Establishing Cellular Models of SARS-CoV2 Infection for COVID-19 Studies

项目摘要 2019年冠状病毒病(新冠肺炎)是美国和全球的公共卫生危机。在撰写本应用程序时，它已导致全球超过476,000名患者和1,000万名感染者死亡。美国是受影响最严重的国家，到目前为止已有超过12.3万人死亡，超过240万人感染。新冠肺炎是由一种新型的被称为严重急性呼吸综合征冠状病毒2(SARS-CoV2)的贝塔冠状病毒(CoV)引起的，据报道它会导致严重肺炎和致命性呼吸衰竭。目前还没有针对新冠肺炎的疫苗或FDA批准的药物，而且对这种新病毒的基础科学知识也很少。因此，迫切需要动物和细胞模型来开始研究疾病机制或测试治疗干预措施。在这里，我们建议探索理解新冠肺炎病毒感染的基本方面：病毒的组织嗜性和宿主的抗病毒先天免疫反应。首先，我们将测试与报告的新冠肺炎症状相关的各种组织中的病毒嗜性，并通过测定新冠肺炎建议进入受体/辅助因子ACE2和TMPRSS2在各种细胞类型中的表达来建立疾病的细胞模型，并使用病毒特异性分析和病毒载量定量技术监测随着时间的推移感染的进展。其次，我们将测试宿主细胞对SARS-CoV2感染的反应，通过测定感染期间促炎症细胞因子和参与抗病毒防御的I型干扰素的表达水平。我们还将评估感染期间的自噬流量，因为病毒通过劫持自噬在细胞中获得生长优势。

Project Summary Coronavirus Disease of 2019 (COVID-19) is a US and global public health crisis. It has led to the deaths of, at the point of writing this application, over 476,000 patients and 10 million infected individuals worldwide. The US was the most affected country, with over 123,000 deaths thus far and over 2.4 M infections. COVID-19 is caused by a novel beta-coronavirus (CoV) known as Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV2), which was reported to cause severe pneumonia and lethal respiratory failure. There is no vaccine or FDA-approved drugs for COVID-19, and there is very little knowledge about this new virus at the basic science level. Thus, animal and cellular models are urgently needed to start to investigate the disease mechanism or test therapeutic interventions. Here, we propose to explore essential aspects of understanding COVID-19 viral infection: tissue tropism of the virus and antiviral innate immune responses of the host. Firstly, we will test viral tropism in various tissues relevant to reported COVID-19 symptoms and establish cellular models of the disease, by determining the expression of COVID-19 suggested entry receptor/cofactors, ACE2 and TMPRSS2, in various cell types and monitor the progress of infection over time using virus-specific assays and viral load quantification techniques. Secondly, we will test host cellular responses to SARS-CoV2 infection by determining the expression levels during infection, of proinflammatory cytokines and type-I interferons involved in antiviral defenses. We will also assess autophagy flux during infection, given that viruses gain growth advantage in cells by hijacking autophagy.