Interaction Between Resident Luminal Bacteria and the Host: Can a Healthy Relationship Turn Sour?

INTRODUCTION The mucosal surfaces and cavities of the gastrointestinal (GI) tract in humans and animals are populated by a complex mixture of non-pathogenic microorganisms. These enteric microorganisms, often referred to as the indigenous or normal microflora, are composed of more than 400 species having spatial differences in population size and relative species predominance along the GI tract. The host has evolved various mechanisms of tolerance to these organisms which allows a peaceful and productive coexistence with resident bacterial flora while allowing it to remain responsive to enteropathogenic bacterial species. This discriminatory ability of the intestine toward its luminal microflora is a pivotal feature of efficient tolerance and homeostatic mechanisms. However, under specific conditions, including genetic susceptibility and certain environmental factors, the host may mount an inappropriate response to its own resident luminal bacteria with devastating consequences if not promptly restrained. Indeed, an emerging hypothesis proposes that human inflammatory bowel disease (IBD) is caused by a loss of tolerance toward the host's indigenous luminal content. Thus, the concept that only pathogenic bacteria initiate inflammatory host responses in the intestine should be reconsidered. Although an extensive literature details the molecular mechanisms of host responses to pathogenic bacteria and the ensuing inflammatory disorders, little is known about the intestinal mucosal responses to the non-pathogenic intestinal microflora. This review covers the state of knowledge of the host responses to the indigenous commensal microflora of the intestine, especially the responses of the intestinal epithelial cells (IEC). It also provides an overview of the potential mechanisms involved in the control of intestinal inflammation and the maintenance of homeostasis with special emphasis on the implications for IBD. PHYSIOLOGICAL AND PATHOPHYSIOLOGICAL IMPACT OF HOST/MICROBIAL INTERACTION IN THE INTESTINE Physiological Impact of Host/Microbial Interaction Savage and Dubos defined the term indigenous microflora as including both microorganisms naturally colonizing a particular habitat (autochthonous flora) and microorganisms ubiquitously present in a given environment (allochthonous flora) but unable to colonize the host except under abnormal circumstances (1–4). The normal intestinal microflora does not establish itself all at the same time and is not uniform throughout the GI tract. At various times after the host's birth, certain microorganisms colonize particular regions of the GI tract with luminal habitats that suit their specific needs. The tremendous impact of the endogenous luminal flora on the development and maturation of gut homeostasis is eloquently illustrated by comparative studies of germ-free and conventionalized animals. Studies in gnotobiotic animals have shown that experimentally colonizing the intestine of germ-free rodents with single species selected from the indigenous flora has profound impacts on the anatomical, physiological, and immunological development of the host, including effects on epithelial cell functions and the composition of the gut-associated lymphoid tissue (GALT) (5–7). It is clear that the immunological responses of conventional animals differ greatly from those of germ-free animals. The number of intraepithelial lymphocytes (IEL) is reduced in germ-free animals, particularly αβ T-cell receptor (α, βTCR)-bearing T cells, which also have reduced cytolytic activity. Lamina propria lymphocytes (LPL) are reduced in number and in reactivity to mitogens (8–10). Lymphoid aggregates such as Payer's patches are small and poorly developed in the intestine of germ-free animals compared with those in the intestine of animals harboring a normal indigenous microflora (11). The importance of the indigenous microflora in establishing mucosal lymphocyte populations has been shown in mice with severe-combined immunodeficiency (SCID) who have been reconstituted with mature thymus-derived T-cells (12). It has been found that the number and function of lymphocyte populations is greatly reduced in T-cell reconstituted SCID mice with a reduced microbial flora consisting of only two non-pathogenic clostridial species. This finding suggests that homing and/or expansion of thymus-derived T cell populations in the intestine may be best driven by a complex mixture of luminal bacterial antigens. MacDonald and Carter have shown that normal bacterial flora must be present in order for mice to mount a delayed-hypersensitivity (DTH) reaction, a finding suggesting that enteric bacteria influence peripheral T-cell function as well as local intestinal T-cell function (13). Pathological Impact of Host/Microbial Interaction. A characteristic feature of the mucosal immune system in the normal host is that that the protective cell-mediated and humoral immune responses against enteropathogenic organisms are allowed to proceed while responses to microorganisms of the indigenous flora are prevented. This complex homeostasis between acquisition of tolerance (unresponsiveness) to the indigenous microflora and protective immune responses to enteropathogens presents an intriguing immunological paradox. Under conditions of chronic intestinal inflammation such as IBD, this homeostasis appears to be disrupted (14). Ulcerative colitis (UC) and Crohn disease (CD), two discrete pathologies of IBD, are spontaneously relapsing, immunologically-mediated disorders of the GI tract (15). Microbial agents of the normal microflora are invoked in many of the current etiologic theories of these disorders (16). Previous studies have shown a greater association of luminal enteric bacteria with the intestinal epithelium in patients with IBD than in normal controls (17). Duchmann et al. have demonstrated that immunologic tolerance to the endogenous microflora exists in normal controls but does not exist in patients with active IBD (18,19). These findings are consistent with the clinical observation that there is a lack of inflammation in bypassed distal ileal or colonic segments in IBD patients after proximal diversion of the fecal stream, but a rapid reactivation of immune responsiveness and inflammation when the proximal effluent is introduced into the bypassed ileum (20–22). In