体内体外实验研究肠道病毒EV71对TGF-β信号通路的调控及其分子机制

Enterovirus 71 (EV71) is one of most prevalent viral pathogens in China. As a major cause of hand, foot, and mouth disease (HFMD) in young children, infection of EV71 is also associated with severe neurological complications and even leads to deaths. Previous studies in clinics and a mouse model have suggested immune overreactions could be the reason for the severe host damage during the EV71 infection. Transforming growth factor-β (TGF-β) signaling pathway plays a pivotal role in regulating host immune response. Little has been known about the impact of EV71 infection on this pathway. By using the microarray analysis, our research for the first time identified that several factors in TGF-β signaling pathway have been significantly affected by EV71 infection. Cells infected with EV71 showed reduced levels of TGF-βR1, while the ‘inhibitory smads’ smad7 was up-regulated. In this project, the regulation mechanism of EV71 infection on TGF-β pathway will be further elucidated to identify which viral protein(s) play a determinal role in TGF-β pathway regulation. Furthermore, the regulation of TGF-β pathway and related impacts will be confirmed in EV71 infected mouse model. These results will help us understand the interaction between the EV71 infection and host immune responses and will provide a theoretical basis for the clinical treatments of the severe EV71 infection.

EV71在我国连续大范围流行，除引起手足口病外，还可以导致严重的神经系统并发症甚至死亡。临床和动物模型的研究认为免疫过激是EV71引起机体系统损伤的主要原因之一。TGF-β通路是机体免疫调节的重要路径。在前期研究中，我们证明EV71感染调控TGF-β通路的多个相关因子，包括下调受体TGF-βR1并且上调主要负性调控因子smad7。因此，我们推测EV71通过调控多个关键通路蛋白来抑制细胞内TGF-β通路，而这可能是导致免疫过激反应的重要原因。本项目中，我们将在细胞水平上研究哪一个或几个病毒蛋白调控TGF-β通路及相关分子，并阐明其分子机理。同时，我们还将在小鼠感染模型中验证EV71病毒与TGF-β通路之间的作用关系，并研究病毒调控TGF-β通路与病毒感染引起的组织损伤之间的相关性。研究结果将进一步揭示EV71感染与宿主免疫之间的相互作用，促进对病毒致病机制的了解，同时为临床治疗提供理论依据。

免疫过激是EV71病毒感染引起神经系统损伤的主要原因之一。转化生长因子β(TGF-β）信号通路是机体免疫调节的重要路径。本研究中，我们首先通过基因芯片、RT-PCR、免疫印迹等证明EV71感染过程中调控TGF-β信号转导通路的多个相关因子，包括下调TGF-βR1，上调主要负性调控因子Smad7。通过构建11个病毒表达载体，我们确认了EV71 的3AB病毒蛋白是调控TGF-βR1以及Smad7的主要病毒蛋白。通过构建Smad7启动子的报告载体，我们阐明了3AB蛋白通过调控Smad7启动子活性来增加Smad7的转录，进化提高感染细胞内Smad7蛋白的表达。同时，我们还利用小鼠感染模型验证了EV71病毒感染对TGF-β通路的抑制以及病毒感染引起炎症因子IL-6的表达以及组织损伤与TGF-β通路之间的相关性。本课题的研究从分子角度揭示EV71病毒感染调控宿主重要免疫通路TGF-β通路的分子机理，并通过构建病毒感染的乳鼠模型解析了EV71感染引起的组织损伤的潜在机理。此研究结果将促进我们对EV71病毒致病机制的了解，同时为未来的临床重症预警及治疗提供理论依据。