Terez-Shea Donohue Department of Medicine Mucosal Biology Research Center University of Maryland School of Medicine

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Terez-Shea Donohue Department of Medicine Mucosal Biology Research Center University of Maryland School of Medicine

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1. Terez-Shea Donohue Department of Medicine Mucosal Biology Research Center University of Maryland School of Medicine

2. GENETIC SUSCEPTIBILITY 126. GENETIC SUSCEPTIBILITY Twin studies provide the most compelling evidence for a genetic influence in IBD. Concordance of disease in monozygotic twins approaches 50% in Crohn’s disease, but ranges from 5-14% for ulcerative colitis. These rates, however, are higher than the concordance rate in dizygotic twins, which is no different from the expected rate in siblings of approximately 5%. Although genetic influences are clear, these studies also indicate important environmental inputs, particularly in ulcerative colitis where no concordance of disease is found in the vast majority of patients with identical genetic background. • Tysk C et al. Ulcerative colitis and Crohn’s disease in an unselected population of monozygotic and dizygotic twins. A study of heritibility and the influence of smoking. Gut 1988;29: 990-6. • Orholm M et al. Concordance of IBS among Danish twins. Results of a nationwide study. Scand J Gastroerterol. 1989;84: 794-7.126. GENETIC SUSCEPTIBILITY Twin studies provide the most compelling evidence for a genetic influence in IBD. Concordance of disease in monozygotic twins approaches 50% in Crohn’s disease, but ranges from 5-14% for ulcerative colitis. These rates, however, are higher than the concordance rate in dizygotic twins, which is no different from the expected rate in siblings of approximately 5%. Although genetic influences are clear, these studies also indicate important environmental inputs, particularly in ulcerative colitis where no concordance of disease is found in the vast majority of patients with identical genetic background. • Tysk C et al. Ulcerative colitis and Crohn’s disease in an unselected population of monozygotic and dizygotic twins. A study of heritibility and the influence of smoking. Gut 1988;29: 990-6. • Orholm M et al. Concordance of IBS among Danish twins. Results of a nationwide study. Scand J Gastroerterol. 1989;84: 794-7.

3. Impact of Race and Ethnicity African American and Caucasiand primaliy had CD while Mexican americans had UC.. P-anca predicotr of pouchitis immune response variations across ethnic groups. African American and Caucasiand primaliy had CD while Mexican americans had UC.. P-anca predicotr of pouchitis immune response variations across ethnic groups.

4. IBD Antibodies Sensitivity varies among population Positive pANCA with negative ASCA IgA/IgG indicates Ulcerative colitis vs Crohn’s disease pANCA – antineutrophil cytoplasmic antibody Positive Crohn’s patients – lack of ileal involvement Caucasian patients Ulcerative colitis patients 59–84% Crohn’s Disease 10-40% controls 10% ASCA – antisaccharomyces cerevisiae antibody Usually measured with IgA or IgG Increased titers indicated ileal involvement Crohn’s Disease 50-60%

5. Genetics of IBD Familial occurrence (15-20% patients have affected relative) Genetic influence is lower in ulcerative colitis than in Crohn’s disease Human gene testing is looking for markers Genes coded by chromosomes (1,5,6,10,12,16, and 19) Nucleotide-binding oligomerization domain Nod2 (CARD15) mutations on chromosome 16 Nod1 (CARD4) Cytokine cluster region on chromosome 5 IBD5 locus on chromosome 5q31 OCTN1 and 2 – carnitine/organic ion transporter thought to be IBD5 susceptibility genes DLG5 – on chromosome 10q23, encodes epithelial scaffolding protein Correlates between genotype and disease phenotype (e.g Nod2 and fibrostentic small intestinal disease) The research on genetic susceptibility of inflammatory bowel diseases (IBD) has been tremendous and over 10 chromosomal regions have been identified by genome-wide scanning. Further fine mapping as well as candidate gene studies have already led to the identification of a number of susceptibility genes including CARD15, DLG5, OCTN1 and 2, NOD1, HLA, and TLR4. The CARD15 gene is undoubtedly replicated most widely and most understood at present. CARD15 is involved in the recognition of bacterial peptidoglycan-derived muramyl dipeptide (MDP) and will stimulate secretion of antimicrobial peptides including alpha-defensins (also called cryptdins) to protect the host from invasion. Genetic research in IBD has advanced our understanding of the clinical heterogeneity of the disease and has started to tackle the complex interactions between genetic risk factors and environmental risk factors in IBD. Genes also interfere with the metabolization of drugs and may influence the clinical response and the drug-related toxicity. Interesting pharmacogenetic data with respect to steroids, azathioprine, and infliximab have been generated in IBD. Overall, it is anticipated that genetic markers in the future will be implemented in an integrated molecular diagnostic and prognostic approach of our patients Many of these genes are involve in mucosal function particularly mucosal barrier function. The genetic influence of Crohn’s disease is greater than that of ulcerative colitis, although in both disorders there is a 15-25% likelihood of a patient having an affected relative, usually a first degree relative. Familial patterns of disease location or phenotype (aggressive fistulizing vs. slowly progressive, stenotic disease) exist in Crohn’s disease. Clinical studies indicate a polygenic susceptibility with a pattern of complex genetic traits. Genome wide searches have implicated almost all chromosomes, but at least three separate studies have confirmed involvement of chromosomes 16, 12, 6 and 5. In 2002, three independent groups reported single nucleotide polymorphisms in the NOD2 (CARD 15) gene on chromosome 16 in Crohn’s disease. This gene recognizes intracellular lipopolysaccharide (endotoxin) from Gram negative bacteria and transduces signals activating NF?B and initiating apoptosis. A separate region on chromosome 5 that regulates production of multiple cytokines was also found to be associated with Crohn’s disease. Finally, multiple studies are finding correlations between genotype and disease phenotype, particularly in natural history and response to therapy. For example NOD2/CARD 15 variants are associated with fibrostenotic small intestinal disease. • Hugot JP et al. Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature 2001;411: 599-603. • Ogura Y et al. A frame shift mutation in NOD2 associated with Crohn’s disease. Nature 2001;411:603-6. • Cuthbert AP, Fisher SA, Mirza MM et al. The contribution of NOD2 gene mutations to the risk and site of disease in inflammatory bowel disease. Gastroenterology 2002;122:867-74. • Ahmad T. Armuzzi A, Bunce M et al. The molecular classification of the clinical manifestations of Crohn’s disease. Gastroenterology 2002;122: 854-66. Population studies confirm a role for genetic traits in suceptibility to IBD. However, inheritance does not follow simple Mendelian genetics and the role for non-genetic, environmental ators is large in UC. The aim of genetic marker studies is to identify the site of the genes that predispose to IBD. Chromosome 6 has MHC containing regions for HLA-DR2 and other genes involves in the regulation of inflammation. MHC major role is to present peptide fragments of antigens to T cells. So abnomalitiy may cause frmation of a pathogenci antibody. Chromosome 2 has IL-1ra . Gene 7 has MUC3 which code for certain classes of intestinal mucin glycoproteins. CF loci is in this chromosome. Prevalence rate for IBD in CF patients is 7 times that in controls. PANCA presnt in about 60-80% of UC and 20-30% of CD patients.The research on genetic susceptibility of inflammatory bowel diseases (IBD) has been tremendous and over 10 chromosomal regions have been identified by genome-wide scanning. Further fine mapping as well as candidate gene studies have already led to the identification of a number of susceptibility genes including CARD15, DLG5, OCTN1 and 2, NOD1, HLA, and TLR4. The CARD15 gene is undoubtedly replicated most widely and most understood at present. CARD15 is involved in the recognition of bacterial peptidoglycan-derived muramyl dipeptide (MDP) and will stimulate secretion of antimicrobial peptides including alpha-defensins (also called cryptdins) to protect the host from invasion. Genetic research in IBD has advanced our understanding of the clinical heterogeneity of the disease and has started to tackle the complex interactions between genetic risk factors and environmental risk factors in IBD. Genes also interfere with the metabolization of drugs and may influence the clinical response and the drug-related toxicity. Interesting pharmacogenetic data with respect to steroids, azathioprine, and infliximab have been generated in IBD. Overall, it is anticipated that genetic markers in the future will be implemented in an integrated molecular diagnostic and prognostic approach of our patients Many of these genes are involve in mucosal function particularly mucosal barrier function. The genetic influence of Crohn’s disease is greater than that of ulcerative colitis, although in both disorders there is a 15-25% likelihood of a patient having an affected relative, usually a first degree relative. Familial patterns of disease location or phenotype (aggressive fistulizing vs. slowly progressive, stenotic disease) exist in Crohn’s disease. Clinical studies indicate a polygenic susceptibility with a pattern of complex genetic traits. Genome wide searches have implicated almost all chromosomes, but at least three separate studies have confirmed involvement of chromosomes 16, 12, 6 and 5. In 2002, three independent groups reported single nucleotide polymorphisms in the NOD2 (CARD 15) gene on chromosome 16 in Crohn’s disease. This gene recognizes intracellular lipopolysaccharide (endotoxin) from Gram negative bacteria and transduces signals activating NF?B and initiating apoptosis. A separate region on chromosome 5 that regulates production of multiple cytokines was also found to be associated with Crohn’s disease. Finally, multiple studies are finding correlations between genotype and disease phenotype, particularly in natural history and response to therapy. For example NOD2/CARD 15 variants are associated with fibrostenotic small intestinal disease. • Hugot JP et al. Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature 2001;411: 599-603. • Ogura Y et al. A frame shift mutation in NOD2 associated with Crohn’s disease. Nature 2001;411:603-6. • Cuthbert AP, Fisher SA, Mirza MM et al. The contribution of NOD2 gene mutations to the risk and site of disease in inflammatory bowel disease. Gastroenterology 2002;122:867-74. • Ahmad T. Armuzzi A, Bunce M et al. The molecular classification of the clinical manifestations of Crohn’s disease. Gastroenterology 2002;122:854-66. Population studies confirm a role for genetic traits in suceptibility to IBD. However, inheritance does not follow simple Mendelian genetics and the role for non-genetic, environmental ators is large in UC. The aim of genetic marker studies is to identify the site of the genes that predispose to IBD. Chromosome 6 has MHC containing regions for HLA-DR2 and other genes involves in the regulation of inflammation. MHC major role is to present peptide fragments of antigens to T cells. So abnomalitiy may cause frmation of a pathogenci antibody. Chromosome 2 has IL-1ra . Gene 7 has MUC3 which code for certain classes of intestinal mucin glycoproteins. CF loci is in this chromosome. Prevalence rate for IBD in CF patients is 7 times that in controls. PANCA presnt in about 60-80% of UC and 20-30% of CD patients.

