论文部分内容阅读
目的用1日龄ICR小鼠传代制备EV71小鼠适应株,研究EV71亲代株与小鼠适应株的体内外感染特点,建立EV71感染ICR小鼠动物模型,为病毒疫苗和抗病毒药物的研究提供实用的动物评价工具。方法用1日龄ICR小鼠进行EV71病毒(Fuyang-0805)的传代,得到小鼠传代株。以一定浓度亲代株和传代株病毒分别接种RD、Vero、SY5Y、Caco-2四种细胞,定量方法检测各时间点不同毒株在四种细胞上的复制数量,CCK8方法测定各时间点细胞的存活率;同时,两毒株分别腹腔注射感染1日龄小鼠,定期安乐死动物,采集肺、小肠、骨骼肌、大脑四种器官组织,进行动物体内病毒半定量和定量分析,同时进行各器官组织病理学观察、免疫组织化学鉴定。结果与亲代毒株相比较,小鼠传代株(EV71-MMP4)表现出更强的肌肉来源细胞嗜性与毒性;同时,两毒株腹腔注射感染1日龄小鼠后,EV71-MMP4感染的小鼠体重增长较正常小鼠体重增长缓慢;半定量和定量RT-PCR显示,在小鼠肌肉中的病毒载量于感染后1d和5d达到高峰。EV71-MMP4感染组感染率较高、病毒组织分布较广、感染持续性较好、病毒载量较高,高剂量病毒感染后小鼠小肠、心肌和骨骼肌可观察到细胞空泡变性、淋巴细胞浸润等病理变化。免疫组织化学显示感染后小鼠骨骼肌有EV71病毒特异分布。结论阜阳EV71小鼠适应株表现出较亲代毒株更好的小鼠易感性、细胞毒性,所建立的动物模型可用于EV71病毒致病机制、感染特点的研究和病毒疫苗及药物的评价。
OBJECTIVE: To establish EV71 mouse adaptive strain by passage on day 1 of ICR mice, to study the characteristics of in vitro and in vivo infection of EV71 parental strain and mouse adaptive strain, and to establish animal model of EV71 infection in ICR mice and to provide evidence for virus vaccine and antiviral drug research Practical animal evaluation tool. Methods The passaged passage of EV71 virus (Fuyang-0805) was performed on 1-day-old ICR mice to obtain the mouse passaged strains. The cells of RD, Vero, SY5Y and Caco-2 were inoculated with a certain concentration of parental strain and subcultured virus respectively. The quantity of replication of different strains on the four kinds of cells at different time points was detected by quantitative method. Survival rate; the same time, the two strains were injected intraperitoneally infected 1 day old mice, regular euthanasia animals, lung, small intestine, skeletal muscle and brain collected four organ tissues for animal virus semi-quantitative and quantitative analysis of the same time, each organ Histopathological observation, immunohistochemical identification. Results EV71-MMP4 showed more muscle-derived cell tropism and toxicity compared with the parent strain. At the same time, EV71-MMP4-infected mice infected with EV71-MMP4 The body weight of mice increased slowly compared with that of normal mice. Semiquantitative and quantitative RT-PCR showed that the viral load in mouse muscle peaked at 1 day and 5 days after infection. The EV71-MMP4 infection group had a higher infection rate, a wider distribution of virus tissues, better infection persistence and higher viral load. Degeneration of cells was observed in the small intestine, myocardium and skeletal muscle of mice infected with high doses of virus, Cell infiltration and other pathological changes. Immunohistochemistry showed that the infected mice skeletal muscle EV71 virus specific distribution. Conclusion Fuyang EV71 mice showed better susceptibility and cytotoxicity to mice than their parental strains. The established animal model can be used for the pathogenesis of EV71 virus, the characteristics of the infection and the evaluation of the virus vaccine and drugs.