Modeling early SARS-CoV-2 pathogenesis in human lung organoids and slice cultures

项目总结 新冠肺炎仍然是一场持续的全球健康危机，部分原因是新出现的变种。基本问题 关于SARS-CoV-2感染导致下肺炎症的部分机制，仍存在争议 由于缺乏体外感染模型。更好地了解变异生物学和免疫途径 参与SARS-CoV-2早期感染的研究可能为开发新的治疗策略提供见解。 抗病毒细胞因子I型干扰素及其下游效应已被认为与新冠肺炎有关。 SARS-CoV-2可能会抑制或促进肺中某些细胞类型的干扰素效应，这可能会有所不同 在病毒变种之间。我的初步工作表明，有机物质和肺切片培养可以用于研究 SARS-CoV-2感染的这些早期事件并确定巨噬细胞是一种激活 感染培养物中的干扰素途径。因此，我建议使用定量聚合酶链式反应，单细胞RNA测序， 和流式细胞术研究病毒变异并深入描述病毒和宿主基因和蛋白质的变化 在感染过程中表达。我将研究高度相关的细胞类型，这些细胞类型可以在类似的 到活体肺的时尚。我将研究干扰素刺激的基因反应，并鉴定细胞类型 它正在被调节，或将成为治疗干预的良好靶点。这一知识将是至关重要的 为我们抗击新冠肺炎大流行的努力做出贡献。 我目前是斯坦福大学传染病系凯瑟琳·布利什博士实验室的研究员 我正在被提升为全职讲师的过程中。斯坦福大学提供 出色的科学环境，为这个项目的成功提供了所有必要的资源 对我来说是可用的。我的长期目标是成为一名独立的内科科学家，专注于 呼吸道病毒致病机制。我的目标是建立一个研究计划，专注于模拟已建立的和 新出现的原发肺组织感染，目标是开发新的治疗方法。 为了实现这一目标，我设计了一份量身定做的职业发展计划，包括正式和 非正式训练。这次培训将增强我在肺部生物学、免疫学和测序方面的专业知识。 分析。这些主题的非正式培训将由我的导师Catherine Blish博士和 我的顾问委员会，卡尔文·郭博士、马克·克拉斯诺博士、苏珊·霍姆斯博士和本·平斯基博士。带着他们的 指导，我将完成提议的项目，提交研究手稿，获得进一步的资金，并获得 一个独立的研究职位。

PROJECT SUMMARY COVID-19 remains an ongoing global health crisis, in part due to emerging variants. Fundamental questions remain about the mechanisms by which SARS-CoV-2 infection drives inflammation in the lower lung, in part due to the lack of in vitro infection models. Better understanding of variant biology and the immune pathways involved in early SARS-CoV-2 infection could offer insights for the development of novel therapeutic strategies. The antiviral cytokines type I interferons and their downstream effects have been implicated in COVID-19. SARS-CoV-2 may inhibit or promote interferon effects in some cell types in the lung, and this may differ between viral variants. My preliminary work shows that organoids and lung slice cultures can be used to study these early events in SARS-CoV-2 infection and identifies macrophages as a cell type that activates the interferon pathway in infected cultures. Thus, I propose using quantitative PCR, single-cell RNA sequencing, and flow cytometry to study viral variants and to deeply profile the changes to viral and host gene and protein expression during infection. I will study highly relevant cell types that can interact with each other in a similar fashion to the in vivo lung. I will study the interferon stimulated gene response and identify the cell types in which it is being modulated or would be good targets for therapeutic intervention. This knowledge will be critical to our efforts to combat the COVID-19 pandemic. I am currently a fellow in Dr. Catherine Blish’s lab in the Division of Infectious Diseases at Stanford University and I am in the process of being promoted to a full time Instructor position. Stanford University offers an outstanding scientific environment, where all the resources necessary to the success of this project are made available to me. My long-term goal is to become an independent physician-scientist, with a research focus in respiratory viral pathogenesis. I aim to establish a research program focused on modeling established and emerging infections in primary lung tissue with a goal of developing new treatments. To achieve this goal, I have designed a tailored career development plan that comprises both formal and informal training. This training will enhance my expertise in lung biology, immunology, and sequencing analysis. Informal training in these topics will be provided by my mentor, Dr. Catherine Blish, and by the rest of my advisory committee, Dr. Calvin Kuo, Dr. Mark Krasnow, Dr. Susan Holmes, and Dr. Ben Pinsky. With their guidance, I will complete the proposed project, submit research manuscripts, obtain further funding, and obtain an independent research position.