Co-culture of epithelial cells and bacteria for investigating host-pathogen interactions
Co-culture of epithelial cells and bacteria for investigating host-pathogen interactions
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DOI：
10.1039/b911367c
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发表时间：
2010-01-01
期刊：
影响因子：
6.1
通讯作者：
Jayaraman, Arul
中科院分区：
文献类型：
--
作者：
Kim, Jeongyun;Hegde, Manjunath;Jayaraman, Arul
人类胃肠道是肠道上皮细胞和非致病性细菌共存的独特环境。这种平衡被病原体进入胃肠道所扰乱。感染过程中的一个关键步骤是病原体通过尿道细菌层附着在上皮细胞上。有人提出，病原体在皮层中遇到的微环境在决定附着和定殖的程度方面起着重要作用。目前用于研究病原体定殖的培养方法不太适合于研究这种假设，因为它们不能以模拟胃肠道微环境的方式共培养细菌和上皮细胞。在这里，我们报告了一个微流体共培养模型的发展，使真核细胞和细菌的独立培养，并测试病原体定植的微环境的影响。开发了一种电致动的系统以形成可逆的岛，其允许细菌生物膜沿着上皮细胞单层的培养物的发展。共培养模型用于在HeLa细胞之间形成一个可降解的大肠杆菌生物膜，然后引入肠出血性大肠杆菌。大肠杆菌（EHEC）进入肠道，其顺序模拟胃肠道感染的事件顺序。利用野生型E.大肠杆菌和tnaA突变体（缺乏吲哚信号）作为肠道细菌，我们证明肠道生物膜微环境是EHEC感染性和毒力的关键决定因素。我们的模型有可能被用于基础研究调查胃肠道信号对肠出血性大肠杆菌毒力的影响，以及用于筛选不同的益生菌菌株，以调节病原体在胃肠道的感染性。
The human gastrointestinal (GI) tract is a unique environment in which intestinal epithelial cells and non-pathogenic (commensal) bacteria co-exist. This equilibrium is perturbed by the entry of pathogens into the GI tract. A key step in the infection process is the navigation of the pathogen through the commensal bacterial layer to attach to epithelial cells. It has been proposed that the microenvironment that the pathogen encounters in the commensal layer plays a significant role in determining the extent of attachment and colonization. Current culture methods for investigating pathogen colonization are not well suited for investigating this hypothesis as they do not enable co-culture of bacteria and epithelial cells in a manner that mimics the GI tract microenvironment. Here we report the development of a microfluidic co-culture model that enables independent culture of eukaryotic cells and bacteria, and testing the effect of the commensal microenvironment on pathogen colonization. A pneumatically-actuated system was developed to form reversible islands that allow development of bacterial biofilm along with culture of an epithelial cell monolayer. The co-culture model used to develop a commensal Escherichia coli biofilm among HeLa cells, followed by introduction of enterohemorrhagic E. coli (EHEC) into the commensal island, in a sequence that mimics the sequence of events in GI tract infection. Using wild-type E. coli and a tnaA mutant (lacks the signal indole) as the commensal bacteria, we demonstrate that the commensal biofilm microenvironment is a key determinant of EHEC infectivity and virulence. Our model has the potential to be used in fundamental studies investigating the effect of GI tract signals on EHEC virulence as well as for screening of different probiotic strains for modulating pathogen infectivity in the GI tract.