Department of Experimental Medicine & Biotechnology, Post Graduate Institute of Medical Education & Research, Chandigarh 160012, India.
Microb Pathog. 2012 May;52(5):278-84. doi: 10.1016/j.micpath.2012.02.002.
Enteroaggregative Escherichia coli (EAEC) is an emerging enteric pathogen, associated with cases of acute and persistent diarrhoea worldwide. The pathogenesis of EAEC is yet to be understood. In intestinal epithelium, an increase in Ca²⁺ has been attributed due to the action of different enteric pathogens. EAEC was shown to increase Ca²⁺ in HEp-2 cells.The present study was undertaken to investigate the effect of EAEC induced increase in Ca²⁺ oncultured human intestinal epithelial cells.
INT-407 cells were infected with EAEC (T8 strain) in the absence and presence of dantrolene (inhibitor of release of Ca²⁺ from intracellular stores)/verapamil (L-type Ca²⁺ channel blocker)/BAPTAAM (Ca²⁺ chelator)/U73122 (PLC inhibitor)/Cytochalasin-D (inhibitor of actin polymerization). Ca²⁺ was estimated using Fura-2/AM. Cytoskeletal rearrangement was assessed by F-actin staining using TRITC-phalloidin. The invasiveness of EAEC-T8 to INT-407 cells was checked by electron microscopy and invasion assay.
A significant increase in Ca²⁺ was observed in EAEC-T8 infected INT-407 cells, which was reduced in presence of dantrolene/verapamil/U73122. EAEC-T8 could induce cytoskeletal F-actin polymerization in INT-407 cells and was found to be invasive in nature. The cytoskeletal rearrangement as well as invasion of EAEC-T8 was attenuated in presence of U73122/dantrolene/BAPTA-AM/verapamil/cytochalasin D.
EAEC induced increase in Ca²⁺ seems to play a major role in host cytoskeletal F-actin rearrangements leading to invasion of the organism.
Our study undoubtedly will lead to an improved understanding of EAEC-pathogenesis.
肠聚集性大肠杆菌（EAEC）是一种新兴的肠道病原体，与世界各地的急性和持续性腹泻病例有关。EAEC 的发病机制尚不清楚。在肠道上皮细胞中，由于不同的肠道病原体的作用，Ca²⁺已经被证明可以增加。EAEC 被证明可以增加 HEp-2 细胞中的Ca²⁺。本研究旨在研究 EAEC 诱导的Ca²⁺增加对培养的人肠道上皮细胞的影响。
INT-407 细胞在不存在和存在丹曲林（细胞内储存的 Ca²⁺释放抑制剂）/维拉帕米（L 型 Ca²⁺通道阻滞剂）/BAPTA-AM（Ca²⁺螯合剂）/U73122（PLC 抑制剂）/细胞松弛素-D（肌动蛋白聚合抑制剂）的情况下感染 EAEC（T8 株）。使用 Fura-2/AM 估计Ca²⁺。通过使用 TRITC-鬼笔环肽进行 F-肌动蛋白染色来评估细胞骨架重排。通过电子显微镜和侵袭试验检查 EAEC-T8 对 INT-407 细胞的侵袭性。
在 EAEC-T8 感染的 INT-407 细胞中观察到Ca²⁺显著增加，在存在丹曲林/维拉帕米/U73122 的情况下减少。EAEC-T8 可以诱导 INT-407 细胞中的细胞骨架 F-肌动蛋白聚合，并被发现具有侵袭性。在存在 U73122/丹曲林/BAPTA-AM/维拉帕米/细胞松弛素 D 的情况下，EAEC-T8 的细胞骨架重排和侵袭性减弱。
EAEC 诱导的Ca²⁺增加似乎在宿主细胞骨架 F-肌动蛋白重排导致机体侵袭中起主要作用。
我们的研究无疑将有助于更好地理解 EAEC 的发病机制。
