Genetics and Environmental Interactions Shape the Intestinal Microbiome to Promote Inflammatory Bowel Disease Versus Mucosal Homeostasis

Abstract

See “A pyrosequencing study in twins shows that gastrointestinal microbial profiles vary with inflammatory bowel disease phenotypes,” by Willing BP, Dicksved J, Halfvarson J, et al, on page 1844. See “A pyrosequencing study in twins shows that gastrointestinal microbial profiles vary with inflammatory bowel disease phenotypes,” by Willing BP, Dicksved J, Halfvarson J, et al, on page 1844. Recent genome-wide genetic association studies and advances in molecular analysis of microbial composition have improved dramatically our understanding of the pathogenesis of chronic idiopathic inflammatory bowel diseases (IBD), particularly Crohn's disease. Crohn's disease seems to be the result of an overly aggressive immune response to a subset of commensal enteric bacteria in a genetically susceptible host, with disease initiated by environmental triggers.1Sartor R.B. Microbial influences in inflammatory bowel diseases.Gastroenterology. 2008; 134: 577-594Abstract Full Text Full Text PDF Scopus (1396) Google Scholar, 2Xavier R.J. Podolsky D.K. Unravelling the pathogenesis of inflammatory bowel disease.Nature. 2007; 448: 427-434Google Scholar, 3Cadwell K. Patel K.K. Maloney N.S. et al.Virus-plus-susceptibility gene interaction determines Crohn's disease gene Atg16L1 phenotypes in intestine.Cell. 2010; 141: 1135-1145Google Scholar, 4Garrett W.S. Gordon J.I. Glimcher L.H. Homeostasis and inflammation in the intestine.Cell. 2010; 140: 859-870Google Scholar, 5Abraham C. Cho J.H. Inflammatory bowel disease.N Engl J Med. 2009; 361: 2066-2078Google Scholar Polymorphisms in ≥75 genes have been associated with Crohn's disease.5Abraham C. Cho J.H. Inflammatory bowel disease.N Engl J Med. 2009; 361: 2066-2078Google Scholar Functionally, these genetic risks can be characterized broadly as abnormalities in innate immune responses, immunoregulation, or mucosal barrier function. Innate immune abnormalities primarily result in defective bacterial killing, either owing to altered production and secretion of antimicrobial peptides by Paneth cells (as described for NOD 2, ATG16L1, XBP-1)6Wehkamp J. Salzman N.H. Porter E. et al.Reduced Paneth cell alpha-defensins in ileal Crohn's disease.Proc Natl Acad Sci U S A. 2005; 102: 18129-18134Google Scholar, 7Kaser A. Lee A.H. Franke A. et al.XBP1 links ER stress to intestinal inflammation and confers genetic risk for human inflammatory bowel disease.Cell. 2008; 134: 743-756Google Scholar, 8Cadwell K. Liu J.Y. Brown S.L. et al.A key role for autophagy and the autophagy gene Atg16l1 in mouse and human intestinal Paneth cells.Nature. 2008; 456: 259-263Google Scholar; defective clearance of intracellular bacteria by cells transfected with the Crohn's NOD2 truncation mutant9Hisamatsu T. Suzuki M. Reinecker H.C. et al.CARD15/NOD2 functions as an anti-bacterial factor in human intestinal epithelial cells.Gastroenterology. 2003; 124: 993-1000Google Scholar and in preliminary studies by NOD2-deficient macrophages10Liu B. Schmitz J.M. Holt L. et al.NOD2 contributes to intracellular bacterial killing by phagocytes.in: 2009: 136Google Scholar; and enhanced intracellular replication of a Crohn's ileitis-derived adherent/invasive Escherichia coli strain in ATG16L1- and IGRM- deficient cells.11Lapaquette P. Glasser A.L. Huett A. et al.Crohn's disease-associated adherent-invasive E. coli are selectively favoured by impaired autophagy to replicate intracellularly.Cell Microbiol. 2010; 12: 99-113Google Scholar These genetically determined abnormalities in bacterial killing by innate immune cells link genetic polymorphisms with abnormal bacterial composition (dysbiosis) that has been described in a subset of patients with Crohn's disease.12Frank D.N. St Amand A.L. Feldman R.A. et al.Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases.Proc Natl Acad Sci U S A. 2007; 104: 13780-13785Google Scholar Very recently, Frank et al13Frank D.N. Robertson C.E. Hamm C.M. et al.Disease phenotype and genotype are associated with shifts in intestinal-associated microbiota in inflammatory bowel diseases.Inflamm Bowel Dis. 2010 Sep 13; ([Epub ahead of print])Google Scholar documented associations of NOD 2 and ATG16L1 genotypes with dysbiosis in Crohn's disease patients. In murine models, deficiencies of NOD2 or the multidrug resistance gene (Mdr1, which encodes P- glycoprotein) have been documented to alter intestinal microbiota.14Nones K. Knoch B. Dommels Y.E. et al.Multidrug resistance gene deficient (mdr1a-/-) mice have an altered caecal microbiota that precedes the onset of intestinal inflammation.J Appl Microbiol. 