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Figures 1 and 2 depict the interaction of invasive microorganisms and/or their enterotoxins with enterocytes that result in epithelial/lymphoid or epithelial/nerve cell communications. Molecular, cellular, and in vivo approaches are used to study the mechanisms and the consequences of this interaction of microbes, enterotoxins, endotoxins, and probiotics with colonocytes as they relate to inflammation and maintenance of the tight junctional barrier. Newly established human fetal primary and conditionally immortalized cell lines and human xenograft approaches have been characterized and will be used to define the molecular mechanism(s) of a developmental epithelial response to bacterial exo- and endotoxin stimuli.
An emerging field of investigation is related to the participation of neuropeptides in the pathophysiology of intestinal inflammation caused by bacteria and bacterial products. Studies from the laboratory of Dr. Harry Pothoulakis, a collaborator from the Gastrointestinal Research Laboratory at Beth Israel Deaconess Hospital, demonstrated for the first time that the neuropeptide corticotropin-releasing hormone (CRH) and its receptors localized on intestinal epithelial cells play a major role in the development of intestinal inflammation mediated by the enterotoxins (toxin A and B) of Clostridium difficile. We also developed the fetal human intestinal xenograft system in SCID mice and used it successfully to study the effect of bacterial enterotoxins in the human intestinal mucosa. Recent evidence also indicates that the human xenograft model can be used to study the contribution of neuropeptides and their receptors in intestinal inflammatory responses in a "human" in vivo setting. Our laboratories use the xenograft model and the recently developed CRH receptor-deficient animals to study the importance of the "CRH system" in C. difficile toxin-mediated intestinal inflammation, and for the first time, in Shigella and Salmonella infection. The ability of probiotics such as Saccharomyces boulardii to inhibit expression of CRH receptors during an inflammatory response is a major focus of these studies. Dr. Pothoulakis has established a presence in this laboratory to study these issues in collaboration with other investigators.
Dr. Allan Walker’s laboratory has been one of the leading laboratories in studying the developmental changes of intestinal host defense in response to invading microorganisms and their toxins. Studies in animal models and, more recently, in human fetal intestinal models suggest that the immature human enterocyte under certain circumstances is not able to protect against bacteria, or responds inappropriately to microbes and enterotoxins. This "inappropriate" response may be central to the pathophysiology of age-related infectious gastrointestinal diseases. Recent exciting results indicate that the inability of the immature gut to discriminate between commensal and pathologic bacteria may be related, at least in part, to reduced expression of IkB by the immature enterocyte. Dr. Walker's laboratory focuses on identifying the mechanisms of developmental regulation of enterocyte responses to colonizing intestinal bacteria and their products and examines the role of Toll-like receptors in this response. These studies are undertaken in collaboration with Drs. Pothoulakis, McCormick, and Dr. Ciarán Kelly, another investigator from the GI laboratory at Beth Israel Deaconess Hospital.
Shigella organisms are responsible for acute bacillary dysentery, one of the most important health concerns worldwide. Dr. Beth McCormick’s laboratory has been studying the mechanisms involved in acute infectious colitis due to shigellosis. These studies utilize the T84 cell model in a detailed examination of Shigella and enterocyte basolateral membrane interface, including signal transduction and chemokine stimulus for neutrophil migration, which is the putative mechanism of Shigella-induced pathogenesis of gut inflammation. Collaborative studies with Dr. H.C. Reinecker have also suggested that S. flexneri regulates several functional components of the tight junctions, including ZO-1 and claudin-1. Dr. McCormick's group continues to examine in detail the molecular mechanisms involved in the modulation of tight junctional proteins by S. flexneri as well as the signaling pathways involved in S. flexneri-induced PMN transepithelial cell migration. Experiments to address the effects of probiotics in inflammatory enterocyte signaling and tight junctional regulation in response to S. flexneri are also performed. These studies are conducted in a collaborative manner with Drs. Walker, Reinecker, and Kelly. Dr. Reinecker is a collaborator on a Program Project grant whose laboratory exists in the GI Unit at MGH.
Dr. Ciarán Kelly has had a close working relationship with Dr. Pothoulakis, and he is well known to investigators within the Mucosal Immunology Laboratory at MGH-East. Dr. Kelly has an exciting new project in probiotics and intestinal inflammation that complements the overall objectives of the laboratory and promises considerable interaction among project leaders and their co-investigators. Dr. Kelly brings extensive experience in mechanisms of gastrointestinal inflammation in Helicobacter pylori gastritis and the immune response in Clostridium difficile infection. Dr. Kelly’s recent discovery is related to the ability of the non-pathogenic yeast Saccharomyces boulardii to release a potent anti-inflammatory factor (SAIF) that blocks proinflammatory gene expression from colonocytes by inhibiting NF-kB and MAP-kinase activation in these cells. His project is highly relevant to the focus of this laboratory on the mechanisms of microbes-mucosal barrier function interactions and the ensuing inflammatory response. He has active collaborations with Dr. Pothoulakis and has begun studies with Dr. McCormick. He has planned extensive collaborations with Drs. Nanthakumar and Walker to determine the effects of S. boulardii and its secreted factor SAIF on regulation of tight junction function and plans experiments with Drs. Pothoulakis and McCormick to examine the effect of this anti-inflammatory factor in C. difficile colitis and Shigella infection using the human intestinal xenograft model.