6. A is for AUTOIMMUNE

7. AUTOIMMUNE DISEASES Genetic susceptibility for the host immune system to recognize, and potentially misinterpret, an environmental antigen presented to the body (eg., within the GI tract). The host must be exposed to the antigen. The antigen must be presented to the mucosal immune system following its paracellular passage (normally prevented by tight junction competency) from the lumen to the submucosa.

8. NOD2 (CARD15) First susceptibility gene for Crohn’s Disease (2002) Strong association for small bowel Phenotype - associated with fibrostenotic small intestinal disease Attributable risk Crohn’s ileitis ~ 30% Ulcerative colitis ~ 6% Risk by population Causcasian - 19.1-29% Ashkenazi Jews 47.4% Sephardic Jews 27.45% African Americans 2.6% Asian (Chinese, Korean, Japanese) - ND recognizes intracellular lipopolysaccharide (endotoxin) from Gram negative bacteria and transduces signals activating NF?B and initiating apoptosisNOD-2 variant associateion with IBD beter in whites. Although phenotypically similar, Isreali arabs have a very low incidence when compare to Isreali jews. recognizes intracellular lipopolysaccharide (endotoxin) from Gram negative bacteria and transduces signals activating NF?B and initiating apoptosisNOD-2 variant associateion with IBD beter in whites. Although phenotypically similar, Isreali arabs have a very low incidence when compare to Isreali jews.

9. Other Genes Involved in Mucosal Function OCTN – organic cation transporter gene located on 5q31, mutations at this loci affect the ability of the transporters to pump xenobiotics and amino acids DLG5 – membrane scaffold protein located on 10q23, muations impart the ability to maintain epithelial polarity Both genes important in epithelial permeability and disruption may facilitate exposure to bacterial products Two other genes associated with Crohn's disease have been identified. The first of these, located on 5q31, encodes the organic cation transporter (OCTN) genes, and mutations at these loci affect the ability of the transporters to pump xenobotics and amino acids across cell membranes (60). In the gut, these genes are expressed in epithelial cells, macrophages, and T cells, correlating closely with their potential function in IBD. The second gene is located on 10q23 and encodes the guanylate kinase DLG5 (61). The mutation in this gene involves a single amino acid substitution that is thought to impair the ability of DLG5 to maintain epithelial polarity. Both genes may be important in epithelial permeability, and disruption of this function could lead to inappropriate exposure of the mucosal immune system to bacterial products. Two other genes associated with Crohn's disease have been identified. The first of these, located on 5q31, encodes the organic cation transporter (OCTN) genes, and mutations at these loci affect the ability of the transporters to pump xenobotics and amino acids across cell membranes (60). In the gut, these genes are expressed in epithelial cells, macrophages, and T cells, correlating closely with their potential function in IBD. The second gene is located on 10q23 and encodes the guanylate kinase DLG5 (61). The mutation in this gene involves a single amino acid substitution that is thought to impair the ability of DLG5 to maintain epithelial polarity. Both genes may be important in epithelial permeability, and disruption of this function could lead to inappropriate exposure of the mucosal immune system to bacterial products.