2009; 107: 557-566Google Scholar, 15Gulati A.S. Kruek L. Sartor R.B. Influence of NOD 2 on the protective intestinal commensal bacterium Faecalibacterium prausnitzii.2010Google Scholar As documented by numerous studies, dysbiosis in Crohn's disease is characterized by dramatic increases in mucosally associated bacteria,16Swidsinski A. Ladhoff A. Pernthaler A. et al.Mucosal flora in inflammatory bowel disease.Gastroenterology. 2002; 122: 44-54Abstract Full Text Full Text PDF Scopus (1091) Google Scholar increased Enterobacteriaceae,12Frank D.N. St Amand A.L. Feldman R.A. et al.Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases.Proc Natl Acad Sci U S A. 2007; 104: 13780-13785Google Scholar including adherent/invasive E coli17Darfeuille-Michaud A. Boudeau J. Bulois P. et al.High prevalence of adherent-invasive Escherichia coli associated with ileal mucosa in Crohn's disease.Gastroenterology. 2004; 127: 412-421Abstract Full Text Full Text PDF Scopus (1099) Google Scholar, 18Baumgart M. Dogan B. Rishniw M. et al.Culture independent analysis of ileal mucosa reveals a selective increase in invasive Escherichia coli of novel phylogeny relative to depletion of Clostridiales in Crohn's disease involving the ileum.The ISME Journal. 2007; 1: 403-418Google Scholar and decreased clostridial groups IV and XIVa, including protective Faecalibacterium prausnitzii.12Frank D.N. St Amand A.L. Feldman R.A. et al.Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases.Proc Natl Acad Sci U S A. 2007; 104: 13780-13785Google Scholar, 19Sokol H. Pigneur B. Watterlot L. et al.Faecalibacterium prausnitzii is an antiinflammatory commensal bacterium identified by gut microbiota analysis of Crohn's disease patients.Proc Natl Acad Sci U S A. 2008; 105: 16731-16736Google Scholar A major unresolved question is whether this dysbiosis represents primary versus secondary changes. Increased numbers of mucosally adherent bacteria in patients with infectious enterocolitis and differences with disease activity in IBD patients,16Swidsinski A. Ladhoff A. Pernthaler A. et al.Mucosal flora in inflammatory bowel disease.Gastroenterology. 2002; 122: 44-54Abstract Full Text Full Text PDF Scopus (1091) Google Scholar reversal of abnormalities with steroid therapy,20Swidsinski A. Loening-Baucke V. Vaneechoutte M. et al.Active Crohn's disease and ulcerative colitis can be specifically diagnosed and monitored based on the biostructure of the fecal flora.Inflamm Bowel Dis. 2008; 14: 147-161Google Scholar and similar alterations of commensal microbiota with nonspecific intestinal injury or infection in mice21Lupp C. Robertson M.L. Wickham M.E. et al.Host-mediated inflammation disrupts the intestinal microbiota and promotes the overgrowth of Enterobacteriaceae.Cell Host Microbe. 2007; 2: 119-129Google Scholar suggest secondary changes. However, dysbiosis associated with genetic polymorphisms,13Frank D.N. Robertson C.E. Hamm C.M. et al.Disease phenotype and genotype are associated with shifts in intestinal-associated microbiota in inflammatory bowel diseases.Inflamm Bowel Dis. 2010 Sep 13; ([Epub ahead of print])Google Scholar evidence of disease transmission with commensal bacterial species recovered from diseased mice22Garrett W.S. Gallini C.A. Yatsunenko T. et al.Enterobacteriaceae act in concert with the gut microbiota to induce spontaneous and maternally transmitted colitis.Cell Host Microbe. 2010; 8: 292-300Google Scholar and immune-mediated colitis in genetically susceptible mice mono-associated with single commensal bacterial species (E coli or Enterococcus faecalis)23Kim S.C. Tonkonogy S.L. Albright C.A. et al.Variable phenotypes of enterocolitis in interleukin 10-deficient mice monoassociated with two different commensal bacteria.Gastroenterology. 2005; 128: 891-906Google Scholar suggest primary, causal effects of dysbiosis on intestinal inflammation. Studies in twins have provided powerful insights into the pathogenesis of IBD and the origin of intestinal microbiota composition. The concordance rate of Crohn's disease in monozygotic twins is 44–55% and only 8–10% in ulcerative colitis,24Halfvarson J. Bodin L. Tysk C. et al.Inflammatory bowel disease in a Swedish twin cohort: a long-term follow-up of concordance and clinical characteristics.Gastroenterology. 2003; 124: 1767-1773Google Scholar suggesting key environmental influences in addition to the obvious genetic factors most notable in Crohn's disease. Turnbaugh et al25Turnbaugh P.J. Hamady M. Yatsunenko T. et al.A core gut microbiome in obese and lean twins.Nature. 