BACTERIAL TOXIN ACTION ON THE DEVELOPING HUMAN GUT

在全球范围内，腹泻病是婴儿和儿童发病和死亡的主要原因之一。我们从临床研究中了解到，早产儿肠道最初的细菌定植不当可能会导致严重的炎症，导致早产儿特有的肠道疾病，例如坏死性小肠结肠炎(NEC)，某些产毒性腹泻更常见，在新生儿期表现更严重。在人类肠道模型(细胞系、器官培养、使用小室和异种移植)的初步研究中，我们提供的数据表明，未成熟的人类肠道通过分泌过量的IL-8(一种中性粒细胞趋化因子)(内毒素)和过度的氯离子分泌(外毒素)来不适当地响应细菌毒素。基于这些观察，我们对这项研究的总体假设是，新生儿细菌性炎症性肠道疾病和某些涉及细菌毒素的分泌性腹泻的发病机制主要是由于未成熟(不适当的)肠细胞对细菌毒素刺激的反应。为了验证这一假设，我们将在人类肠道模型中使用两种毒素-肠道细胞“串扰”范式来表征未成熟肠道与成熟肠道中的上皮反应以及这种反应的机制。因此，我们的具体目标是：(1)通过检测内毒素-LBP-CD14与Toll样受体的相互作用和受体后信号转导事件(主要是导致NFkappaB激活的IL-1信号转导通路)，以IL-8分泌为效应反应来检测内毒素与胎儿肠道细胞的相互作用；(2)通过检测毒素结合和受体后反应，通过PGE2和5-羟色胺介导的cAMP、GSalpha、核糖化因子、效应表达和磷酸化等途径来研究外毒素与胎儿肠道细胞的相互作用。在研究了内毒素和外毒素与发育中的肠道相互作用的每一步后，我们将试图利用已知的成熟(营养)因子，将发育调节的步骤本身作为效应器反应来调节任何可识别的发育调节步骤。这些研究可能为使用一种特定的营养因子或营养因子组合预防早产儿和新生儿的毒素性腹泻提供依据。

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 (cells lines, organ culture, Ussing chambers, and xenotransplants), 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 (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 toll-like receptors (TLRs) and postreceptor signal transduction events (principally the IL-1 signal transduction pathway leading to NFkappaB activation) and (2) to study exotoxin-fetal enterocyte interaction using C1- secretion as the effector response by examining toxin binding and postreceptor 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 exotoxoin with the developing intestine, we will attempt to modulate any identifiable step that is developmentally regulated by using known maturational (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 toxigenic diarrhea in premature and neonatal infants.