10. AUTOIMMUNE DISEASES Genetic susceptibility for the host immune system to recognize, and potentially misinterpret, an environmental antigen presented to the body (eg., within the GI tract). The host must be exposed to the antigen. The antigen must be presented to the mucosal immune system following its paracellular passage (normally prevented by tight junction competency) from the lumen to the submucosa.

11. 123. NORMAL MUCOSAL DEFENSES Normally an intact mucus layer and epithelial tight junctions combine to prevent uptake of luminal antigen and translocation of viable bacteria. Goblet cells secrete mucus which covers the epithelial cell as well as intestinal trefoil factor, which increases the viscosity of mucus as well as stimulates epithelial restitution, which is the ability of epithelial cells to rapidly migrate to cover a denuded surface, thereby restoring epithelial integrity. Secreted growth factors (keratinocyte growth factor [KGF] and transforming growth factor ? [TGF?]) from fibroblasts and lamina propria mononuclear cells also enhance epithelial restitution. In addition, secreted products, including defensins made by epithelial cells and Paneth cells and secretory IgA (sIgA) from plasma cells, provide protection against bacteria. Defensins lyse bacteria while secretory IgA complexes bacterial and dietary antigens to prevent uptake. Finally, protective prostaglandins, including PGE2, from mesenchymal cells (fibroblasts, smooth muscle cells and myofibroblasts) as well as activated immune cells enhance epithelial barrier function, stimulate secretion of chloride which washes away luminal pathogens and increase mucosal blood flow.123. NORMAL MUCOSAL DEFENSES Normally an intact mucus layer and epithelial tight junctions combine to prevent uptake of luminal antigen and translocation of viable bacteria. Goblet cells secrete mucus which covers the epithelial cell as well as intestinal trefoil factor, which increases the viscosity of mucus as well as stimulates epithelial restitution, which is the ability of epithelial cells to rapidly migrate to cover a denuded surface, thereby restoring epithelial integrity. Secreted growth factors (keratinocyte growth factor [KGF] and transforming growth factor ? [TGF?]) from fibroblasts and lamina propria mononuclear cells also enhance epithelial restitution. In addition, secreted products, including defensins made by epithelial cells and Paneth cells and secretory IgA (sIgA) from plasma cells, provide protection against bacteria. Defensins lyse bacteria while secretory IgA complexes bacterial and dietary antigens to prevent uptake. Finally, protective prostaglandins, including PGE2, from mesenchymal cells (fibroblasts, smooth muscle cells and myofibroblasts) as well as activated immune cells enhance epithelial barrier function, stimulate secretion of chloride which washes away luminal pathogens and increase mucosal blood flow.

12. HOST DEFENSE ALERT Show one at a timeShow one at a time

13. Stimulation Of Immune Responses In The Normal Host 139. STIMULATION OF IMMUNE RESPONSES IN THE NORMAL HOST The abiity to discriminate between commensal and pathogenic bacteria may be due to continues exposure. Normal hosts, which are resistant to inflammation, have controlled uptake of luminal antigens across the specialize epithelium of Peyer’s patches and colonic lymphoid aggregates to initiate a regulated immune response. These antigens cross microfold (M) cells in the dome epithelium without being processed and then are degraded and presented by dendritic cells within the Peyer’s patch. Naïve (TH0) T helper cells differentiate into regulatory T cell subsets, TH3 and T regulatory 1 (TR1), which preferentially secrete TGF? and IL-10, respectively. These immunosuppressive cytokines inhibit the activity of macrophages and T helper 1 cells, thereby preventing secretion of IL-12 and interferon ? (IFN?). This regulatory tone of the intestine prevents induction of pathologic immune responses to commensal bacteria and dietary antigens.139. STIMULATION OF IMMUNE RESPONSES IN THE NORMAL HOST The abiity to discriminate between commensal and pathogenic bacteria may be due to continues exposure. Normal hosts, which are resistant to inflammation, have controlled uptake of luminal antigens across the specialize epithelium of Peyer’s patches and colonic lymphoid aggregates to initiate a regulated immune response. These antigens cross microfold (M) cells in the dome epithelium without being processed and then are degraded and presented by dendritic cells within the Peyer’s patch. Naïve (TH0) T helper cells differentiate into regulatory T cell subsets, TH3 and T regulatory 1 (TR1), which preferentially secrete TGF? and IL-10, respectively. These immunosuppressive cytokines inhibit the activity of macrophages and T helper 1 cells, thereby preventing secretion of IL-12 and interferon ? (IFN?). This regulatory tone of the intestine prevents induction of pathologic immune responses to commensal bacteria and dietary antigens.

14. ETIOLOGIC HYPOTHESES 122. ETIOLOGIC HYPOTHESES There are four widely accepted etiologic hypotheses for these idiopathic disorders. Each of these hypotheses involve microbial agents. The simplest and most amenable to curative treatment is that a viable pathogen persistently infects the intestine. The most widely studied pathogen is Mycobacterium paratuberculosis which causes naturally occurring Johne’s disease, a chronic progressive granulomatous enterocolitis in ruminants (cattle, sheep, goats and deer). Intestinal Helicobacter species (non H. pylori species) can cause colitis in immunodeficient mice and some genetically engineered mice, including those with IL-10 deficiency, but have not been implicated in human disease. Measles, mumps and Listeria have their advocates, but have not been widely reproduced. Similarly, toxigenic E. coli have been isolated from patients with both ulcerative colitis and Crohn’s disease but need to be more thoroughly investigated. Dysbiosis is an altered balance of protective bacteria (Lactobacillus and Bifidobacterium species) and aggressive commensal organisms (including Bacteroides, Enterococcus, inherent/invasive E. coli). Defective mucosal integrity could lead to enhanced uptake of commensal luminal bacteria which could then overwhelm the normally protective activities of the mucosal immune system. Decreased barrier function could be a result of altered mucus, increased permeability, decreased epithelial metabolic activity, or impaired mucosal healing after injury. Dysregulated immune responses to ubiquitous luminal antigens could lead to tissue injury. This dysregulation could be a result of loss of protective responses leading to defective tolerance to luminal antigens, overally aggressive cell mediated immune function or dysregulated apoptosis of macrophages or T lymphocytes. 122. ETIOLOGIC HYPOTHESES There are four widely accepted etiologic hypotheses for these idiopathic disorders. Each of these hypotheses involve microbial agents. The simplest and most amenable to curative treatment is that a viable pathogen persistently infects the intestine. The most widely studied pathogen is Mycobacterium paratuberculosis which causes naturally occurring Johne’s disease, a chronic progressive granulomatous enterocolitis in ruminants (cattle, sheep, goats and deer). Intestinal Helicobacter species (non H. pylori species) can cause colitis in immunodeficient mice and some genetically engineered mice, including those with IL-10 deficiency, but have not been implicated in human disease. Measles, mumps and Listeria have their advocates, but have not been widely reproduced. Similarly, toxigenic E. coli have been isolated from patients with both ulcerative colitis and Crohn’s disease but need to be more thoroughly investigated. Dysbiosis is an altered balance of protective bacteria (Lactobacillus and Bifidobacterium species) and aggressive commensal organisms (including Bacteroides, Enterococcus, inherent/invasive E. coli). Defective mucosal integrity could lead to enhanced uptake of commensal luminal bacteria which could then overwhelm the normally protective activities of the mucosal immune system. Decreased barrier function could be a result of altered mucus, increased permeability, decreased epithelial metabolic activity, or impaired mucosal healing after injury. Dysregulated immune responses to ubiquitous luminal antigens could lead to tissue injury. This dysregulation could be a result of loss of protective responses leading to defective tolerance to luminal antigens, overally aggressive cell mediated immune function or dysregulated apoptosis of macrophages or T lymphocytes.

