Interaction of Shiga toxin with primary human intestinal and renal epithelial cells: glycovesicles as novel toxin inhibitors

肠出血性大肠杆菌产生的志贺毒素（Stx）可导致危及生命的并发症，如溶血性尿毒症综合征（HUS），这被认为主要是由于Stx介导的肾和脑内皮细胞损伤。我们最近发现，除了内皮细胞外，人结肠和肾上皮细胞系也表达Stx糖鞘脂（GSL）受体globotriaosylceramide （Gb3Cer）和globotetraosylceramide (Gb4Cer)，并且对临床上高度相关的Stx1a和Stx2a亚型也很敏感。最新的结果表明，研究较少的人结肠和肾脏的原代上皮细胞也表现出Stx受体。该项目的目的是探索两种类型的原代上皮细胞(1)表达Gb3Cer-和gb4cer -脂质形式，以及与stx结合的GSLs是否与脂质筏相关，(2)stx - gsl -相互作用如何使用来自上皮细胞的类似脂质筏的膜实时进行，(3)Stx1a和Stx2a如何以及在多大程度上损害原代结肠和肾脏上皮细胞。(4)新开发的与Stx结合的新糖脂能否竞争性地抑制Stx介导的细胞损伤以及Stx与真正GSL受体的分子相互作用，这些新糖脂将作为糖囊插入囊泡中。该项目可以促进开发预防和治疗肠出血性大肠杆菌介导感染的新方法。

Shiga toxin (Stx) produced by EHEC causes life-threatening complications such as the hemolytic uremic syndrome (HUS), which are thought to be mainly attributed to Stx-mediated injury of endothelial cells of the kidney and the brain. We could recently show that besides endothelial cells human colonic and renal epithelial cell lines do express the Stx glycosphingolipid (GSL) receptors globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer) and are sensitive towards the clinically highly relevant Stx1a and Stx2a subtypes as well. Latest results indicate that still less investigated primary human epithelial cells of the colon and the kidney also exhibit Stx receptors. The aim of this project is to explore for both types of primary epithelial cells (1) which Gb3Cer- and Gb4Cer-lipoforms are expressed and whether the Stx-binding GSLs are associated with lipid rafts, (2) how the Stx-GSL-interaction proceeds in real time using lipid raft-analogous membranes derived from the epithelial cells, (3) how and to which extent Stx1a and Stx2a damage primary colon and kidney epithelial cells, and (4) whether the Stx-mediated cellular damage and the molecular interaction of Stx with the genuine GSL receptors can be competitively inhibited by using newly developed Stx-binding neoglycolipids, which will be applied inserted in vesicles as glycovesicles. The project could promote the development of novel options for prevention and therapy of EHEC-mediated infections.