Verotoxin 1 binding to intestinal crypt epithelial cells results in localization to lysosomes and abrogation of toxicity
Verotoxin 1 binding to intestinal crypt epithelial cells results in localization to lysosomes and abrogation of toxicity
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DOI：
10.1046/j.1462-5822.2003.00254.x
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发表时间：
2003-02-01
影响因子：
3.4
通讯作者：
Smith, DGE
中科院分区：
文献类型：
--
作者：
Hoey, DEE;Sharp, L;Smith, DGE
肠出血性大肠杆菌（EHEC）是一组与人类严重疾病后遗症相关的细菌，而维罗毒素（VTs）是肠出血性大肠杆菌（EHEC）的重要毒力因子。VTs的强大细胞毒活性在致病性中是重要的，导致表达受体Gb3 （globotriaosylneuroide）的细胞死亡。肠出血性大肠杆菌，特别是血清型O157:H7，经常在没有疾病的情况下定植于水库宿主（如牛），然而，这种宿主无毒的基础尚不清楚。本研究的目的是评估VT与肠上皮之间的相互作用，肠上皮是宿主与肠道生物之间的主要界面。牛肠上皮细胞在体外原代细胞培养中表达Gb3，特异性定位于增殖的隐窝细胞，与肠黏膜原位免疫组织学观察结果一致。这些细胞中受体的表达与体内人肠上皮中Gb3的缺失形成对比。尽管受体表达，VT对牛上皮细胞没有细胞毒活性。VT的亚细胞定位表明该毒素被排除在内质网之外，而定位于溶酶体，与细胞毒性的消除相对应。用甲基- β -环糊精处理原代细胞培养物不影响VT细胞内运输，这表明这些细胞中的Gb3与脂筏无关，而是随机分布在膜中。Gb3异构体、膜分布和VT运输的结合与其他抗细胞毒性的受体阳性细胞的观察结果相关。这些研究表明，肠上皮是VT相互作用的重要决定因素，对宿主和人类肠出血性大肠杆菌感染的不同后果具有重要意义。
Verotoxins (VTs) are important virulence factors of enterohaemorrhagic Escherichia coli (EHEC), a group of bacteria associated with severe disease sequelae in humans. The potent cytotoxic activity of VTs is important in pathogenicity, resulting in the death of cells expressing receptor Gb3 (globotriaosylceramide). EHEC, particularly serotype O157:H7, frequently colonize reservoir hosts (such as cattle) in the absence of disease, however, the basis to avirulence in this host has been unclear. The objective of this study was assessment of interaction between VT and intestinal epithelium, which represents the major interface between the host and enteric organisms. Bovine intestinal epithelial cells expressed Gb3 in vitro in primary cell cultures, localizing specifically to proliferating crypt cells in corroboration with in situ immunohistological observations on intestinal mucosa. Expression of receptor by these cells contrasts with the absence of Gb3 on human intestinal epithelium in vivo . Despite receptor expression, VT exhibited no cytotoxic activity against bovine epithelial cells. Sub-cellular localization of VT indicated that this toxin was excluded from endoplasmic reticulum but localized to lysosomes, corresponding with abrogation of cytotoxicity. VT intracellular trafficking was unaffected by treatment of primary cell cultures with methyl-beta-cyclodextrin, indicating that Gb3 in these cells is not associated with lipid rafts but is randomly distributed in the membrane. The combination of Gb3 isoform, membrane distribution and VT trafficking correlate with observations of other receptor-positive cells that resist verocytotoxicity. These studies demonstrate that intestinal epithelium is an important determinant in VT interaction with major implications for the differential consequences of EHEC infection in reservoir hosts and humans.