15. AUTOIMMUNE DISEASES Genetic susceptibility for the host immune system to recognize, and potentially misinterpret, an environmental antigen presented to the body (eg., within the GI tract). The host must be exposed to the antigen. The antigen must be presented to the mucosal immune system following its paracellular passage (normally prevented by tight junction competency) from the lumen to the submucosa. In many cases, increased intestinal permeability appears to precede disease and causes an abnormality in antigen delivery that triggers the multi-organ process leading to the autoimmune response. In many cases, increased intestinal permeability appears to precede disease and causes an abnormality in antigen delivery that triggers the multi-organ process leading to the autoimmune response.

16. IBD - Defective Mucosal Integrity CD may be considered as a defective immune response, no longer only as hyperresponsiveness of the mucosal immune system Nod2/ CARD15 expression suggest that macrophages and epithelial cells could be the site of a primary pathophysiologic defect, and T-cell activation might just be a secondary effect inducing chronification of the inflammation, perhaps as a backup mechanism to a defective innate immunity

17. Tight Junction Proteins and IBD claudinclaudin

18. B is for BACTERIA

19. Role of Bacteria in the Pathogenesis of Chronic Immune Mediated Intestinal Inflammation 128. ROLE OF BACTERIA IN THE PATHOGENESIS OF CHRONIC IMMUNE MEDIATED INTESTINAL INFLAMMATION The most compelling evidence for a role of bacteria in the pathogenesis of experimental inflammation is provided by the observation that there is a total absence of immune activation and colitis in the absence of luminal bacteria in at least 11 different animal models. Genetically susceptible mice, rats, guinea pigs and tamarins raised in a sterile (germ-free) environment fail to develop colitis, yet littermates colonized with normal enteric bacteria exhibit macrophage and T cell activation, leading to the onset of chronic colitis. • Rath HC et al. Normal luminal bacteria especially bacteroides mediate chronic colitis, gastritis and arthritis in HLA-?27/human B2 microgloblin species transgenic rats. Journal of Clinical Investigation 1996;98:945. • Sellon RK et al. Resident luminal bacteria are necessary for the development of spontaneous colitis and immune system activation in IL-10 deficient mice. Infection and Immunity 1998;66:5224-31.128. ROLE OF BACTERIA IN THE PATHOGENESIS OF CHRONIC IMMUNE MEDIATED INTESTINAL INFLAMMATION The most compelling evidence for a role of bacteria in the pathogenesis of experimental inflammation is provided by the observation that there is a total absence of immune activation and colitis in the absence of luminal bacteria in at least 11 different animal models. Genetically susceptible mice, rats, guinea pigs and tamarins raised in a sterile (germ-free) environment fail to develop colitis, yet littermates colonized with normal enteric bacteria exhibit macrophage and T cell activation, leading to the onset of chronic colitis. • Rath HC et al. Normal luminal bacteria especially bacteroides mediate chronic colitis, gastritis and arthritis in HLA-?27/human B2 microgloblin species transgenic rats. Journal of Clinical Investigation 1996;98:945. • Sellon RK et al. Resident luminal bacteria are necessary for the development of spontaneous colitis and immune system activation in IL-10 deficient mice. Infection and Immunity 1998;66:5224-31.

20. Influence Of Luminal Contents On Reactivation Of Crohn’s Disease 130. INFLUENCE OF LUMINAL CONTENTS ON REACTIVATION OF CROHN’S DISEASE An elegant study by D’Haens et al has demonstrated the critical role of luminal contents in reactivation of Crohn’s disease. One year after segmental resection of the distal ileum and cecum with primary anastomosis recurrence of inflammation, as determined by endoscopic evidence of mucosal ulceration in the neoterminal ileum is present in 85-90% of patients. However, following a similar ileal resection with primary anastomosis but with proximal diversion of luminal contents by a diverting ileostomy, no recurrence is noted by endoscopy. Importantly, recurrence of inflammation is noted less than one month after restoration of exposure to the fecal stream following takedown of the proximal diverting ostomy. D’Haens et al demonstrated that reinfusion of ileostomy contents into the diverted distal ileum led to histologic evidence of inflammation within one week. These observations strongly implicate intestinal luminal contents as the source of the constant antigenic stimulation that leads to recurrent inflammation in the vast majority of Crohn’s disease patients. • D’Haens GR et al. Early lesions of recurrent Crohn’s disease caused by infusion of intestinal luminal contents into excluded ileum. Gastroenterology 1998;114:262.130. INFLUENCE OF LUMINAL CONTENTS ON REACTIVATION OF CROHN’S DISEASE An elegant study by D’Haens et al has demonstrated the critical role of luminal contents in reactivation of Crohn’s disease. One year after segmental resection of the distal ileum and cecum with primary anastomosis recurrence of inflammation, as determined by endoscopic evidence of mucosal ulceration in the neoterminal ileum is present in 85-90% of patients. However, following a similar ileal resection with primary anastomosis but with proximal diversion of luminal contents by a diverting ileostomy, no recurrence is noted by endoscopy. Importantly, recurrence of inflammation is noted less than one month after restoration of exposure to the fecal stream following takedown of the proximal diverting ostomy. D’Haens et al demonstrated that reinfusion of ileostomy contents into the diverted distal ileum led to histologic evidence of inflammation within one week. These observations strongly implicate intestinal luminal contents as the source of the constant antigenic stimulation that leads to recurrent inflammation in the vast majority of Crohn’s disease patients. • D’Haens GR et al. Early lesions of recurrent Crohn’s disease caused by infusion of intestinal luminal contents into excluded ileum. Gastroenterology 1998;114:262.

