Immunopathogenesis of SARS-CoV-2 infection

我们已经编写了一份动物研究提案，该提案已获得NIAID动物护理和使用委员会的批准。 我们尚未开始研究，因为BSL 3和ABSL 3设施直到2020年夏季才开放。 我们正在等待BSL 3审查委员会批准我们的提案。 在第一年，C57 BL/6背景的WT和huACE 2转基因小鼠将感染SARS-CoV和SARS-CoV-2的临床分离株。 在第二年，还将对适应小鼠组织培养繁殖的SARS-CoV-2毒株（mSARS-CoV-2）与SARS-CoV-2的原始临床分离株进行比较，以确定其是否对小鼠ACE 2具有更大的亲和力，以及在表达小鼠ACE 2的C57 BL/6 WT小鼠中是否具有更大的毒力。 对于每种类型的实验，将检测四种病毒剂量，以确定是否可以为每种病毒开发无症状、轻度毒力或更高毒力的模型。 将对雌性和雄性动物进行检测，基于小鼠中的其他传染病模型，预计在冠状病毒疾病的临床进展和组织病理学方面存在差异，并且在感染SARS-CoV和SARS-CoV-2的人类中观察到性别差异。 因此，它们将被视为单独的队列，但在下表中的数量中合并。在该实验和所有后续实验中，将通过以下方式评估临床疾病进展：从咽拭子、血液和粪便中回收病毒;感染前和感染后多个时间间隔的血清抗体滴度、血象、血液化学和血清细胞因子;以及感染后一个月的身体外观、体重减轻和濒死和存活进展。 最迟在感染后1个月，通过CO2和颈椎脱位对动物实施安乐死。 根据我们的经验，为了观察存活率的显著差异，我们将在每个实验中需要20只小鼠/群组，并且由于实验之间的存活率的差异而重复实验3次。 这些数字应足以观察到其他临床数据的显著差异。 将需要较少的未感染小鼠，因为预期这些对照之间的差异较小。 对于本实验和后续实验，在下表中以黄色突出显示作为E列管理的动物。

We have written an Animal Study Proposal that has been approved by the NIAID Animal Care and Use Committee. We have not yet begun research because the BSL3 and ABSL3 facilities were not open until summer 2020. We are awaiting approval of our proposal by the BSL3 review committee. In the 1st Year, WT and huACE2 transgenic mice on a C57BL/6 background will be infected with clinical isolates of SARS-CoV and SARS-CoV-2. In the 2nd Year a strain of SARS-CoV-2 that has been adapted to mouse tissue culture propagation (mSARS-CoV-2) will also be compared with the original clinical isolate of SARS-CoV-2 to determine whether it has greater affinity for mouse ACE2 and greater virulence in C57BL/6 WT mice expressing mouse ACE2. For each type of experiment four virus doses will be tested to determine if models for asymptomatic, mild virulence or higher virulence can be developed for each virus. Both females and males will be tested and are expected to have differences in clinical progression of coronaviral disease and histopathology, based on other infectious disease models in mice, and differences between the sexes observed in humans infected with SARS-CoV and SARS-CoV-2. As a result, they will be treated as separate cohorts but are combined in the numbers in the table below. In this and all subsequent experiments, clinical disease progression will be assessed by virus recovery from throat swabs, blood and stool; serum antibody titers, hemograms, blood chemistries and serum cytokines before and at multiple intervals after infection; as well as physical appearance, weight loss, and progression to moribundity and survival for a month after infection. Animals will be euthanized by CO2 and cervical dislocation at the latest one-month post-infection. In our experience, to observe significant differences in survival we will need 20 mice/cohort in each experiment and repeat the experiment 3 times due to the variance in survival between experiments. These numbers should be sufficient to observe significant differences in the other clinical data as well. Fewer non-infected mice will be needed since there is expected to be less variance among these controls. Animals to be managed as Column E are highlighted in yellow in the tables below for this and subsequent experiments.