In Project 1, a new research project, we will study the interactions of bacterial endo- exotoxin with the developing gut. Diarrheal disease constitutes one of the major causes of morbidity and mortality in infants and children on a global scale. We know from clinical studies that an inappropriate initial bacterial colonization of the premature intestine may result in severe inflammation leading to an intestinal disease unique to the premature, e.g.-necrotizing enterocolitis (NEC) and that certain toxigenic diarrheas occur more commonly and are manifested more severely in the neonatal period. In preliminary studies in human intestinal models (cell lines, organ culture, Ussing chambers, and neonatal period. In preliminary studies in human intestinal models (cell lines, organ culture, Ussing, and xenograph transplants), we provide data that the immature human intestine inappropriately responds to bacterial toxin by secreting excessive IL-8, a chemokine for neutrophils, (endotoxin) and by excessive chloride secretion (endotoxin and by excessive chloride secretion (exotoxin). Based on these observations, our overall hypothesis for this research proposal is that the pathogenesis of neonatal bacterial inflammatory intestinal diseases and certain secretory diarrheas involving bacterial toxins is principally due to an immature (inappropriate) enterocyte response to the bacterial toxin stimulation. In order to test this hypothesis, we will use two toxin-enterocyte """"""""crosstalk"""""""" paradigms in human intestinal models to characterize the epithelial response and the mechanisms of this response in the immature compared to the mature intestine. Accordingly, our specific aim are: (1) to examine endotoxin interaction with the fetal enterocyte using IL-8 secretion as the effector response by examining LPS-LBP-CD14 interaction with the fetal enterocyte using IL-8 secretion as the effector response by examining LPS-LBP-CD14 interaction with tool-like receptors (TLRs) and post- receptor signal transduction events (principally the IL-1 signal transduction pathway leading to NfkappaB activation) and (2) to study exotoxin-fetal enterocyte interaction using Cl-secretion as the effector response by examining toxin binding and post-receptor responses via cAMP, Gsalpha, ribosylation factors, effector expression and phosphorylation and other pathways mediated by PGE2 and 5-HT. Having examined each step in the interaction of endo- and exotoxin with the developing intestine we will attempt we will attempt to modulate any identifiable that is developmentally regulated by using known mutational (trophic) factors using the developmentally regulated step itself as the effector response. These studies may provide the basis for using a specific trophic factor or combination of trophic factors in the prevention of NEC and toxigenic diarrhea in premature and neonatal infants.
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