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Tuesday, April 29, 2014
Stefania Senger, PhD, Instructor in Pediatrics, Harvard Medical School, Mucosal Immunology and Biology Research Center, Massachusetts General Hospital for Children
The epithelial lining of the intestine is characterized by its extraordinary abilities both to continually turn over in a highly regulated fashion under homeostatic conditions and to rapidly renew itself after damage from infectious diarrheal diseases, radiation, and antineoplastic chemotherapy. It is not just a physical barrier to non-self antigens, but can actively adjust to challenging environments, contributing to innate immune response and shaping the adaptive one. Intestinal epithelial renewal is the product of a population of rapidly dividing intestinal stem-progenitor cells (ISCs), which give rise to all the intestinal epithelial cell lineages. Great interest surrounds intestinal stem cell modulation as it has become increasingly evident that stem cell related signaling is altered in numerous pathological settings including cancerous and non-cancerous human intestinal pathologies.
1. Impact of microbiota derived metabolites on gut mucosal molecular pathways and effect on early steps of celiac disease pathogenesis.
Celiac disease is an immune-mediated disorder affecting genetically predisposed subjects, caused by the ingestion of gluten. HLAII DQ2/DQ8 molecules are strictly required to develop celiac disease; however other and as yet unidentified genetic factors may be necessary for the onset of the disease. In celiac patients the small intestine shows blunting of villi and lengthening of crypts, that implies increase in the proliferative compartment over the mature differentiated epithelium. Alteration of stem cell related pathways are consistent with the observed histology.
Preliminary data generated in our laboratory together with published data support the hypothesis that the intestinal stem cell niche is indeed perturbed in these patients.
These alterations are unlikely to be due to genetic makeup alone, since the frequency of celiac disease has increased steadily in the past 40 years from 0.25% to 1% of the general population suggesting non-genetic factors in play.
Further, recent studies have shown the importance of microbiota produced metabolites in the genetic control of the host cell programs, providing evidence for a role for commensal bacteria in regulating intestinal stem cell differentiation.
We hypothesize that environmental triggers derived from commensal bacterial composition together with genetic makeup of the host can tilt the balance between tolerance and immune response in subjects genetically predisposed to celiac disease.
To test this hypothesis, I make use of an in vitro 3-D (organoid) primary intestinal culture system derived from intestinal stem cells. (Fig.1). The organoid represents the sole epithelial component of the small intestine. Using mouse organoids, I have tested the effect of metabolites that we previously identified to be altered in children with celiac disease, on intestinal stem cell differentiation and barrier function. Our preliminary data support the notion that metabolites can, in a dose dependant way, promote differentiation and contribute to the maintenance of a competent gut barrier function. We intend to extend the study to individuals with celiac disease and healthy control derived intestinal organoids to test the hypothesis that patient derived organoids respond differently to metabolites stimuli compared to healthy controls. This study could potentially lead to discovery of new genetic alterations that contribute to celiac disease and will increase our understanding on the early crosstalk between host mucosa and microbiota.
2. Identification of intestinal pattern recognition receptors (PRR)/pathways involved in epithelial modulation and surveillance against Shigella and Enteropathogenic Escherichia Coli (EPEC)
There is increasing evidence that interactions between host gastrointestinal mucosa and enteric pathogens are engaged by PRRs in different ways during the host-bacteria “cross-talk”. PRRs are expressed on several cell types and importantly on polarized cells, when engaged, can activate various downstream effectors based on their sub-cellular distribution (apical or basolateral). Of note, TLR4 has been found to be expressed on intestinal stem cells and contribute to stem cell apoptosis and differentiation. In this context my research focuses on investigating how exposure to pathogens and pathogen derived toxins, regulates early epithelial responses such as trans-epithelial and trans cellular permeability, epithelial production of pro-inflammatory cytokines, intercellular tight junctions (TJ) dynamics and late epithelial responses like regulation of intestinal stem cell functions. The contribution of PRR like PAR2 and TLRs to these responses will also be investigated. Gut derived organoid tissues and ex-vivo explants of wild type and KO mouse intestinal tracts, have been instrumental in my studies. The results of this research will contribute to understand how TLRs and/or PAR2 signaling are involved in intestinal mucosal surveillance against Shigella and EPEC and their effect on mucosal epithelium differentiation.
This study has been funded by NIDDK: Enteric Research Investigational Network Cooperative Research Centers (ERIN).
Figure 1: Intestinal organoid derived from a single, isolated murine crypt culture. The staining with a lysozyme antibody (left) shows mature, differentiated Paneth’s cells, properly localized at the bottom of the crypt-like domains. The same organoid is depicted (center) in phase light microscopy
Vanuytsel T, Senger S, Fasano A, Shea-Donohue T. Major signaling pathways in intestinal stem cells. Biochim Biophys Acta. 2013 Feb;1830(2):2410-26.
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