不同H1N1流感病毒感染A549细胞的生物学特征及对细胞基因表达的影响研究

发布时间：2018-11-21 20:24

【摘要】：2009甲型H1N1流感疫情在数月内遍及全球,流感大流行影响了社会经济和人群健康。虽然大多数病例的自限性病征类似于季节性流感,但仍有部分既往健康的年轻患者迅速进展为重症甚至死亡。甲型H1N1流感病毒致病机制的研究对于疾病防控有重要意义。本研究的目的是：本研究旨在比较甲型H1N1流感病毒与季节性H1N1流感病毒感染人呼吸道上皮细胞系的生物学特征；并利用表达谱芯片在基因层面分析2009甲型H1N1流感大流行的代表株A/California/07/2009(H1N1)与季节性流感病毒代表株A/广东宝安/51/2008(H1N1)可能存在的致病机理的差异。本课题的主要内容包括： 1.比较甲型H1N1流感病毒A/California/07/2009(H1N1)与季节性H1N1流感病毒A/广东宝安/51/2008(H1N1)以相同病毒量感染人呼吸道上皮细胞系A549细胞的生物学特征,包括感染率、病毒进入细胞的效率、感染后不同时间点的病毒复制、释放、细胞病变等。 2.利用博奥22K人类全基因组寡核苷酸微阵列芯片,得到A/California/07/2009(H1N1)与A/广东宝安/51/2008(H1N1)流感病毒感染A549细胞的基因表达谱,利用聚类分析和微阵列显著性分析筛选出每个时间点在两病毒感染的细胞中表达有显著性差异的基因,继而通过多种生物信息学公共数据库对这些差异表达基因的生物学功能注释,了解2009甲型H1N1流感病毒与季节性H1N1流感病毒对宿主细胞各种生物过程的影响和可能存在的致病机理的差异,解释甲型H1N1流感病毒感染机体所致的异于季节性流感的机体反应的原因。本课题的主要结果： 1.A549细胞对A/California/07/2009(H1N1)和A/广东宝安/51/2008(H1N1)流感病毒均易感,且两种病毒均可在A549细胞内有效复制。然而其感染过程的早期阶段存在差异。A/California/07/2009(H1N1)病毒感染A549细胞时,进入细胞的效率偏低,但依然能在细胞内高效复制,使得其感染率峰值及病毒释放与A/广东宝安/51/2008(H1N1)相当。 2.两种H1N1流感病毒感染A549细胞均表现出对宿主细胞的免疫抑制作用和抗凋亡作用,然而2009甲型H1N1流感病毒表现出的免疫抑制和抗凋亡作用弱于季节性流感病毒。多数差异表达基因变化趋势一致,不同点并非质的差别,而在于量的差异。仅有少数基因的变化趋势在两种病毒感染的细胞中不同。 3.甲型H1N1流感病毒可以通过多种机制抑制宿主细胞的抗病毒反应,包括通过抑制细胞因子的产生、抑制天然免疫细胞(如NK)的活化、逃避巨噬细胞的识别,达到对宿主天然免疫反应的逃避。
[Abstract]:The 2009 H1N1 pandemic has affected social, economic and human health around the world for several months. Although the self-limited symptoms of most cases are similar to seasonal influenza, there are still some previously healthy young patients who rapidly develop severe or even die. It is important to study the pathogenic mechanism of influenza A H1N1 virus for disease prevention and control. The purpose of this study was to compare the biological characteristics of human respiratory epithelial cell lines infected with influenza A H1N1 virus and seasonal H1N1 influenza virus. The possible pathogenicity of A/California/07/2009 (H1N1) and A / Guangdong Baoan / 51 / 2008 (H1N1), the representative strain of 2009 H1N1 influenza pandemic, was analyzed by using the expression microarray at the gene level. The main contents of this topic are as follows: 1. To compare the biological characteristics of human respiratory epithelial cell line A549 infected with A H1N1 influenza virus (H1N1) and seasonal H1N1 influenza virus A / Guangdong Baoan / 51 / 2008 (H1N1) in the same amount, including the infection rate. The efficiency of virus entry into cells, virus replication, release, cytopathia and so on at different time points after infection. 2. The gene expression profiles of A549 cells infected with A/California/07/2009 (H1N1) and A / Guangdong Baoan / 51 / 2008 (H1N1) influenza virus were obtained by using the Boo 22K human genome oligonucleotide microarray chip. Cluster analysis and microarray significance analysis were used to screen the genes with significant difference at each time point in the cells infected by the two viruses. And then annotate the biological functions of these differentially expressed genes through a variety of bioinformatics public databases, To understand the effects of 2009 H1N1 influenza virus and seasonal H1N1 influenza virus on various biological processes of host cells and the possible pathogenetic mechanisms. Explain why the influenza A H1N1 virus infects the body, which is different from seasonal influenza. The main results of this study were as follows: 1.A549 cells were susceptible to A/California/07/2009 (H1N1) and A / Guangdong Baoan / 51 / 2008 (H1N1) influenza viruses, and the two viruses could replicate effectively in A549 cells. However, there are differences in the early stages of the infection. When A/California/07/2009 (H1N1) virus infects A549 cells, it is inefficient to enter the cells, but can still replicate efficiently in the cells. The peak infection rate and virus release are comparable to A / Guangdong Baoan / 51 / 2008 (H1N1). 2. Both H1N1 influenza viruses infected A549 cells showed immunosuppressive and anti-apoptotic effects on host cells, but the immunosuppressive and anti-apoptotic effects of 2009 H1N1 influenza virus were weaker than those of seasonal influenza viruses. The variation trend of most differentially expressed genes is the same, the difference is not the qualitative difference, but the quantitative difference. Only a few genes have a different trend in cells infected by both viruses. 3. Influenza A (H1N1) virus can inhibit the antiviral response of host cells through a variety of mechanisms, including inhibiting cytokines production, inhibiting the activation of innate immune cells (such as NK), and evading macrophage recognition. To escape the innate immune response of the host.
【学位授予单位】：中国疾病预防控制中心
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R373

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