Persistent Transmissible Gastroenteritis Virus Infection Enhances Enterotoxigenic Escherichia coli K88 Adhesion by Promoting Epithelial-Mesenchymal Transition in Intestinal Epithelial Cells
Persistent Transmissible Gastroenteritis Virus Infection Enhances Enterotoxigenic Escherichia coli K88 Adhesion by Promoting Epithelial-Mesenchymal Transition in Intestinal Epithelial Cells
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持续传染性胃肠炎病毒感染通过促进肠上皮细胞中的上皮-间质转化来增强产肠毒素大肠杆菌 K88 粘附
DOI：
10.1128/jvi.01256-17
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发表时间：
2017-11-01
影响因子：
5.4
通讯作者：
Yang, Qian
中科院分区：
文献类型：
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作者：
Xia, Lu;Dai, Lei;Yang, Qian
传染性胃肠炎病毒（TGEV）是一种以腹泻为特征的冠状病毒，发病率高，2周龄以下仔猪死亡率为100%。感染TGEV的猪通常会被其他病原体继发感染，从而加重腹泻的严重程度，但其机制尚不清楚。在这里，我们假设持续的TGEV感染刺激上皮-间质转化（EMT），因此产肠毒素大肠杆菌（ETEC）更容易粘附在生成细胞上。肠上皮细胞是TGEV和ETEC感染的主要靶点。我们发现TGEV可以持续感染猪肠柱状上皮细胞（IPEC-J2）并引起EMT，这与关键细胞特征的多重变化一致。感染细胞呈现成纤维细胞样形状；间充质标记物水平增加，上皮标记物相应减少；白细胞介素-1β (IL-1β)、IL-6、IL-8、转化生长因子β (TGF-β)、肿瘤坏死因子α (TNF-α) mrna表达水平升高；迁徙和入侵行为的增加。进一步的实验表明，通过TGF-β激活磷脂酰肌醇3-激酶(PI3K)/Akt和细胞外信号调节激酶（ERK）信号通路对于tgev介导的EMT过程至关重要。在经历EMT的细胞中，细胞摄取也发生了改变。tgev感染的细胞具有更高水平的整合素α5和纤维连接蛋白，并表现出增强的ETEC K88粘附。逆转EMT可降低ETEC K88粘附，抑制整合素α5和纤维连接蛋白的表达。总之，这些结果表明，TGEV感染诱导IPEC-J2细胞的EMT，增加ETEC K88在肠道的粘附，促进双重感染。传染性胃肠炎病毒（TGEV）引起猪腹泻，并经常继发其他病原体感染。在这项研究中，我们发现持续的TGEV感染诱导猪肠柱状上皮细胞（IPEC-J2）发生EMT，并增强继发性病原体ETEC K88的粘附。进一步的实验表明，整合素α5和纤维连接蛋白在tgev增强的ETEC K88粘附中起重要作用。逆转EMT可降低整合素α5和纤维连接蛋白的表达，降低ETEC K88的粘附。我们得出结论，TGEV感染触发EMT并促进双重感染。我们的研究结果为继发性感染提供了新的见解，并表明靶向抗emt治疗可能对继发性感染的预防和治疗具有重要意义。
ABSTRACT Transmissible gastroenteritis virus (TGEV) is a coronavirus characterized by diarrhea and high morbidity rates, and the mortality rate is 100% in piglets less than 2 weeks old. Pigs infected with TGEV often suffer secondary infection by other pathogens, which aggravates the severity of diarrhea, but the mechanisms remain unknown. Here, we hypothesized that persistent TGEV infection stimulates the epithelial-mesenchymal transition (EMT), and thus enterotoxigenic Escherichia coli (ETEC) can more easily adhere to generating cells. Intestinal epithelial cells are the primary targets of TGEV and ETEC infections. We found that TGEV can persistently infect porcine intestinal columnar epithelial cells (IPEC-J2) and cause EMT, consistent with multiple changes in key cell characteristics. Infected cells display fibroblast-like shapes; exhibit increases in levels of mesenchymal markers with a corresponding loss of epithelial markers; have enhanced expression levels of interleukin-1β (IL-1β), IL-6, IL-8, transforming growth factor β (TGF-β), and tumor necrosis factor alpha (TNF-α) mRNAs; and demonstrate increases in migratory and invasive behaviors. Additional experiments showed that the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) signaling pathways via TGF-β is critical for the TGEV-mediated EMT process. Cellular uptake is also modified in cells that have undergone EMT. TGEV-infected cells have higher levels of integrin α5 and fibronectin and exhibit enhanced ETEC K88 adhesion. Reversal of EMT reduces ETEC K88 adhesion and inhibits the expression of integrin α5 and fibronectin. Overall, these results suggest that TGEV infection induces EMT in IPEC-J2 cells, increasing the adhesion of ETEC K88 in the intestine and facilitating dual infection. IMPORTANCE Transmissible gastroenteritis virus (TGEV) causes pig diarrhea and is often followed by secondary infection by other pathogens. In this study, we showed that persistent TGEV infection induces an EMT in porcine intestinal columnar epithelial cells (IPEC-J2) and enhances the adhesion of the secondary pathogen ETEC K88. Additional experiments suggest that integrin α5 and fibronectin play an important role in TGEV-enhanced ETEC K88 adhesion. Reversal of EMT reduces the expression of integrin α5 and fibronectin and also reduces ETEC K88 adhesion. We conclude that TGEV infection triggers EMT and facilitates dual infection. Our results provide new insights into secondary infection and suggest that targeted anti-EMT therapy may have implications for the prevention and treatment of secondary infection.