21. Regulation Of Experimental Colitis By T Cell Subsets 142. REGULATION OF EXPERIMENTAL COLITIS BY T CELL SUBSETS Cong and Elson have demonstrated that bacterial antigen-specific T cell clones can both induce and prevent experimental colitis. These investigators isolated TH1 and T regulatory 1 (TR1) clones from C3H HeJ Bir mice with colitis. When stimulated in vitro with lysates of cecal contents, the TH1 clones produced interferon ? and the TR1 clones produced IL-10. When transferred into immunodeficient (SCID) mice, the TH1 cells produced colitis and the TR1 cells had no pathologic effects. However, when these cells were co-transferred into immunodeficient mice no disease ensued, suggesting that the IL-10- secreting TR1 cells secreting IL-10 could protect against the detrimental effects of TH1 cells. These results suggest that bacterial antigen-specific regulatory cells mediate protection in normal hosts. • Cong Y et al. Enteric bacterial antigen specific TR, cells regulate CD4+ T cell induced colitis. Gastroenterology 2000;118:683A.142. REGULATION OF EXPERIMENTAL COLITIS BY T CELL SUBSETS Cong and Elson have demonstrated that bacterial antigen-specific T cell clones can both induce and prevent experimental colitis. These investigators isolated TH1 and T regulatory 1 (TR1) clones from C3H HeJ Bir mice with colitis. When stimulated in vitro with lysates of cecal contents, the TH1 clones produced interferon ? and the TR1 clones produced IL-10. When transferred into immunodeficient (SCID) mice, the TH1 cells produced colitis and the TR1 cells had no pathologic effects. However, when these cells were co-transferred into immunodeficient mice no disease ensued, suggesting that the IL-10- secreting TR1 cells secreting IL-10 could protect against the detrimental effects of TH1 cells. These results suggest that bacterial antigen-specific regulatory cells mediate protection in normal hosts. • Cong Y et al. Enteric bacterial antigen specific TR, cells regulate CD4+ T cell induced colitis. Gastroenterology 2000;118:683A.

22. Role Of Individual Bacteria In The Pathogenesis Of Experimental Colitis 131. ROLE OF INDIVIDUAL BACTERIA IN THE PATHOGENESIS OF EXPERIMENTAL COLITIS HLA B27/?2 microglobulin transgenic rats raised under sterile (germ-free) conditions do not develop colitis. However, when colonized with normal specific pathogen free commensal cecal bacteria, they develop aggressive disease involving the colon, antrum, and duodenum within one month. When colonized (monoassociated) with a single bacterial strain, Bacteroides vulgatus, these susceptible rats develop moderate colitis but no gastroduodenal disease. However, when similarly mono-associated with E. coli, they exhibit neither disease nor T cell activation. However, some bacteria, including Lactobacillus GG, actually prevent inflammation by attenuating the colitis in HLA B27 transgenic rats colonized with normal cecal bacteria. These results indicate that all commensal bacterial species are not identical in their abilities to induce disease: some are aggressive, such as Bacteroides vulgatus; some are neutral, including E. coli; and some, like Lactobacillus species, are protective. • Rath HC et al. Differential induction of colitis and gastritis in HLA ?27 transgenic rats selectively colonized with bacteroides vulgatus and Eschericia coli. Infection and Immunity 1999;67:2969. • Dieleman LA et al. Lactobacillus GG prevents recurrence of colitis in HLA B27 transgenic rats after antibiotic treatment. Gastroenterology 2000;118:A814.131. ROLE OF INDIVIDUAL BACTERIA IN THE PATHOGENESIS OF EXPERIMENTAL COLITIS HLA B27/?2 microglobulin transgenic rats raised under sterile (germ-free) conditions do not develop colitis. However, when colonized with normal specific pathogen free commensal cecal bacteria, they develop aggressive disease involving the colon, antrum, and duodenum within one month. When colonized (monoassociated) with a single bacterial strain, Bacteroides vulgatus, these susceptible rats develop moderate colitis but no gastroduodenal disease. However, when similarly mono-associated with E. coli, they exhibit neither disease nor T cell activation. However, some bacteria, including Lactobacillus GG, actually prevent inflammation by attenuating the colitis in HLA B27 transgenic rats colonized with normal cecal bacteria. These results indicate that all commensal bacterial species are not identical in their abilities to induce disease: some are aggressive, such as Bacteroides vulgatus; some are neutral, including E. coli; and some, like Lactobacillus species, are protective. • Rath HC et al. Differential induction of colitis and gastritis in HLA ?27 transgenic rats selectively colonized with bacteroides vulgatus and Eschericia coli. Infection and Immunity 1999;67:2969. • Dieleman LA et al. Lactobacillus GG prevents recurrence of colitis in HLA B27 transgenic rats after antibiotic treatment. Gastroenterology 2000;118:A814.

23. GERMETICS

24. Nod2 - present in macrophages, dendritic and Paneth cells, and can be induced in enterocytes. Its activation by bacterial muramyl dipeptide induces expression of proinflammatory mediators. Nod1 - expressed in intestinal epithelial cells and activates proinflammatory cytokine production in response to a peptidoglycan motif in gram-negative bacteria. CARD15/NOD2, there are additional "innate" pathways by which commensal and pathogenic bacteria can directly interact with cells of the intestinal mucosa (eg, toll-like receptors). CARD15/NOD2, there are additional "innate" pathways by which commensal and pathogenic bacteria can directly interact with cells of the intestinal mucosa (eg, toll-like receptors).

25. Nod2 and Innate Immunity Transduces signals activating NF?? and initiating apoptosis CARD15/NOD2 function and NF?? activation indicate that an inflammatory reaction of the intestinal mucosa as a response of the innate immune system may be necessary for the maintenance of gut homeostasis. CARD15/NOD2, there are additional "innate" pathways by which commensal and pathogenic bacteria can directly interact with cells of the intestinal mucosa (eg, toll-like receptors). CARD15/NOD2, there are additional "innate" pathways by which commensal and pathogenic bacteria can directly interact with cells of the intestinal mucosa (eg, toll-like receptors).

26. PANETH CELLS Paneth cells are upregulated in response to mucosal stressors such as IR or enteric infection. They serve an important role in innate immunity particularly in enteric antibacterial defense. They contain antimicrobial molecules including ?-defensins (cryptidins) and mice deficient in cryptidins have impaired clearance of Gram-negative bacteria (Wilson CL et al., 1999). Nucleotide-binding oligomerization domain (NOD) 2 is present in macrophages, dendritic and Paneth cells, and can be induced in enterocytes. Mutations in NOD2 are highly associated with the development of Crohn’s disease (Hugot JP et al., 2001). In Paneth cells, NOD2 was located in the cytosol in close proximity to the granules that contain antimicrobial peptides (Ogura Y et al., 2003). Recent studies show that Paneth cells also express TLR-9 and the decreased expression of TLR-9 is consistent with degranulation of Paneth cells (Rumio et al., 2004). In small intestinal Crohn's disease, Paneth cell depletion occurs in the most heavily inflamed areas, but occasional specimens show increased numbers and Paneth cell degranulation has been observed in cystic fibrosis [11]. Paneth cells are upregulated in response to mucosal stressors such as IR or enteric infection. They serve an important role in innate immunity particularly in enteric antibacterial defense. They contain antimicrobial molecules including ?-defensins (cryptidins) and mice deficient in cryptidins have impaired clearance of Gram-negative bacteria (Wilson CL et al., 1999). Nucleotide-binding oligomerization domain (NOD) 2 is present in macrophages, dendritic and Paneth cells, and can be induced in enterocytes. Mutations in NOD2 are highly associated with the development of Crohn’s disease (Hugot JP et al., 2001). In Paneth cells, NOD2 was located in the cytosol in close proximity to the granules that contain antimicrobial peptides (Ogura Y et al., 2003). Recent studies show that Paneth cells also express TLR-9 and the decreased expression of TLR-9 is consistent with degranulation of Paneth cells (Rumio et al., 2004). In small intestinal Crohn's disease, Paneth cell depletion occurs in the most heavily inflamed areas, but occasional specimens show increased numbers and Paneth cell degranulation has been observed in cystic fibrosis [11].

