COVID-19 and Kidney Injury: Urinary Transcriptomics of Kidney Injury in Novel Nonhuman Primate Models of SARS-CoV-2

摘要/摘要 严重急性呼吸综合征冠状病毒-2(SARS CoV-2)是新冠肺炎的病原体，是 当前全球大流行的原因是前所未有的公共卫生问题。肺是最 SARS-CoV-2严重损伤了器官，但肾脏似乎也受到了损害。在90%的患者中 新冠肺炎机械通气时，急性肾损伤起病早，导致预后差且高 住院死亡率。 血管紧张素转换酶-2(ACE2)是肾素-血管紧张素系统的一员，它反对血管紧张素II 动作并施加有益的CV效果。越来越多的证据表明，ACE-2是一种重要的 SARS-CoV-2受体。由于ACE2表达于近端小管、足细胞、系膜细胞、 鲍曼氏囊和集合管的壁上皮，新冠肺炎的作用需要进一步阐明。 在AKI的发展中。在之前的SARS疫情中，动物模型得到了广泛的应用。 冠状病毒用于模拟疾病进展并测试疫苗和治疗方法。非人灵长类(NHP)是 非常适合于模拟人类呼吸道病毒感染，主要是因为它们在解剖学和 免疫反应。查德·J·罗伊博士(杜兰国立大学传染病空气生物学主任 灵长类研究中心，该项目的联合调查者)最近产生了小说和翻译 新冠肺炎鼻腔/气管内接种非典型肺炎冠状病毒模型，能够 复制严重新冠肺炎患者的快速临床恶化。在这项先导性探索性研究中， 我们建议评估两个新的NHP(恒河猴和非洲绿)模型的尿转录组。 SARS-2-CoV-2感染及其与AKI早期标志物的关系我们的总体假设是， 由于血管紧张素转换酶2可能在新冠肺炎诱导的肾损伤中起关键作用，因此尿液中转录水平的变化 信号与尿液早期肾损伤标志物的改变有关，并反映了 SARS-CoV-2对肾脏ACE2表达细胞及肾功能的影响使用尿液转录组进行评估 从早期感染期间收集的尿液中分离的表达ACE2的细胞的单细胞RNA测序， 新冠肺炎进展、恢复期、尿中肾损伤标志物(表皮生长因子、单核细胞趋化蛋白-1、 NGAL)，以及肾功能的临床参数，我们提出了以下具体目标：测试 假设SARS-CoV2对肾脏的影响是通过尿转录组分的变化来证实的 伴有SARS-CoV-2感染的NHP的早期肾脏损伤标志物的分泌通过以下方式： A)评估尿液中肾脏损伤标志物在早期感染、进展和 新冠肺炎的恢复期。B)确定尿中排泄的早期肾损伤标志物 C)检测尿液的变化 新冠肺炎期间，在单细胞分辨率下血管紧张素转换酶2表达细胞的转录转录。的成功之路 拟议的研究将有助于阐明基于基因表达的细胞类型浓缩分析在 尿作为新冠肺炎进展过程中肾脏损害的初始信号。

ABSTRACT/SUMMARY The severe acute respiratory syndrome coronavirus-2 (SARS CoV-2), etiologic agent of the COVID-19, is the cause of the current worldwide pandemic with unprecedented public health issues. The lung is the most severely injured organ by SARS-CoV-2, but the kidney appears to be damaged as well. In 90% of patients with COVID-19 on mechanical ventilation, acute kidney injury (AKI) occurs early, leading to poor prognosis and high in-hospital mortality. Angiotensin (Ang) converting ezyme-2 (ACE2), a member of the renin-angiotensin system, opposes Ang II actions and exerts beneficial CV effects. There is mounting evidence that ACE-2, acts as a major entry receptor for SARS-CoV-2. Because ACE2 is expressed in proximal tubules, podocytes, mesangial cells, parietal epithelium of Bowman’s capsule and collecting ducts, it is critical to elucidate the effects of COVID-19 in the development of AKI. Animal models have been used extensively during previous outbreaks of SARS- CoV to model disease progression and to test vaccines and therapeutics. Non-human primates (NHPs) are ideally suited to model respiratory viral infections in humans primarily due to their similarities in anatomy and immunologic responses. Chad J. Roy, PhD (Director of Infectious Disease Aerobiology, Tulane National Primate Research Center, and Co-Investigator of this project) has recently generated novel and translational NHP models of COVID-19 using SARS-CoV-2 infection via nasal/intratracheal inoculation, which is able to reproduce the rapid clinical deterioration seen in people with severe COVID-19. In this pilot exploratory study, we propose to assess urine transcriptomics in two novel NHP (Rhesus macaque and African green) models of SARS-2-CoV-2 infection and their associations with early markers of AKI. Our overall hypothesis is that, because ACE2 may play a critical role in COVID-19-induced kidney injury, changes in urinary transcriptional signatures are associated with alterations of urinary early kidney injury markers and reflect the effects of SARS-CoV-2 on kidney ACE2-expressing cells and renal function. Using urine transcriptomics assessed by single-cell RNA-sequencing in ACE2-expressing cells isolated from urines collected during early infection, progression, and recovery phases of COVID-19, urinary measurements of kidney injury markers (EGF, MCP-1, NGAL), and clinical parameters of kidney function, we propose the following Specific aim: To test the hypothesis that the effects of SARS-CoV2 on the kidney are evidenced by changes of urine transcriptomics and accompanied by early excretion of kidney injury markers in NHPs with SARS-CoV-2 infection by mean of: A) To assess the temporal changes of urinary kidney injury markers during early infection, progression, and recovery phases of COVID-19. B) To determine whether urinary excretion of early kidney injury markers precede clinical parameters of kidney impairment and AKI; and C) To examine changes of urine transcriptomics in ACE2-expressing cells at a single-cell resolution during COVID-19. The success of the proposed study will help to elucidate the value of gene expression–based cell type enrichment analyses in urine as initial signals of renal injury during the progression of COVID-19.