2009; 457: 480-484Google Scholar reported no significant differences in microbial diversity between stool samples of monozygotic and dizygotic twin pairs and their mother, although there was a trend toward greater similarity in monozygotic compared with dizygotic twins. In this issue of Gastroenterology, Willing et al26Willing B.P. Dicksved J. Halfvarson J. et al.A pyrosequencing study in twins shows that gastrointestinal microbial profiles vary with inflammatory bowel disease phenotypes.Gastroenterology. 2010; 139: 1844-1854Google Scholar examined 40 twin pairs divergent or concordant for Crohn's disease or ulcerative colitis by pyrosequencing of polymerase chain reaction-expanded 16s ribosomal RNA means. They determined that disease phenotype outweighed genotype effects on the fecal and mucosally associated microbiome. Unfortunately, this study was not able to conclusively compare the diversity of microbiota in monozygotic versus dizygotic twins with IBD owing to small numbers of samples (11 pairs of dizygotic twins, only 6 of whom had Crohn's disease), although no obvious changes were noted. Consistent with previous reports, Crohn's disease patients exhibited decreased microbial diversity and ileal mucosal samples had increased Enterobacteriaceae, including E coli and Ruminococcus gnavus concentrations. Of potential pathophysiologic importance, Ruminococcus gnavus has mucolytic activities.27Png C.W. Linden S.K. Gilshenan K.S. et al.Mucolytic bacteria With increased prevalence in IBD mucosa augment in vitro utilization of mucin by other bacteria.Am J Gastroenterol. 2010 Jul 20; ([Epub ahead of print])Google Scholar Two genera of core commensals were significantly decreased in ileal Crohn's disease: The Ruminococcaceae family, including Faecalibacterium, and Roseburia, which is a member of the Firmicutes phylum. These genera that were decreased in ileal Crohn's disease are prominent producers of short-chain fatty acids, most notably butyrate, which has important protective activities in the intestine. Particularly novel features of this study include the large number of twin pairs; the observation that dysbiosis was most marked in ileal Crohn's disease, where the bacterial composition significantly differed from that of colonic Crohn's disease, ulcerative colitis, and normals; and comparison of microbiota samples in several locations in the same patient, showing close similarities of dysbiosis in ileal mucosal and fecal samples. Unfortunately, such twin studies cannot distinguish genetic from early environmental influences that persist long after twins separate and live in different environments. Genetic and environmental stimuli not only contribute to the pathogenesis of Crohn's disease, but also strongly influence composition of the intestinal microbiota28Hansen J. Gulati A. Sartor R.B. The role of mucosal immunity and host genetics in defining intestinal commensal bacteria.Curr Opin Gastroenterol. 2010 Sep 24; ([Epub ahead of print])Google Scholar (Figure 1). In addition to the effects of individual genes mentioned, host factors strongly determine bacterial composition, as demonstrated by cross colonization of microbiota in different species.29Rawls J.F. Mahowald M.A. Ley R.E. et al.Reciprocal gut microbiota transplants from zebrafish and mice to germ-free recipients reveal host habitat selection.Cell. 2006; 127: 423-433Google Scholar Well-documented environmental contributors to shaping of the gut bacterial composition include transfer of maternal microbiota, diet, and antibiotics. Transfer of Lactobacilli and Bifidobacteria by breast feeding and vaginal delivery provide initial colonization of the infant.30Martin R. Heilig G.H. Zoetendal E.G. et al.Diversity of the Lactobacillus group in breast milk and vagina of healthy women and potential role in the colonization of the infant gut.J Appl Microbiol. 2007; 103: 2638-2644Google Scholar, 31Martin R. Jimenez E. Heilig H. et al.Isolation of bifidobacteria from breast milk and assessment of the bifidobacterial population by PCR-denaturing gradient gel electrophoresis and quantitative real-time PCR.Appl Environ Microbiol. 