Background. Enteroaggregative Escherichia coli (EAEC) is an evolving etiological agent of acute and persistent diarrhoea worldwide. The previous study from our laboratory has reported the apoptosis-inducing activity of EAEC in human small intestinal and colonic epithelial cell lines. In the present investigation, we have explored the underlying mechanism of EAEC-induced apoptosis in human intestinal epithelial cell lines. Methods. INT-407 and HCT-15 cells were infected with EAEC-T8 and EAEC-pT8 (plasmid cured strain of EAEC-T8) separately. Cells cultured in the absence of bacteria served as a negative control in all the experiments. For the subsequent experiments, the molecular mechanism(s) of epithelial cell aposptosis was measured in EAEC infecting both the cell lines by flow cytometry, real-time PCR and Western blotting. Results and conclusions. EAEC was found to activate the intrinsic/mitochondrial apoptotic pathway in both the cell lines through upregulation of pro-apoptotic Bax and Bak, un-alteration/reduction in the level of anti-apoptotic Bcl-2 and Bcl-XL, decrease in mitochondrial transmembrane potential, accumulation of cytosolic cytochrome c leading to activation of procaspase-9 and procaspase-3, which ultimately resulted in DNA fragmentation and apoptosis. Further, an increased expression of Fas, activation of procaspase-8 and upregulation of pro-apoptotic Bid in the EAEC-infected cells indicated the involvement of extrinsic apoptotic pathway too in this process. Our finding has undoubtedly led to an increased understanding of EAEC pathogenesis, which may be helpful to develop an improved strategy to combat the infection.

中文翻译：

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肠聚集性大肠杆菌诱导人肠上皮细胞凋亡的分子机制

背景。肠聚集性大肠杆菌(EAEC) 是全球范围内急性和持续性腹泻的一种不断发展的病原体。本实验室前期研究报道了EAEC对人小肠和结肠上皮细胞系的凋亡诱导活性。在本研究中，我们探索了 EAEC 诱导人肠上皮细胞系凋亡的潜在机制。方法。分别用EAEC-T8和EAEC-pT8(EAEC-T8的质粒治愈株)感染INT-407和HCT-15细胞。在没有细菌的情况下培养的细胞作为所有实验的阴性对照。在随后的实验中，通过流式细胞术、实时PCR和蛋白质印迹法测量了感染两种细胞系的EAEC中上皮细胞凋亡的分子机制。结果和结论。发现 EAEC 通过上调促凋亡 Bax 和 Bak、不变/降低抗凋亡 Bcl-2 和 Bcl-X L水平、减少线粒体凋亡，激活两种细胞系中的内在/线粒体凋亡途径。跨膜电位，胞浆细胞色素c的积累导致 procaspase-9 和 procaspase-3 的激活，最终导致 DNA 断裂和细胞凋亡。此外，EAEC 感染细胞中 Fas 表达增加、procaspase-8 激活和促凋亡 Bid 上调表明外源性凋亡途径也参与了这一过程。我们的发现无疑增加了对 EAEC 发病机制的了解，这可能有助于制定改进的对抗感染策略。