27. Nod2 Functions in Mucosal Defense Nod2 expressed in cytosol of Paneth cells Mutations in Nod2 may confer reduced ability to kill gut bacteria. Mice deficient in NOD2 show increase susceptibility to Listeria moncytogenes Nod2 is expressed in the cytosol of gut epithelial cells, macrophages, and DC (8). After ligand binding, Nod2 oligomerizes and recruits RICK/RIP, which leads to phosphorylation and degradation of I B and activation of NF- B (50). A consequence of mutations in Nod2 may therefore be a decreased ability to kill gut bacteria (51). Consistent with this, monocytes from patients with common Crohn's disease mutations show defective activation of NF- B and IL-8 secretion when stimulated with MDP (52). Interestingly, Nod2-deficient mice do not develop spontaneous gut inflammation (53, 54), and their macrophages show normal NF- B activation and pro-inflammatory cytokine production to ligands for TLR3, 4, and 9. However, they secrete large amounts of IL-12 in response to TLR2 ligands (55). Concomitant activation through Nod2 inhibits TLR2 induction of IL-12 in cells from wild-type mice but has no effect in Nod2-deficient mice, suggesting that enhanced IL-12 production in Crohn's disease may be due to a failure of Nod2 to negatively regulate TLR2 signaling, which then facilitates a Th1 response through IL-12. It is not known whether Nod2-mediated negative regulation of TLR2 signaling occurs in Crohn's patients bearing the common mutations. In other studies, macrophages from mice engineered to express the 3020insC Nod2 mutation show increased activation of NF- B and increased IL-1ß and IL-6 production when activated with MDP, which suggests that Nod2 mutations may, in some situations, lead to a gain of function and increased pro-inflammatory cytokine production (56). Nod2-deficient mice are also unusually susceptible to intestinal infection with the pathogen Listeria monocytogenes (54) and have been found to be deficient in cryptdin 4 and 10, bacteriacidal defensins produced by Paneth cells found in small intestinal crypts (57). In the human gut, Nod2 is highly expressed in Paneth cells (58), and reduced defensin expression has been reported in Crohn's patients with Nod2 mutations (59). Nod2 mutations may thus also predispose to Crohn's disease indirectly by reducing defensin-mediated innate antimicrobial immunity. Paneth cells are upregulated in response to mucosal stressors such as IR or enteric infection. They serve an important role in innate immunity particularly in enteric antibacterial defense. They contain antimicrobial molecules including ?-defensins (cryptidins) and mice deficient in cryptidins have impaired clearance of Gram-negative bacteria (Wilson CL et al., 1999). Nucleotide-binding oligomerization domain (NOD) 2 is present in macrophages, dendritic and Paneth cells, and can be induced in enterocytes. Mutations in NOD2 are highly associated with the development of Crohn’s disease (Hugot JP et al., 2001). In Paneth cells, NOD2 was located in the cytosol in close proximity to the granules that contain antimicrobial peptides (Ogura Y et al., 2003). Recent studies show that Paneth cells also express TLR-9 and the decreased expression of TLR-9 is consistent with degranulation of Paneth cells (Rumio et al., 2004). In small intestinal Crohn's disease, Paneth cell depletion occurs in the most heavily inflamed areas, but occasional specimens show increased numbers and Paneth cell degranulation has been observed in cystic fibrosis [11]. Mice deficient in NOD2 show increase susceptibility to Listeria moncytogenes Nod2 is expressed in the cytosol of gut epithelial cells, macrophages, and DC (8). After ligand binding, Nod2 oligomerizes and recruits RICK/RIP, which leads to phosphorylation and degradation of I B and activation of NF- B (50). A consequence of mutations in Nod2 may therefore be a decreased ability to kill gut bacteria (51). Consistent with this, monocytes from patients with common Crohn's disease mutations show defective activation of NF- B and IL-8 secretion when stimulated with MDP (52). Interestingly, Nod2-deficient mice do not develop spontaneous gut inflammation (53, 54), and their macrophages show normal NF- B activation and pro-inflammatory cytokine production to ligands for TLR3, 4, and 9. However, they secrete large amounts of IL-12 in response to TLR2 ligands (55). Concomitant activation through Nod2 inhibits TLR2 induction of IL-12 in cells from wild-type mice but has no effect in Nod2-deficient mice, suggesting that enhanced IL-12 production in Crohn's disease may be due to a failure of Nod2 to negatively regulate TLR2 signaling, which then facilitates a Th1 response through IL-12. It is not known whether Nod2-mediated negative regulation of TLR2 signaling occurs in Crohn's patients bearing the common mutations. In other studies, macrophages from mice engineered to express the 3020insC Nod2 mutation show increased activation of NF- B and increased IL-1ß and IL-6 production when activated with MDP, which suggests that Nod2 mutations may, in some situations, lead to a gain of function and increased pro-inflammatory cytokine production (56). Nod2-deficient mice are also unusually susceptible to intestinal infection with the pathogen Listeria monocytogenes (54) and have been found to be deficient in cryptdin 4 and 10, bacteriacidal defensins produced by Paneth cells found in small intestinal crypts (57). In the human gut, Nod2 is highly expressed in Paneth cells (58), and reduced defensin expression has been reported in Crohn's patients with Nod2 mutations (59). Nod2 mutations may thus also predispose to Crohn's disease indirectly by reducing defensin-mediated innate antimicrobial immunity. Paneth cells are upregulated in response to mucosal stressors such as IR or enteric infection. They serve an important role in innate immunity particularly in enteric antibacterial defense. They contain antimicrobial molecules including ?-defensins (cryptidins) and mice deficient in cryptidins have impaired clearance of Gram-negative bacteria (Wilson CL et al., 1999). Nucleotide-binding oligomerization domain (NOD) 2 is present in macrophages, dendritic and Paneth cells, and can be induced in enterocytes. Mutations in NOD2 are highly associated with the development of Crohn’s disease (Hugot JP et al., 2001). In Paneth cells, NOD2 was located in the cytosol in close proximity to the granules that contain antimicrobial peptides (Ogura Y et al., 2003). Recent studies show that Paneth cells also express TLR-9 and the decreased expression of TLR-9 is consistent with degranulation of Paneth cells (Rumio et al., 2004). In small intestinal Crohn's disease, Paneth cell depletion occurs in the most heavily inflamed areas, but occasional specimens show increased numbers and Paneth cell degranulation has been observed in cystic fibrosis [11].