2009; 75: 965-969Google Scholar Maternal influences are elegantly demonstrated by uterine implantation studies of 2 different mouse strains into the same foster mother that showed no differences in fecal microbiota profiles in the offspring with different genetic background.32Friswell M.K. Gika H. Stratford I.J. et al.Site and strain-specific variation in gut microbiota profiles and metabolism in experimental mice.PLos One. 2010; 5: e8584Google Scholar Relocation studies showed that environmental changes before 4 weeks of age affected fecal microbiota, although bacterial profiles were stable in adult mice. These studies indicate that although host genotype helps to shape intestinal microbial composition, early life environmental influences alter the final microbiota profile. Intestinal bacteria depend on dietary and host-derived nutrients for survival; therefore, it is not surprising that bacterial composition and gene expression are profoundly influenced by diet.33Turnbaugh P.J. Ley R.E. Mahowald M.A. et al.An obesity-associated gut microbiome with increased capacity for energy harvest.Nature. 2006; 444: 1027-1031Google Scholar, 34Turnbaugh P.J. Ridaura V.K. Faith J.J. et al.The effect of diet on the human gut microbiome: a metagenomic analysis in humanized gnotobiotic mice.Sci Transl Med. 2009; 1: 6ra14Google Scholar, 35Sonnenburg J.L. Xu J. Leip D.D. et al.Glycan foraging in vivo by an intestine-adapted bacterial symbiont.Science. 2005; 307: 1955-1959Google Scholar Dietary factors potentially relevant to IBD pathogenesis and treatment include iron, which enhances experimental colitis and increases concentrations of iron-dependent E coli, Klebsiella, and Bacteroides species in mice,36Matsuura M. Liu B. Carroll I. et al.Selective alteration of growth and virulence for iron-dependent bacterial species; possible novel mechanisms of detrimental effects of dietary iron on intestinal inflammation.2010Google Scholar sucrose and fructose) which potentiate experimental colitis and alters colonic microbiota; Whitehead and Sartor, unpublished observations), and undigested fiber and prebiotics, such as inulin and fructose oligosaccharides, which enhance growth of Bifidobacterium and are substrates for butyrate production.37Guarner F. Prebiotics, probiotics and helminths: the 'natural' solution?.Dig Dis. 2009; 27: 412-417Google Scholar Recent data demonstrate that use of antibiotics that are relevant to Crohn's disease, such as repeated cycles of ciprofloxacin, a combination of ciprofloxacin and metronidazole, or the combination of amoxicillin, metronidazole, and bismuth results in long-term changes in microbial composition and decreased diversity.38Dethlefsen L. Relman D.A. Microbes and health sackler colloquium: incomplete recovery and individualized responses of the human distal gut microbiota to repeated antibiotic perturbation.Proc Natl Acad Sci U S A. 2010 Sep 16; ([Epub ahead of print])Google Scholar, 39Antonopoulos D.A. Huse S.M. Morrison H.G. et al.Reproducible community dynamics of the gastrointestinal microbiota following antibiotic perturbation.Infect Immun. 2009; 77: 2367-2375Google Scholar, 40Swidsinski A. Loening-Baucke V. Bengmark S. et al.Bacterial biofilm suppression with antibiotics for ulcerative and indeterminate colitis: consequences of aggressive treatment.Arch Med Res. 2008; 39: 198-204Google Scholar These alterations could potentially enhance susceptibility to IBD in genetically predisposed individuals. Future studies in larger twin cohorts and carefully designed family units undergoing genetic profiling can further differentiate the relative influences of genetic and environmental factors in IBD. Parallel animal model studies can perform mechanistic studies of the influence of IBD-related genes on complex microbial populations and use gnotobiotic mice colonized with selected groups of IBD-relevant bacterial species to identify specific effects of genes on bacterial composition, gene expression, and function. Likewise, the effects of various Western dietary components on composition, gene expression, and function of commensal bacteria of particular importance to IBD can be defined in specific pathogen–free and gnotobiotic studies. These studies will lay the foundation for therapeutic approaches to both treat and prevent onset and relapses of intestinal inflammation in genetically susceptible hosts using physiologic, nontoxic manipulations to correct the dysbiosis of IBD. A Pyrosequencing Study in Twins Shows That Gastrointestinal Microbial Profiles Vary With Inflammatory Bowel Disease PhenotypesGastroenterologyVol. 139Issue 6PreviewThe composition of the gastrointestinal microbiota is thought to have an important role in the etiology of inflammatory bowel diseases (IBDs) such as Crohn's disease (CD) and ulcerative colitis (UC). Interindividual variation and an inability to detect less abundant bacteria have made it difficult to correlate specific bacteria with disease. Full-Text PDF