EV71诱导人神经细胞SH-SY5Y自噬的分子机制

Mechanism of autophagy induced by EV71 in human neuroblastoma cell SH-SY5Y

摘要:

【背景】EV71感染所致的重症手足口病易导致神经系统并发症，使患儿预后较差，甚至死亡。【目的】从EV71可诱导神经细胞自噬这一现象出发，探索该病毒诱导神经细胞自噬的miRNA机制，探讨EV71损伤神经细胞可能的分子机制。【方法】通过RT-PCR及Western blot技术，在感染EV71病毒的人神经母细胞瘤细胞SH-SY5Y中检测细胞自噬变化；通过芯片分析细胞感染前后差异表达的miRNA分子，再使用miRNA mimics调节工具明确与EV71诱导神经细胞自噬有关的miRNA分子。【结果】EV71可诱导SH-SY5Y细胞自噬增加，下调细胞内miRNA29b (miR29b)分子的表达水平；当上调细胞内miR29b的表达后，EV71诱导细胞自噬增加的现象可被逆转，病毒复制水平下降。【结论】EV71诱导神经细胞自噬是通过下调miR29b分子的表达水平实现；miR29b不仅与自噬相关，它与EV71病毒复制也存在密切关系。因此，该研究不仅有助于阐明EV71导致神经系统损伤的具体分子机制，还为miR29b成为治疗EV71感染可能的新药物靶点奠定了理论基础。

Abstract:

[Background] Enterovirus 71 (EV71) is likely to cause severe hand, foot, and mouth disease (HFMD) that is sometimes associated with severe central nervous system complications, resulting in poor prognosis and even death of children. [Objective] To reveal the possible miRNA mechanism of EV71 increasing autophagy in human neuroblastoma cells, which might be related to EV71 induced neurological damage. [Methods] Western blot and qRT-PCR were used to demonstrate autophagy in EV71 infected SH-SY5Y cells. The differentially expressed miRNAs in SH-SY5Y cells with/without EV71 infection were detected by qPCR-Array. The functions of those miRNAs were analyzed by bioinformatics method. Then miRNA mimics was used to identify miRNAs associated with EV71-induced neuronal autophagy. [Results] EV71 could increase autophagy in SH-SY5Y cells and decrease the expression of miRNA29b in cells. Moreover, transfection of miRNA29b mimics in cells could not only reverse the down-regulated expression of miRNA29b induced by EV71, but also decrease the cellular autophagy induced by EV71 and the level of viral replication. [Conclusion] The results indicated that EV71 could induce autophagy in SH-SY5Y cells by down-regulating the expression of miRNA29b. Moreover, miRNA29b might be related to virus replication. Therefore, this study not only helps elucidate the specific molecular mechanism of neurological damage caused by EV71, but also lays a theoretical foundation for miRNA29b to become a new potential drug target for the treatment of EV71 infection.