Elucidating Virus:Host Interactions of Human Parainfluenza Virus 3 in a Novel Human Lung Tissue Explant Model

项目概要： 人副流感病毒3型（HPIV3）在美国构成了一个主要的公共卫生问题，因为它是 是五岁以下儿童住院治疗的第二大原因，也是儿童死亡率高的原因。 免疫力低下的成年人。研究HPIV 3传统上使用细胞系，小型啮齿动物 模型和人气道上皮模型，所有这些模型在提供 对人类肺部疾病的深入了解。为了填补这一知识空白，我们开发了一种新的肺 使用整个人肺组织外植体的模型。在初步数据中，我们利用了最近开发的GFP- 标记HPIV 3感染人肺组织，并显示HPIV 3可以在该系统中稳健复制， 通过流式细胞术检测GFP阳性细胞的存在来评估。我们假设， 在这种人肺组织模型中的HPIV 3感染将阐明新型病毒：宿主相互作用并提供见解 人类HPIV3的发病机制。在目标1中，我们将使用GFP标记的HPIV 3感染人肺 组织，以鉴定感染的细胞类型，并使用 单细胞RNA测序。在目的2中，我们将通过以下方式表征HPIV 3诱导的炎症反应： 使用多重ELISA检查趋化因子/细胞因子的诱导，并鉴定这些诱导的程度。 诱导发病的炎性介质由感染细胞和/或旁观者细胞产生。这些 这些数据将为HPIV3如何引起疾病提供新的见解，并有可能发现新的 免疫或抗病毒靶点。该项目提供了一个独特的培训机会，因为它将分子 克隆、传统病毒学技术、尖端单细胞RNA测序技术、翻译 研究和高维数据分析，并就如何成为一个成功的 学术科学家

PROJECT SUMMARY: Human parainfluenza virus 3 (HPIV3) constitutes a major public health concern in the United States as it is the second largest cause of hospitalization in children under the age of five and responsible for high mortality in immunocompromised adults. Studying HPIV3 has traditionally been conducted using cell lines, small rodent models and human airway epithelial models, all of which have limitations with regards to their ability to provide insights into human disease in the lung. To address this gap in knowledge, we have developed a novel lung model using whole human lung tissue explants. In preliminary data, we utilized a recently developed GFP- tagged HPIV3 to infect human lung tissue and show that HPIV3 can replicate robustly in this system, as assessed by the presence of GFP-positive cells detected by flow cytometry. We hypothesize that evaluation of HPIV3 infection in this human lung tissue model will elucidate novel virus:host interactions and provide insights into the pathogenesis of HPIV3 in humans. In Aim 1, we will use GFP-tagged HPIV3 to infect human lung tissues to identify the infected cell types and evaluate the transcriptional changes induced by infection using single cell RNA sequencing. In Aim 2, we will characterize the inflammatory response induced by HPIV3 by examining the induction of chemokines/cytokines using multiplex ELISA, and identify the extent to which these inflammatory mediators that induce morbidity are produced from infected cells and/or bystander cells. These data will provide novel insights into how HPIV3 causes disease and have the potential to identify novel immunotherapeutic or antiviral targets. This project offers a unique training opportunity, as it meshes molecular cloning, traditional virology techniques, cutting edge single cell RNA sequencing technologies, translational research and high dimensional data analysis with thorough mentorship on how to become a successful academic scientist.