The inverse correlation between exposure to helminthes and the incidence of certain immune-mediated diseases, including inflammatory bowel diseases (IBD), has been evidenced from epidemiological studies. The distribution of several pathogenic helminth infections coincides geographically with many devastating microbial diseases, such as HIV, malaria, and tuberculosis. Infections with intestinal helminths and enteric bacterial pathogens, such as enteropathogenic Escherichia coli (EPEC), continue to be a major global health threat, especially for children. However, the nature of the interaction and the exact mechanism by which helminth modulates the host’s response to concurrent pathogens are still unclear. Dr. Hai Ning Shi’s laboratory investigates the role of helminth parasites in evoking a T helper cell intestinal response and its effect on bacterial invasion. He is particularly interested in exploring the mechanisms by which the helminth parasite modulates intestinal mucosal response to enteric bacteria and bacteria-associated intestinal inflammation using a co-infection model system. This system involves two murine enteric infectious agents that induce distinct Th responses: (i) the helminth Heligmosomoides polygyrus (Th2) and (ii) the Gram-negative bacterium Citrobacter rodentium (Th1). He has shown that helminth co-infection results in an impaired host protection and the development of more severe C. rodentium-mediated intestinal inflammation by a STAT 6 (Th2) dependent mechanism. He has also shown that the helminth modulates host response via the effect of dendritic cells. In collaboration with Dr. Walker, Dr. Shi also examines the impact of intestinal colonization of bacteria (probiotics) during early life on the development and regulation of mucosal T cell responses (Th1, Th2 and T regs) and explores the mechanisms by which probiotics modulate host protection against enteric pathogens. Using both in vivo and in vitro approaches, he also examines and defines the conditions under which dysregulation of intestinal mucosal response to luminal antigens triggers the development of intestinal inflammatory responses that ultimately result in chronic inflammatory disease. His research will provide greater insight about how intestinal microorganisms may alter the regulatory mechanisms of mucosal immunity, which may be instrumental in the establishment of effective preventive and therapeutic approaches for the treatment of Th1- and Th2-mediated diseases and for the design of effective intestinal vaccines.
Dr. Bobby Cherayil’s laboratory studies innate host defenses to gastrointestinal infection, using Salmonella typhimurium as a model intracellular bacterial pathogen. S. typhimurium is an important cause of acute gastroenteritis in humans as a result of its ability to provoke a vigorous inflammatory response in the intestine. Although self-limited in most individuals, this disease can be potentially life threatening in the very young, the elderly, and the immunocompromised, and is a significant cause of morbidity at all ages. Other types of Salmonella, such as S. typhi and S. paratyphi, cause a systemic febrile illness, typhoid fever, which is responsible for a large number of deaths, particularly in the developing world. In this disease, the organism lives and multiplies for extended periods within host macrophages at systemic sites such as the liver, spleen, and bone marrow. Both the intestinal and systemic phases of Salmonella infection can be studied conveniently in the mouse. One of the projects in the laboratory is directed at elucidating the molecular details of the intestinal inflammatory response to Salmonella. Dr. cherayil's group has shown recently that interferon gamma (IFN
) plays an important role in regulating the Salmonella-induced inflammatory response in the intestine. Along with Dr. Allan Walker’s group, they showed that this function is developmentally regulated. They are currently carrying out experiments to investigate the role of some of the IFN-regulated genes in intestinal inflammation, and together with Dr. Haining Shi’s laboratory, they are also attempting to identify the cellular source of IFN in the intestine. Dr. Pothoulakis’s group is assisting with some of these studies. The other important question being addressed relates to the mechanisms involved in the intracellular survival of Salmonella. Specifically, Dr. Cherayil is interested in understanding how the organism adapts to its intracellular niche, and conversely, how the host attempts to restrict the growth of the pathogen. His group has started to analyze the role of iron metabolism and autophagocytosis in these processes. In collaboration with Dr. Marianne Wessling-Resnick and her group at the Harvard School of Public Health, they have found recently that changes in expression or function of the macrophage iron efflux protein ferroportin have significant effects on intracellular growth of Salmonella, probably by changing the amount of iron available to the bacteria. Autophagocytosis also appears to play a role in the Salmonella-macrophage interaction, and the Cherayil lab is starting to explore this further with the help of the extensive mutant collection available in Dr. Beth McCormick’s laboratory. It is their hope that these investigations will shed light on important aspects of Salmonella pathogenesis, and that this information will be useful in devising new ways to treat and prevent the diseases caused by this organism.