addition, a mechanistic role for the enteric non-pathogenic bacterial flora in the pathogenesis of chronic mucosal inflammation has been shown in various animal models of experimental colitis (23,24). Selective intestinal colonization by defined bacterial species in germ-free rodents with experimental colitis triggers disease, although not all enteric bacteria are equal in their capacity to induce chronic mucosal inflammation. For example, reconstitution studies of gnotobiotic HLA-B27 transgenic rats (25,26) and studies in guinea pigs with carrageenen induced colites (27) implicate Bacteroides vulgatus as particularly important to the induction of colitis but have not found any pathological response after colonization with Escherichia coli. Conversely B. vulgatus appears to protect against the development of E. coli-induced experimental colitis in IL-2−/− mice (28), suggesting that different bacterial species have differing effects in initiating and perpetuating intestinal inflammation in different genetically susceptible hosts. The absence of experimental colitis and pathological immune responses to any of the bacterial species in wild-type animals demonstrates the non-pathogenic nature of normal microflora and, most importantly, suggests that the normal host develops immunosuppressive mechanisms to control the constant challenge to the immune system by antigens from commensal microorganisms. Protective Effects of Host/Microbial Interaction. It appears that bacteria of the endogenous microflora not only affect the induction of immune maturation and defense mechanisms of the naïve gut but also induce protective mechanisms in the host with considerable therapeutic relevance for experimental colitis and IBD. For example, Madsen et al. have shown that specific-pathogen free (SPF) IL-10−/− mice colonized at 2 weeks of age display changes in bacterial colonization with simultaneous increases in aerobic adherent and translocated bacteria and reduced levels of Lactobacillus reuteri. Rectal administration of endogenous L. reuteri enhances mucosal barrier function and attenuates the development of colitis in SPF IL-10−/− mice at 4 weeks of age (29). Similar protective effects of lactobacilli have been demonstrated in a rat model of methotrexate-induced enterocolitis. These rats have decreased weight loss, decreased intestinal permeability, and decreased myeloperoxidase levels after oral administration of L. plantarum compared to untreated controls (30). Evidence of the protective activity of certain lactic acid bacteria in human IBD is provided by studies from Gionchetti et al. The authors show that a combination of 8 different lactic acid bacteria, including lactobacilli, bifidobacteria, and streptococci (VSL#3), inhibits relapse of chronic pouchitis (15% recurrence rate in the VSL#3 treated group vs. 100% in the placebo group), inhibits mucosal TNFα and up-regulates IL-10 production in treated pouches (31–33). Non-pathogenic gram-negative E. coli have also been shown to have protective effects against inflammation in patients with ulcerative colitis (34). The mechanisms ensuring hyporesponsiveness to indigenous microflora in normal hosts or the mechanisms by which this microflora protects the host during chronic intestinal inflammation and the role of the intestinal epithelium in regulating mucosal immune responses are virtually unknown. Reaching an understanding of the interplay between host and the luminal endogenous bacteria and the molecular mechanisms controlling intestinal tolerance promises to improve our ability to treat intestinal inflammation. The inflammatory process observed in the intestine is the consequence of the dysregulated activation of mucosal cells such as IECs, mesenchymal cells, macrophages, neutrophils, dendritic cells, and lymphocytes. Although the individual contribution of these cell types to the inflammatory process are different, IECs are the likely primary target of the proinflammatory constituents in the intestinal lumen. BACTERIAL CROSS-TALK TO IECs The host is isolated and protected from the potentially toxic intestinal luminal content by a single layer of IECs. These cells are the first line of defense against noxious luminal agents and, as has been shown recently, are important players in intestinal responses induced by microbial products (35,36). There is accumulating evidence that the collaboration between enteric bacteria and the intestinal epithelium helps create a complex ecosystem by modifying host epithelial cell biology. For example, Bacteroides thetaiotoomicron a prominent species of the normal anaerobic, gram-negative microflora of the distal intestinal tract provides nutrients and binding receptors for other bacterial species by inducing the production of specific fucosylated glycoconjugates by the host (37). Furthermore, the colonization of germ-free mice with B. thetaiotoomicron induces a characteristic profile of genes in epithelial cells, which enhances mucosal barrier function, nutrient absorption, xenobiotic metabolism, differentiation, defense, and angiogenesis (38–40). Remarkably, the single bacterial strain B. thataiotoomicron and, to a lesser extent, E. coli or Bacteroides infantis can recapitulate changes in the phenotype and function of the gut epithelium normally induced by the complete mixed conventional microflora. These findings support the concept that there may be bacteria-specific effects in the crosstalk between organisms and host epithelium. Adherent and invasive pathogenic bacteria, bacterial cell wall components, and cytokines all can stimulate IECs to release proinflammatory products including chemokines, cytokines, and adhesion molecules (35,36). The a role in the induction of many of the cytokines, chemokines, and adhesion molecules involved in inflammatory disorders, including IBD The production and of these proinflammatory molecules in the of the have a on the inflammatory to the activation of propria cells dendritic cells, and and to the of peripheral and cells which in the inflammatory process the potential of bacteria and bacterial products are the most important given their content in the intestinal lumen. Although the and only of luminal and the ileum only the number of microorganisms in the to of colonic with the intestinal epithelium and host epithelial cells are in to luminal pathogenic and non-pathogenic microorganisms. 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