Phenotypic and functional analysis of immune cells during severe COVID-19

2019年新型冠状病毒病（COVID-19）的后果给全球人民、公共卫生和卫生保健系统带来了巨大挑战。为了控制SARS-CoV2大流行，了解宿主对这种病毒的反应至关重要。一方面，确定免疫激活机制将有助于确定疾病过程中的治疗靶点，特别是因为免疫病理导致COVID-19期间的组织损伤。另一方面，深入表征针对SARS-CoV2的免疫记忆反应将有助于设计有效的保护性疫苗接种策略。在这里，我们建议将尸检组织中的多重组织学与空间分解转录组学分析相结合，以表型表征和绘制严重COVID-19期间的免疫反应。我们将分析作为疾病靶器官的肺的各个时间点，以及同一患者对应的次级淋巴器官即引流淋巴结和扁桃体的时间点。我们将在3d培养模型中使用功能性荧光寿命成像，以验证先天机制（如NETosis）是组织损伤的一个驱动力的假设，并评估巨噬细胞和中性粒细胞之间的通信。由于假设针对SARS-CoV2的先天免疫反应失调也会影响适应性免疫反应，因此我们计划表征组织中的T细胞和B细胞反应，重点关注组织特异性免疫记忆的产生，以及促进这一过程的细胞和分子微环境的表征。本提案以协同的方式，将三位申请人在病理学、免疫学和最新成像技术方面的专业知识结合起来，以阐明严重COVID-19的免疫病理机制，并探索驻留记忆细胞的组织特异性保护潜力。

The consequences of the novel coronavirus disease 2019 (COVID-19) are posing great challenges to the people, public health and healthcare systems worldwide. In order to get the SARS-CoV2 pandemic under control, it is of utmost importance to understand the host response against this virus. On the one hand, characterizing the mechanisms of immune activation will help to identify therapeutic targets in the course of the disease, especially since immune pathology is contributing to tissue damage during COVID-19. On the other hand, an in-depth characterization of the immunological memory response against SARS-CoV2 will help to design efficient, protective vaccination strategies. Here, we propose to combine multiplexed histology with spatially resolved transcriptomic analyses in autopsy tissues, in order to phenotypically characterize and map the immune response during severe COVID-19. We will analyze various time points in the lung as a target organ of the disease, as well as in the corresponding secondary lymphoid organs of the same patients namely the draining lymph nodes and tonsils. We will employ functional fluorescence lifetime imaging in 3D-culture models in order to test the hypothesis that innate mechanisms such as NETosis are one driving force for tissue damage, and assess the communication between macrophages and neutrophils. Since a dysregulated innate immune response against SARS-CoV2 is hypothesized to also impact on the adaptive immune response, we are planning to characterize T cell and B cell responses in the tissues, with a focus on the generation of tissue-specific immunological memory, and on the characterization of the cellular and molecular microenvironment that fosters this process.In a synergistic way, this proposal combines the expertise of the three applicants with respect to pathology, immunology and state-of-the-art imaging technology, in order to elucidate the mechanisms of immune pathology underlying severe COVID-19, and to explore the potential of tissue-specific protection by resident memory cells.