infection with Helicobacter hepaticus. Infect.Immun., 65, 3126–3131.



38. Shomer, N.H., Dangler, C.A., Schrenzel, M.D. & Fox, J.G. (1997) Helicobacterbilis-induced inflammatory bowel disease in scid mice with defined flora. Infect.Immun., 65, 4858–4864.

39. Kullberg, M.C., Ward, J.M., Gorelick, P.L., Caspar, P., Hieny, S., Cheever, A.,Jankovic, D. & Sher, A. (1998) Helicobacter hepaticus triggers colitis in specific-pathogen-free interleukin-10 (IL-10)-deficient mice though an IL-12-and gamma interferon-dependent mechanism. Infect. Immun., 66, 5157–5166.

40. Martin, H.M. & Rhodes, M.J. (2000) Bacteria and Inflammatory BowelDisease. Curr. Opin. Infect. Dis., 13, 503–509.

41. Campieri, M. & Gionchetti, P. (2001) Bacteria as the cause of ulcerative colitis.Gut, 48, 132–135.

42. Caradonna, L., Amati, L., Magrone, T., Pellegrino, N.M., Jirillo, E. & Caccavo, D. (2000) Enteric bacteria, lipopolysaccharides and related cytokinesin inflammatory bowel disease: biological and clinical significance. J. Endotoxin. Res., 6, 205–214.

43. Blaser, M.J. (1999) Hypothesis: the changing relationships of Helicobacter pyloriand humans: Implications for health and disease. J. Infect. Dis., 179, 1523–1530.

44. Totten, P.A., Fennell, C.L., Tenover, P.C., Wezenberg, J.M., Perine, P.L.,Stamm, W.E. & Holmes, K.K. (1985) Campylobacter cinaedi andCampylobacter fennelliae two new campylobacter species associated withenteric disease in homosexual men. J. Infect. Dis., 151, 131–139.

45. Saunders, K.E., Shen, Z., Dewhirst, F.E., Paster, B.J., Dangler, C.A. & Fox, J.G.(1999) Novel intestinal Helicobacter species isolated from Cotton-TopTamarins (Saguinus Oedipus) with Chronic Colitis. J. Clin. Microbiol., 37,146–151.

46. Danon, S.J., Goh, K.L., Parasakthi, N., Neilan, B.A., Grehan, M.J. & Lee, A.(2000) The human colon: a reservoir for Helicobacter species. Gut, 47, A2.

47. Oliveira, A.G., Queiroz, D.M.M.M., Rocha, G.A., Rocha, A.M.C., Santos, A.,Sana, M.G.P., Moura, S.B., Correa, P.R.V. & Camargos, E.R.S. (2001)Isolation of Helicobacter strains from the colonic mucosa of inflammatorybowel disease (IBD) patients. Gut, 49, A1.

48. Grehan, M.J. & Danon, S.J. (2001) Absence of Helicobacter species in theileum and colon of Australians with and without inflammatory bowel disease(IBD) – Should we be looking at other mucus-associated bacteriaGastroenterology, 120 (Supp. 1), A-337.

49. Masseret, E., Boudeau, J., Colombel, J.F., Neut, C., Desreumaux, P., Joly, B.,Cortot, A. & Darfeuille-Michaud, A. (2001) Genetically related Escherichia.coli strains associated with Crohn’s disease. Gut, 48, 320–325.

50. Glasser, A.L., Boudeau, J., Barnich, N., Perruchot, M.H., Colombel, J.F. &Darfeuille-Michaud, A. (2001) Adherent invasive Escherichia coli strains frompatients with Crohn’s disease survive and replicate within the macrophage with-out inducing host cell death. Infect. Immun., 69, 5529–5537.

51. Prindiville, T.P., Sheikh, R.A., Cohen, S.H., Tang, Y.J., Cantrell, M.C. & Silva, J.Jr. (2000) Bacteroides fragilis enterotoxin gene sequences in patientswith inflammatory bowel disease. Emerg. Infect. Dis., 6, 171–174.

52. Duchmann, R., May, E., Heike, M., Knoll, C.P., Neurath, K.H. & Meyer zumBuschenfelde, K.H. (1999) T cell specificity and cross reactivity towards entero-bacteria, Bacteroides, Bifidobacterium and antigens from resident intestinalflora in humans. Gut, 44, 812–818.



53. Muyzer, G. & Smalla, K. (1998) Application of denaturing gradient gel electro-phoresis (DGGE) and temperature gradient gel electrophoresis (TGGE) inmicrobial ecology. Antonie van Leeuwenhoek, 73, 127–141.

54. Rhodes, J.M. (1997) Colonic mucus and ulcerative colitis. Gut, 40,807–808.

55. Buisine, M.P., Desreumaux, P., Leteurtre, E., Copin, M.C., Colombel, J.F.,Porchet, N. & Aubert, J.P. (2001) Mucin gene expression in intestinal epithelialcells in Crohn’s disease. Gut, 49, 544–551.