28. Nod1 and Innate Immunity Nod-1 acts as a cytoplasmic sensor for bacteria and their products Important in defense against H. pylori and Shigella flexneri Associated with asthma Early diagnosis (< 25 years old) was strongly associated with NOd1, Also associated with EIM (large and small joint arithritis, ankylosing spondylitis, occular inflammation, primary sclerosing cholanagitis)Early diagnosis (< 25 years old) was strongly associated with NOd1, Also associated with EIM (large and small joint arithritis, ankylosing spondylitis, occular inflammation, primary sclerosing cholanagitis)

29. Toll Receptors and Their Ligands Toll-like receptors (TLR) and their ligands. HSPs, heat shock proteins 60 and 70. TLR cannnot distinguish between commensal and pathogenicbacteria. Must balance between tolereance to the endogenous bacteria and host defense. This may be accomplished by the continu Toll-like receptors (TLR) and their ligands. HSPs, heat shock proteins 60 and 70. TLR cannnot distinguish between commensal and pathogenicbacteria. Must balance between tolereance to the endogenous bacteria and host defense. This may be accomplished by the continu

30. Myeloid differentiation factor 88 (MyD88)-dependent and MyD88-independent pathways are shown. Both pathways lead to the activation of NF- B and the mitogen-activated protein (MAP) kinase cascade and the production of IFN- . The Toll-IL-1 receptor domain-containing, adapter-inducing IFN- protein (TRIF)-related adapter molecule (TRAM)/TRIF adapter proteins also activate interferon regulatory factor (IRF)-3 and IRF-7 that leads to the production of IFN- . The cytokines that are directly induced by TLR2 and TLR4 signaling are shown in gray, and the chemokines that are induced by IFN- are shown in blue. TIRAP, Toll-IL-1 receptor domain-containing adapter protein; IKK, I B kinase; TBK1, TANK-binding kinase 1; STAT1, signal transducers and activators of transcription 1; iNOS, inducible nitric oxide synthase. TANK, TRAF-associated NF- B activator; IRAKs, IL-1 receptor-associated kinase; TRAF6, tumor necrosis factor (TNF) receptor-associated factor 6; Tak, transforming growth factor- (TGF- )-activated kinase; Tab, Tak-1-binding protein; IP-10, interferon inducible protein; MCP-5, monocyte chemoattractant protein; and IFNAR, interferon- receptor. Myeloid differentiation factor 88 (MyD88)-dependent and MyD88-independent pathways are shown. Both pathways lead to the activation of NF- B and the mitogen-activated protein (MAP) kinase cascade and the production of IFN- . The Toll-IL-1 receptor domain-containing, adapter-inducing IFN- protein (TRIF)-related adapter molecule (TRAM)/TRIF adapter proteins also activate interferon regulatory factor (IRF)-3 and IRF-7 that leads to the production of IFN- . The cytokines that are directly induced by TLR2 and TLR4 signaling are shown in gray, and the chemokines that are induced by IFN- are shown in blue. TIRAP, Toll-IL-1 receptor domain-containing adapter protein; IKK, I B kinase; TBK1, TANK-binding kinase 1; STAT1, signal transducers and activators of transcription 1; iNOS, inducible nitric oxide synthase. TANK, TRAF-associated NF- B activator; IRAKs, IL-1 receptor-associated kinase; TRAF6, tumor necrosis factor (TNF) receptor-associated factor 6; Tak, transforming growth factor- (TGF- )-activated kinase; Tab, Tak-1-binding protein; IP-10, interferon inducible protein; MCP-5, monocyte chemoattractant protein; and IFNAR, interferon- receptor.

31. C is for CLINICAL RELEVANCE

32. Potential Clinical Applications Of Genomics In IBD Patients 159. POTENTIAL CLINICAL APPLICATIONS OF GENOMICS IN IBD PATIENTS It is probable that in the future we will select therapy based on the genetic background of Crohn’s disease and ulcerative colitis patients. This concept is based on the premise that each genetic subset may predictably respond to individual treatment, which would provide a means to optimize in a cost effective manner the choice of medications to avoid toxicity and enhance clinical response. An additional impact of genomics is to identify subjects at high risk of developing disease. If the genotype of the proband with IBD is known, siblings and offspring could be screened for this gene. Patients found to be at high risk could receive prophylactic therapy which would be tailored by the nature of the genetic abnormality as discussed above. This approach offers the possibility of preventing the onset of these disorders. markers of inflammation such as C-reactive protein are useful to stratify patients likely to respond to biologic therapies and to follow response to treatment. In the future, functional genomics and proteomics will be used to rapidly screen patients for subclinical characteristics that can predict disease course and response to therapy 159. POTENTIAL CLINICAL APPLICATIONS OF GENOMICS IN IBD PATIENTS It is probable that in the future we will select therapy based on the genetic background of Crohn’s disease and ulcerative colitis patients. This concept is based on the premise that each genetic subset may predictably respond to individual treatment, which would provide a means to optimize in a cost effective manner the choice of medications to avoid toxicity and enhance clinical response. An additional impact of genomics is to identify subjects at high risk of developing disease. If the genotype of the proband with IBD is known, siblings and offspring could be screened for this gene. Patients found to be at high risk could receive prophylactic therapy which would be tailored by the nature of the genetic abnormality as discussed above. This approach offers the possibility of preventing the onset of these disorders. markers of inflammation such as C-reactive protein are useful to stratify patients likely to respond to biologic therapies and to follow response to treatment. In the future, functional genomics and proteomics will be used to rapidly screen patients for subclinical characteristics that can predict disease course and response to therapy

33. Hypothetical Regulation Of Clinical Phenotype And Therapeutic Response In IBD Patients By Genotype 158. HYPOTHETICAL REGULATION OF CLINICAL PHENOTYPE AND THERAPEUTIC RESPONSE IN IBD PATIENTS BY GENOTYPE It is likely that Crohn’s disease and ulcerative colitis are heterogeneous groups of disorders with common clinical features mediated by multiple genetic defects. It is probable, however, that each genetic subset may have a unique natural history and response to individual therapeutic agents. For example, genetic defect A may have an aggressive fistulizing course of disease and selectively respond to one drug while genetic defect B may have an entirely different natural history and therapeutic response. If this hypothesis is true, one could logically predict outcomes to different drugs and select a highly effective therapeutic regimen based on genotype. Early observations suggest that this hypothesis is true since Crohn’s disease patients with NOD2 (CARD 15) genetic abnormalities tend to have earlier onset, fibrostenotic disease localized to the small bowel compared to Crohn’s disease patients who have normal NOD2 genes.158. HYPOTHETICAL REGULATION OF CLINICAL PHENOTYPE AND THERAPEUTIC RESPONSE IN IBD PATIENTS BY GENOTYPE It is likely that Crohn’s disease and ulcerative colitis are heterogeneous groups of disorders with common clinical features mediated by multiple genetic defects. It is probable, however, that each genetic subset may have a unique natural history and response to individual therapeutic agents. For example, genetic defect A may have an aggressive fistulizing course of disease and selectively respond to one drug while genetic defect B may have an entirely different natural history and therapeutic response. If this hypothesis is true, one could logically predict outcomes to different drugs and select a highly effective therapeutic regimen based on genotype. Early observations suggest that this hypothesis is true since Crohn’s disease patients with NOD2 (CARD 15) genetic abnormalities tend to have earlier onset, fibrostenotic disease localized to the small bowel compared to Crohn’s disease patients who have normal NOD2 genes.

