INTESTINAL TOXIN RECEPTORS AND PATHOGENESIS OF DIARRHEA

这项建议旨在确定 肠道对志贺菌毒素的反应。我们最近确认了 一种年龄相关的成熟模式，出现在 志贺氏菌的糖脂结合部位，球状三糖神经酰胺(Gb3) 毒素在兔小肠中的分布，并已获得初步数据 这表明毒素的分泌作用依赖于 Gb3的出现。这项建议的主要目标是 鉴定Gb3的发育规律并证实 Gb3是小肠中的功能性受体。这个 人类新生儿对临床志贺氏菌病的相对抵抗力可能 与受体的发育模式有关 小肠中的志贺氏菌毒素。这些实验将 包括兔小鼠血清中Gb3纯化和定量 肠道在不同的年龄，以建立年龄依赖性， Gb3中的数量差异是 肠道对志贺氏菌毒素的功能性反应。 糖脂将在DEAE-Sephadex上提取和纯化 色谱柱和Florisil或Unisil柱，并用 高效液相色谱柱。Gb3在近端和近端的定位和分布 小肠远端，位于隐窝、绒毛细胞和亚细胞内 分数，将在不同的年龄学习。这些实验将 描绘毒素受体的特定膜结构域并 将因此解释观察到的反应的变化 小肠不同程度的毒素。为了确认 Gb3是参与毒素介导的功能性受体 过程中，Gb3胶束将被插入耐火材料中 新生兔的肠道，以使它们对 毒素和Gb3脂质体的分泌作用 将使用抗Gb3的单抗来阻断毒素- 回肠结扎后的体液反应。监管机构 参与Gb3发育模式的机制将是 通过测量特定的半乳糖基转移酶活性进行评估 自初步数据显示Gb3水平下降以来，不同年龄的人 它的前体在新生儿中水平很高。荷尔蒙 对Gb3合成的调控也将进行研究。 所描述的研究将阐明发育控制 Gb3在不同日龄兔体内的作用。这些研究可能 为志贺氏菌毒素的发病机制提供新的见解 中介性腹泻，是发展中国家和发展中国家的一个主要健康问题 发达国家。

This proposal is designed to identify the developmental pattern of intestinal response to shigella toxin. We have recently identified an age-dependent, maturational pattern in the appearance of the glycolipid binding site, globotriaosylceramide (Gb3), for shigella toxin in rabbit small intestine, and have obtained preliminary data which suggest that the secretory effects of the toxin depend upon the appearance of Gb3. The major goals of this proposal are to characterize the developmental regulation of Gb3 and to confirm that Gb3 is the functional receptor in the small intestine. The relative resistence of human neonates to clinical shigellosis could be related to a developmental pattern of the receptors for shigella toxin in the small intestine. These experiments will include purification and quantitation of Gb3 from rabbit small intestine at various ages in order to establish age-dependent, quantitative difference in Gb3 as the basis for variations in functional responsiveness of the intestine to shigella toxin. Glycolipids will be extracted and purified on DEAE-Sephadex chromatography and Florisil or Unisil columns and quantitated by HPLC. The localization and distribution of Gb3 in proximal and distal small intestine, in crypt and villus cells and in subcellular fractions, will be studied at various ages. These experiments will delineate the specific membrane domain of the toxin receptor and will thus explain the observed variations in the response to the toxin at various levels of the small intestine. In order to confirm that Gb3 is the functional receptor involved in the toxin mediated process, Gb3 micelles will be inserted into the refractory intestine of newborn rabbits in order to render them sensitive to the secretory effects of the toxin, and Gb3 liposomes and monoclonal anti-Gb3 antibodies will be used to block the toxin- mediated fluid response in ligated ileal loops. The regulatory mechanisms involved in the developmental patter of Gb3 will be evaluated by measuring specific galactosyltransferase activity at various ages since preliminary data reveal decreased levels of Gb3 with high levels of its precursors in neonates. Hormonal modulation of the regulation of Gb3 synthesis will also be studied. The studies described will elucidate the developmental control and function of Gb3 in rabbits at various ages. These studies may provide new insights into the pathogenesis of shigella toxin mediated diarrhea, a major health problem in both developing and developed countries.