56. Favier, C., Neut, C., Mizon, C., Cortot, A., Colombel, J.F. & Mizon, J. (1997)Fecal b-D-galactosidase production and Bifidobacteria are decreased in Crohn’sdisease. Dig. Dis. Sci., 42, 817–822.

57. Madsen, K.L., Doyle, J.S., Jewell, L.D., Tavernini, M.M. & Fedorak, R.N.(1999) Lactobacillus species prevents colitis in interleukin 10 gene-deficientmice. Gastroenterology, 116, 1107–1114.

58. Gionchetti, P., Rizzello, F., Venturi, A. & Campieri, M. (2000) Probiotics ininfective diarrhoea and inflammatory bowel diseases. J. Gastroenterol.Hepatol., 15, 489–493.

59. Alvarez-Olmos, M.I. & Oberhelman, R.A. (2001) Probiotic agents and infectious diseases: a modern perspective on a traditional therapy. Clin. Infect.Dis., 32, 1567–1576.

60. Ulisse, S., Gionchetti, P., D’Alo, S., Russo, F.P., Pesce, I., Ricci, G., Rizzello, F., Helwig, U., Cifone, M.G., Campieri, M. & De Simone, C. (2001)Expression of cytokines, inducible nitric oxide synthase, and matrix metallo-proteinases in pouchitis: effects of probiotic treatment. Am. J. Gastroenterol.,96, 2691–2699.

61. Blumberg, R.S., Saubermann, L.J. & Strober, W. (1999) Animal models ofmucosal inflammation and their relation to human inflammatory bowel disease.Curr. Opin. Immunol., 11, 648–656.

62. Okamoto, S., Watanabe, M., Yamazaki, M., Yajima, T., Hayashi, T., Ishii, H.,Mukai, M., Yamada, T., Watanabe, N., Jameson, BA. & Hibi, T. (1999) A syn-thetic mimetic of CD4 is able to suppress disease in a rodent model of immunecolitis. Eur. J. Immunol., 29, 355–366.

63. Pizarro, T.T., Arseneau, K.O. & Cominelli, F. (2000) Lessons from geneticallyengineered animal models XI. Novel mouse models to study pathogenic mech-anisms of Crohn’s disease. Am. J. Physiol. Gastrointest. Liver Physiol., 278,G665–669.

64. Plevy, S.E., Landers, C.J., Prehn, J., Carramanzana, N.M., Deem, R.L., Shealy, D. & Targan, S.R (1997) A role for TNF alpha and mucosal Th1 cytokines in the pathogenesis of Crohn’s disease. J. Immunol., 159,6276–6282.

65. Neurath, M.F., Finotto, S., Fuss, I., Boirivant, M., Galle, P.R. & Strober, W.(2001) Regulation of T cell apoptosis in IBD: to die or not to die that is themucosal question. Trends Immunol., 22, 21–26.

66. Ueyama, H., Kiyohara, T., Sawada, N., Isozaki, K., Kitamura, S., Kondo, S.,Miyagawa, J., Kanayama, S., Shinomura, Y., Ishikawa, H., Ohtani, T., Nezu, R., Nagata, S. & Matsuzawa, Y. (1998) High Fas ligand expression onlymphocytes in lesions of ulcerative colitis. Gut, 43, 48–55.

67. Kohout, P. & Koref, P. (2001) Intestinal permeability in relatives of patientswith Crohn’s disease. Vnitr. Lek., 47, 371–374.



68. Irvine, E.J. & Marshall, J.K. (2000) Increased intestinal permeability precedesthe onset of Crohn’s disease in a subject with familial risk. Gastroenterology,119, 1740–1744.

69. Zareie, M., Singh, P.K., Irvine, E.J., Sherman, P.M., McKay, D.M. & Perdue,M.H. (2001) Monocyte/macrophage activation by normal bacteria and bacterialproducts. Implications for altered epithelial function in Crohn’s disease. Am. J.Pathol., 158, 1101–1109.

70. Gassler, N., Rohr, C., Schneider, A., Kartenbeck, J., Bach, A., Obermuller, N.,Otto, H.F. & Autschbach, F. (2001) Inflammatory bowel disease is associatedwith changes of enterocyte junctions. Am. J. Physiol. Gastrointest. LiverPhysiol., 281, G216–G228.

71. Cario, E. & Podolsky, D.K. (2000) Differential alteration in intestinal epithelialcell expression of Toll-like receptor 3 (TLR 3) and TLR4 in inflammatorybowel disease. Infect. Immun., 68, 7010–7117.

72. Abreu, M.T., Vora, P., Faure, E., Thomas, L.S., Arnold, E.T. & Arditi, M.(2001) Decreased expression of Toll-like receptor 4 and MD2 correlate withintestinal epithelial cell protection against dysregulated proinflammatory geneexpression in response to bacterial lipopolysaccharide. J. Immunol., 167,1609–1617.



inflammatory bowel disease: is the intestine a trojan horse?

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