34. BALANCE OF COMMENSAL BACTERIAL COMPONENTS 132. BALANCE OF COMMENSAL BACTERIAL COMPONENTS Results in animal models of colitis as well as in some clinical studies suggest that a balance exists between proinflammatory and protective bacteria in the distal intestine. Injurious bacteria include Bacteroides vulgatus, which selectively induce disease in HLA B27 transgenic rats, Enterococcus faecalis, which induce colitis in IL-10 knockout mice, and enteroadherent/invasive E. coli, which are found in higher frequencies in the neoterminal ileum of Crohn’s disease patients undergoing relapse after segmental resection and primary anastomosis. Protective bacteria include the probiotic (beneficial bacteria) species Lactobacillus, Bifidobacterium and nonpathogenic E. coli which have been found to prevent and treat experimental colitis and prevent recurrence of pouchitis or ulcerative colitis. • Rath HC et al. Differential induction of colitis and gastritis in HLA ?27 transgenic rats selectively colonized with bacteroides vulgatus and Eschericia coli. Infection and Immunity 1999;67:2969. • Kim SC et al. IL-10 deficient mice monoassociated with non pathogenic Enterococcus faecalis develop chronic colitis. Gastroenterology 2001;120-A822 (abstract). • Darfeuille-Michaud A et al. Presence of adherent E. coli stains in ileal mucosa of patients with Crohn’s disease. Gastroenterology 1998;115:1405. • Gionchetti P et al. Oral Bacteriotherapy as maintenance treatment in patients with chronic pouchitis: a double-blind, placebo-controlled trial. Gastroenterology 2000;119:305-309. • Kruis W et al. Double-blind comparison of an oral E. coli preparation and mesalamine in maintaining remission of ulcerative colitis. Alimentary Pharmacology and Therapeutics 1997;11:853-8.132. BALANCE OF COMMENSAL BACTERIAL COMPONENTS Results in animal models of colitis as well as in some clinical studies suggest that a balance exists between proinflammatory and protective bacteria in the distal intestine. Injurious bacteria include Bacteroides vulgatus, which selectively induce disease in HLA B27 transgenic rats, Enterococcus faecalis, which induce colitis in IL-10 knockout mice, and enteroadherent/invasive E. coli, which are found in higher frequencies in the neoterminal ileum of Crohn’s disease patients undergoing relapse after segmental resection and primary anastomosis. Protective bacteria include the probiotic (beneficial bacteria) species Lactobacillus, Bifidobacterium and nonpathogenic E. coli which have been found to prevent and treat experimental colitis and prevent recurrence of pouchitis or ulcerative colitis. • Rath HC et al. Differential induction of colitis and gastritis in HLA ?27 transgenic rats selectively colonized with bacteroides vulgatus and Eschericia coli. Infection and Immunity 1999;67:2969. • Kim SC et al. IL-10 deficient mice monoassociated with non pathogenic Enterococcus faecalis develop chronic colitis. Gastroenterology 2001;120-A822 (abstract). • Darfeuille-Michaud A et al. Presence of adherent E. coli stains in ileal mucosa of patients with Crohn’s disease. Gastroenterology 1998;115:1405. • Gionchetti P et al. Oral Bacteriotherapy as maintenance treatment in patients with chronic pouchitis: a double-blind, placebo-controlled trial. Gastroenterology 2000;119:305-309. • Kruis W et al. Double-blind comparison of an oral E. coli preparation and mesalamine in maintaining remission of ulcerative colitis. Alimentary Pharmacology and Therapeutics 1997;11:853-8.

35. PROBIOTICS A preparation of, or a product containing, viable, defined microorganisms in sufficient numbers to alter the microflora by implantation or colonization in a compartment of the host, and exert beneficial effects on host health. Lactobacillus Lactobacillus rhamnosus GG (LGG) (ATCC 53103) is now widely used as a probiotic in dairy products.. Bifidobacterium Immunobiotics – therapeutic manipulation of microflora Oral LGG adheres and colonizes the human intestine Stimulate gut antigen presenting cells to promote protection and switch regulatory mechanisms The role of commensal anaerobic bacteria in host nutrition, colonic health and gut maturation is starting to be examined. The resident intestinal microflora is complex and involves more than 400 bacterial species. Establishment and maintenance of a microflora is highly dependable on food intake and style of diet. are bacterial strains with probiotic properties originating from human microflora.The role of commensal anaerobic bacteria in host nutrition, colonic health and gut maturation is starting to be examined. The resident intestinal microflora is complex and involves more than 400 bacterial species. Establishment and maintenance of a microflora is highly dependable on food intake and style of diet. are bacterial strains with probiotic properties originating from human microflora.

37. SUMMARY Autoimmune Bacteria Clinical relevance Probiotic is a food term s good against ddiarrhea die to due to rotavirus, C. Difficile, post antibiotic therapydisease with a genetic influence play an important role in the pathogenesis and also in the treatment of IBD targeting therapies toward improvement of mucosal barrier function Probiotic is a food term s good against ddiarrhea die to due to rotavirus, C. Difficile, post antibiotic therapydisease with a genetic influence play an important role in the pathogenesis and also in the treatment of IBD targeting therapies toward improvement of mucosal barrier function

38. Gene-Environment Interaction 124. PATHOGENESIS OF IBD Although genetic susceptibility, luminal antigenic drive, and environmental triggers are each important, animal models demonstrate that no single factor is sufficient to induce chronic relapsing, immune-mediated intestinal inflammation. Chronic inflammatory bowel diseases depend on the interaction of these essential components, each of which is necessary but not sufficient to induce disease.124. PATHOGENESIS OF IBD Although genetic susceptibility, luminal antigenic drive, and environmental triggers are each important, animal models demonstrate that no single factor is sufficient to induce chronic relapsing, immune-mediated intestinal inflammation. Chronic inflammatory bowel diseases depend on the interaction of these essential components, each of which is necessary but not sufficient to induce disease.

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