Targeting SARS-CoV-2 induced lung immunopathology using novel genetic mouse models

肺免疫病理是SARS-CoV-2相关发病率和死亡率的主要原因 感染。越来越多的证据表明，SARS-CoV-2相关的肺损伤不仅导致 由病毒引起，也由过度产生的促炎细胞因子引起，称为细胞因子风暴。 尽管现有的疫苗和抗病毒药物可以预防感染，但这些策略并不特别有效。 靶向免疫介导的病理学。因此，将抗病毒宿主防御机制从 由这些机制诱导的免疫病理学为新冠肺炎提供了一种新的治疗策略 治疗。然而，为了制定这样的战略，更好地理解 使用新冠肺炎病的动物模型调节SARS-CoV-2诱导的免疫病理学是至关重要的。这个 关键差距是有限的治疗方法，具体针对免疫介导的病理和 重述人类肺部疾病的动物模型的可用性。用小鼠模型进行实验 表达SARS-CoV-2受体的人ACE2(HACE2)证实了病毒对大脑的入侵和 多器官病理，肺病理有限，不能完全概括急性呼吸道疾病 严重新冠肺炎病患者的窘迫综合征。我们正在进行的结果表明 淋巴毒素β受体(LT-R)缺陷小鼠对SARS-CoV-2诱导的保护作用 免疫病理学。我们还发现，LTβR在另一种模型中促进细胞因子风暴和肺损伤 呼吸道疾病、流感感染。这项提议的目标是开发新的小鼠模型 模拟新冠肺炎患者肺部疾病，检测LTR抑制剂阻断SARS冠状病毒感染的效果。 2.致肺损伤。我们的中心假设是hACE2在肺泡II型上皮细胞中的表达 细胞是SARS-CoV-2诱导的肺部疾病所必需的细胞，LT-R拮抗剂抑制SARS-CoV-2诱导的肺部疾病。 CoV-2诱导的肺免疫病理改变。为了检验这一假设，我们提出了两个具体目标。在目标1中， 我们将使用CRISPR-CRISPR建立在II型肺泡上皮细胞中可调节hACE2表达的小鼠。 Cas9系统。我们将用SARS-CoV-2经鼻腔感染这些小鼠，并评估肺的免疫病理学 和细胞因子的表达。在目标2中，我们将测试LTR拮抗剂对SARS冠状病毒的阻断效果。 2诱导免疫病理。我们将优化LTR抑制剂的剂量和时机，并评估肺 免疫病理学、病毒复制、细胞因子产生和保护性免疫。这个建议很有新意 并具有重要意义，因为它将产生新的动物模型，为研究SARS-CoV-2的发病机制提供更深层次的 了解病毒诱导的免疫病理调节机制并测试其可行性 靶向新型免疫调节剂LT-R无限制抑制SARS-CoV-2诱导的免疫病理 保护豁免权。

Lung immunopathology is a major cause of the morbidity and mortality associated with the SARS-CoV-2 infection. Accumulating evidence suggests that SARS-CoV-2-associated lung damage is caused not only by the virus, but also by excessive production of proinflammatory cytokines, known as cytokine storm. Although available vaccines and antiviral drugs protect against infection, these strategies do not specifically target immune-mediated pathology. Therefore, uncoupling anti-viral host defense mechanisms from the immunopathology induced by these mechanisms, represents a novel therapeutic strategy for COVID-19 treatment. However, to develop such strategies, a better understanding of the fundamental mechanisms that regulate SARS-CoV-2-induced immunopathology using animal models of COVID-19 disease is critical. The critical gap is limited therapeutic approaches that specifically target immune-mediated pathology and availability of animal models that recapitulate human lung disease. Experiments with mouse models expressing SARS-CoV-2 receptor, human ACE2 (hACE2) demonstrated virus invasion to the brain and multiple organ pathology with limited lung pathology, which does not fully recapitulate acute respiratory distress syndrome in patients with severe COVID-19 disease. Our ongoing results demonstrate that lymphotoxin beta receptor (LTR)-deficient mice are protected from SARS-CoV-2-induced immunopathology. We also found that LTβR promotes cytokine storm and lung damage in another model of respiratory disease, influenza infection. The objective of this proposal is to develop novel mouse models that mimic lung disease of COVID-19 patients and to test the efficacy of LTR inhibitor to block SARS-CoV- 2-induced lung damage. Our central hypothesis is that expression of hACE2 in type II alveolar epithelial cells is required for SARS-CoV-2 induced lung disease and that LTR antagonist inhibits SARS- CoV-2 induced lung immunopathology. To test this hypothesis, we propose two specific aims. In Aim 1, we will generate mice with regulated hACE2 expression in type II alveolar epithelial cells using CRISPR- Cas9 system. We will infect these mice intranasally with SARS-CoV-2 and evaluate lung immunopathology and cytokine expression. In Aim 2, we will test the effectiveness of LTR antagonist to block SARS-CoV- 2-induced immunopathology. We will optimize dose and timing of LTR inhibitors and evaluate lung immunopathology, viral replication, cytokine production, and protective immunity. This proposal is innovative and significant, as it will generate novel animal models to study SARS-CoV-2 pathogenesis, provide deeper understanding of the mechanisms regulating virus-induced immunopathology and test the feasibility of targeting novel immune regulator, LTR, to inhibit SARS-CoV-2 induced immunopathology without limiting protective immunity.