Enterohemorrhagic Escherichia coli reduce mucus and intermicrovillar bridges in human stem cell-derived colonoids.
Enterohemorrhagic Escherichia coli reduce mucus and intermicrovillar bridges in human stem cell-derived colonoids.
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DOI：
10.1016/j.jcmgh.2015.10.001
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发表时间：
2016-01-01
影响因子：
7.2
通讯作者：
Kovbasnjuk O
中科院分区：
文献类型：
--
作者：
In J;Foulke-Abel J;Zachos NC;Hansen AM;Kaper JB;Bernstein HD;Halushka M;Blutt S;Estes MK;Donowitz M;Kovbasnjuk O
肠出血性大肠杆菌（EHEC）在美国每年引起超过70，000次食源性腹泻。由于该疾病的初始无症状阶段和缺乏合适的动物模型，在危及生命的出血性结肠炎和溶血性尿毒综合征之前的早期事件顺序尚未完全了解。我们确定了EHEC与人结肠上皮细胞相互作用的初始分子事件。将来自成人近端结肠干细胞的人类集落发育成单层以研究EHEC-上皮相互作用。通过跨上皮电阻测量监测单层融合和分化。单层细胞顶部感染肠出血性大肠杆菌，并使用生化和成像方法评估上皮损伤随时间的进展。人类结肠样培养物概括了隐窝和表面结肠细胞的差异蛋白表达模式。产生粘液的分化的结肠样单层优先被肠出血性大肠杆菌定植。EHEC定植后，在结肠样单层的顶端表面形成特征性的附着和消退病变。肠出血性大肠杆菌在感染的早期阶段以粘蛋白2（结肠粘液的主要成分）和原钙粘蛋白24（PCDH 24）（微绒毛驻留蛋白）为目标。EHEC分泌的丝氨酸蛋白酶EspP通过PCDH 24还原启动刷状缘损伤。人结肠样单层是一个相关的病理生理模型，允许肠道感染期间的早期分子事件的研究。集落样单层提供了进入顶端和基底侧表面的途径，从而提供了优于三维培养的优势，以可控和易处理的方式研究宿主-病原体相互作用。在没有肠道细菌的情况下，EHEC减少结肠粘液并影响刷状缘细胞骨架。
Enterohemorrhagic Escherichia coli (EHEC) causes over 70,000 episodes of foodborne diarrhea annually in the United States. The early sequence of events that precede life-threatening hemorrhagic colitis and hemolytic uremic syndrome is not fully understood due to the initial asymptomatic phase of the disease and the lack of a suitable animal model. We determined the initial molecular events in the interaction between EHEC and human colonic epithelium. Human colonoids derived from adult proximal colonic stem cells were developed into monolayers to study EHEC-epithelial interactions. Monolayer confluency and differentiation were monitored by transepithelial electrical resistance measurements. The monolayers were apically infected with EHEC, and the progression of epithelial damage over time was assessed using biochemical and imaging approaches. Human colonoid cultures recapitulate the differential protein expression patterns characteristic of the crypt and surface colonocytes. Mucus-producing differentiated colonoid monolayers are preferentially colonized by EHEC. Upon colonization, EHEC forms characteristic attaching and effacing lesions on the apical surface of colonoid monolayers. Mucin 2, a main component of colonic mucus, and protocadherin 24 (PCDH24), a microvillar resident protein, are targeted by EHEC at early stages of infection. The EHEC-secreted serine protease EspP initiates brush border damage through PCDH24 reduction. Human colonoid monolayers are a relevant pathophysiologic model that allow the study of early molecular events during enteric infections. Colonoid monolayers provide access to both apical and basolateral surfaces, thus providing an advantage over three-dimensional cultures to study host–pathogen interactions in a controllable and tractable manner. EHEC reduces colonic mucus and affects the brush border cytoskeleton in the absence of commensal bacteria.