A spatially resolved molecular atlas of acute viral myocarditis at single-cell resolution

病毒性心肌炎是一种很难研究和诊断的异质性疾病。因为 急性病毒性心肌炎的异质性及其诊断的难度和低敏感性 心内膜心肌活检(被认为是诊断病毒性心肌炎的金标准)， 对这种疾病的分子发病机制的了解，特别是在婴儿和新生儿中 心内膜心肌活检较少进行。心脏内对病毒做出反应的特定细胞 感染、它们反应的性质以及这种反应的时空分布还不是很清楚。 从细胞和分子水平更好地了解心脏对病毒感染的时空反应 水平将提供对驱动活跃的炎症过程的病理过程的迫切需要的洞察 这是在心脏受到病毒感染后发生的。 对病毒性心肌炎的分子发病机制缺乏了解的部分原因是缺乏工具来 在单细胞分辨率下研究复杂自然组织中的病毒感染。在这里，我们将使用创新的空间 解析转录学、单细胞RNA测序(scRNA-seq)工具和生物信息学，与 经典病毒学技术，以及研究哺乳动物正病毒(REOV)心肌炎的小鼠模型。 我们有三个目标：在目标1中，我们将研究决定REOV感染结果的病毒和宿主因素 心脏组织。在目标2中，我们将确定下垂在REOV诱导的心肌炎中的作用。在目标3中，我们 将把我们的高分辨率心肌炎单细胞图谱与液体活检的原理结合起来，基于 无细胞RNA开发高度特异的病毒性心肌炎血液生物标记物，从而解决紧急和 未得到满足的医疗需求。我们预计我们的研究将为病毒的病理生物学提供前所未有的洞察力 心肌炎。我们的实验将阐明在复杂的心脏组织中感染了哪些细胞类型，并将揭示 感染成功与否取决于细胞状态和细胞环境。我们将阐明它的作用 病毒性心肌炎中的内皮细胞，我们将探索免疫细胞反应是否以及如何 主场防守要防主场受伤。我们将近距离探索感染细胞对未感染旁观者细胞的影响 物理上的接近，我们将绘制出调节这种旁观者效应的细胞相互作用。我们还将 探讨感染和未感染的先天免疫反应的空间和细胞类型的异质性 心脏组织。 这些研究的成功实施将带来研究病毒的新方法和分子工具 心肌炎和其他病毒性疾病，并可能确定新的诊断方法和治疗目标 急性病毒性心肌炎。

Viral myocarditis is a heterogeneous disease that is difficult to study and diagnose. Because of the heterogeneous nature of acute viral myocarditis and the difficulty and low diagnostic sensitivity of endomyocardial biopsies (considered the gold standard for diagnosis of viral myocarditis), there is limited knowledge of the molecular pathogenesis of this disease, particularly in infants and neonates where endomyocardial biopsies are less often performed. The specific cells within the heart that respond to viral infection, the nature of their responses, and the spatiotemporal distribution of such responses are not well known. A better understanding of the spatiotemporal response of the heart to viral infection at the cellular and molecular level will provide much needed insight into the pathological processes that drive the active inflammatory process that ensues following viral infection of the heart. The lack of understanding of the molecular pathogenesis of viral myocarditis is in part due to the lack of tools to investigate viral infection in complex native tissues at single cell-resolution. Here we will use innovative spatially resolved transcriptomics, single-cell RNA sequencing (scRNA-seq) tools and bioinformatics, in conjunction with classical virology techniques, and mouse models to study myocarditis in mammalian orthoreovirus (REOV). We have three aims: In Aim 1, we will study the viral and host factors that define the outcome of REOV infection of cardiac tissues. In Aim 2, we will determine the role of pyroptosis in REOV-induced myocarditis. In Aim 3, we will combine our high-resolution single-cell atlas of myocarditis with the principles of liquid biopsies based on cell-free RNA to develop highly specific blood biomarkers of viral myocarditis, thereby addressing an urgent and unmet medical need. We anticipate our studies will provide unprecedented insight into the pathobiology of viral myocarditis. Our experiments will clarify which cell types are infected in complex cardiac tissues and will reveal how infection success depends on both cell state and cellular environment. We will elucidate the role for endothelial cells in viral myocarditis and we will explore whether and how immune cell responses switch from host defense to host injury. We will explore the effects of infected cells on uninfected bystander cells in close physical proximity, and we will map the cellular interactions that mediate this bystander effect. We will also explore the spatial and cell type heterogeneity of innate immune responses within infected and uninfected cardiac tissues. Successful implementation of these studies will lead to new approaches and molecular tools to study viral myocarditis and other viral diseases and may identify novel diagnostic approaches and therapeutic targets for acute viral myocarditis.