The Program in Glycobiology, under the direction of Dr. David Newburg, studies biologically active glycans relevant to developmental mucosal immunology. For example, this program discovered that the human milk glycans constitute an innate immune system whereby the mother, through her milk, protects her infant from disease during infancy. Glycans include glycoproteins, glycolipids, glycopeptides, glycosaminoglycans, mucins, and oligosaccharides. They are expressed in especially high amounts on the cell surface, particularly the intestinal mucosal cell surface, and in human milk. Such glycans are important in cellular communication, but they are also used by many pathogens to identify, bind, and infect their host target cell. Human milk is especially rich in glycans, and many of the human milk glycans inhibit pathogens by inhibiting their ability to bind to their host cell receptors. This human milk research is part of a longstanding collaboration with Dr. Guillermo Ruiz-Palacios in Mexico City, and Dr. Ardythe Morrow in Cincinnati. The glycobiology program specializes in developing new methods by using state-of-the-art technology for the analysis of glycans, for measuring their metabolism, and for determining genetic control mechanisms of their expression. The program also measures biologically active glycans from exogenous sources, such as probiotics, in collaboration with Dr. Ciarán Kelly, and botanicals. This program also works closely with Dr. Nanda Nanthakumar in defining the ontogeny of glycans in the intestinal mucosa and their relationship to inflammatory bowel diseases.
Starting at birth, the gastrointestinal tract is colonized by microflora that interact with epithelial cells through constant reciprocal communication, termed microbial-epithelial crosstalk. The microflora is required for normal function of the intestine and, thus, for the well being of an individual. Dr. Nanda Nanthakumar's laboratory focuses on the microbial-epithelial crosstalk at birth, during weaning, and during the recovery from antibiotic therapy. His group has identified a mechanism of the crosstalk associated with the intestinal innate immune system. In collaboration with the immunologists Drs. Cherayil, Shi, and Walker, Dr. Nanthakumar investigates the role of bacterial-epithelial crosstalk in innate immunity. An important consequence of bacterial-epithelial crosstalk is changes in epithelial glycosylation. The ontogeny of intestinal glycosylation and its role in establishing the gut microbial ecosystem is being studied in collaboration with the glycobiologist Dr. David Newburg. They investigate genetic variation of human glycan expression as a determinant of variation in the composition of microbial ecosystems, and of altered susceptibility to enteric pathogens. Oral consumption of specific intestinal microbes exert health benefits by preventing or reducing the severity of enteric pathogen infection; these microbes are termed probiotics. The mechanism whereby probiotics alter crosstalk between microflora and intestinal mucosa, thought to underlie the ability of probiotics to reduce pathogenic infections, is being investigated in collaboration with the Walker and McCormick laboratories. Premature infants are born with an immature gut; rapid colonization can elicit inappropriate intestinal inflammation resulting in necrotizing enterocolitis (NEC) and other inflammatory bowel diseases. In collaboration with the Walker laboratory, Dr. Nanthakumar's group developed human models for studying the etiology and pathophysiology of NEC. With collaborations by clinical investigators from Chicago and Chile, a novel pathway was identified that may be important for the onset of NEC. Using specific cellular, molecular, genomic and proteomic approaches, the Nanthakumar lab is investigating various facets of bacteria-epithelial crosstalk between the host intestinal epithelium with mutualistic, pathogenic, and probiotic bacteria.
Dr. Verena Göbel’s laboratory investigates epithelial morphogenesis and growth regulation in the nematode Caenorhabditis elegans. One specific interest of the laboratory is in the development of the intestine. C. elegans is a well-characterized genetic model organism with its major internal organs formed by different types of tubes consisting of distinct, yet simple, polarized epithelia. The intestine of this transparent roundworm, a single-layered bilaterally symmetrical epithelium of only twenty cells, is particularly well suited to study tubulogenesis. The invariant pattern of C. elegans cell division is completely mapped, facilitating developmental studies. Its well established genetic resources and its short lifespan allow for the efficient performance of genetic screens, a powerful approach to identify physiologically relevant molecules. The pioneering role of invertebrates in delineating pathways relevant to human disease has been well documented and continues to be expanded.
Dr. Göbel’s laboratory has recently identified a number of cytoskeletal genes that are required to build the apical/luminal and basolateral membranes of the intestine and other tubular organs, and are needed to form and stabilize membrane microdomains such as microvilli. The characterization of the exact roles of these genes in the process of tubulogenesis is currently under investigation. A forward, genome-wide RNA interference (RNAi) screen on C. elegans tubulogenesis is also underway to identify additional morphogenesis and growth regulatory genes involved in this process. The emphasis of this screen lies on intestinal morphogenesis and is expected to identify the major developmental genes for the intestine. In collaboration with Dr. Allan Walker and Dr. Frank Ruemmele, Hopital Necker-Enfants Malades, Paris, identified mutant phenotypes and their underlying genetic defects will be used to focus a search for the underlying defective genes of inherited human intestinal failure syndromes.
Each of these investigators provides a detailed biosketch and research background and interests in an individual section of this website.