Human and mouse transcriptome profiling identifies cross-species homology of mononuclear phagocytes

人类和小鼠转录组分析鉴定单核吞噬细胞的跨物种同源性

• 批准号: 10318670
• 负责人: Claudia V Jakubzick
• 金额: $ 98.4万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-15 至 2027-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10318670
• 关键词: Air; Allergens; Alveolar Macrophages; Antigens; Bacterial Infections; Bioinformatics; Biological Assay; Biological Models; Categories; Dendritic Cells; Disease; Exposure to; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Markers; Grant; Heart; Heterogeneity; Human; Immune response; Knockout Mice; Langerhans cell; Liver; Lung; Microbe; Microscopy; Mononuclear; Morphology; Mus; Mycoses; Natural Immunity; Organ; Phagocytes; Play; Prevention strategy; Proteins; Respiratory System; Role; Skin; System; Time; Tissues; Transgenic Organisms; Validation; Virus Diseases; adaptive immune response; adaptive immunity; base; cell type; conditional knockout; draining lymph node; falls; in vitro Model; in vivo; interstitial; lymph nodes; macrophage; monocyte; mouse model; particle; single-cell RNA sequencing; therapy development; trafficking; transcriptome; transcriptome sequencing; treatment strategy; virtual

Project Summary/Abstract The mononuclear phagocyte (MP) system plays a fundamental role in both innate and adaptive immunity. It includes three broad classes of MPs extensively characterized in the mouse: (1) macrophages, including alveolar macrophages, Langerhans cells, and three distinct subtypes of interstitial macrophages; (2) tissue-trafficking monocytes; and (3) dendritic cells (DCs), which fall into two main types (DC1 and DC2), though DC2 can be further subdivided. All these MPs, except AMs and LCs, which are unique to lung and skin, reside in multiple organs, including the heart, skin, liver, and gut. MP subtypes demonstrate a clear division of labor during innate and adaptive immunity with little to virtually no functional redundancy, which means that specific interactions among them are crucial for optimal immune responses against viral, bacterial, and fungal infections. Currently, however, multiple fundamental gaps for the identification and understanding of how these MPs function in human organs limit our ability to develop prevention and treatment strategies across diseases. This project will investigate cross-species and cross-tissue homologies at the cellular, gene expression and functional levels. We will obtain fresh human and mouse tissue from multiple organs (lung, skin, and their draining lymph nodes), and employ three broad approaches. First, we will use both bulk RNA sequencing (RNA-seq) and single-cell RNA sequencing (scRNA-seq) to identify cross-species and cross-tissue homology. RNA-seq provides sequencing depth (i.e., whole-transcriptome coverage), and scRNA-seq provides the ability to confirm bulk homologous MP subtypes and examine the heterogeneity within previously defined MP subtypes. Thus, bioinformatics analyses will identify clusters of homologous MP cell types and align them across species. Second, for each cluster identified, we will identify genes conserved across species and tissues, and those that are unique to a given homologous MP subtype, termed marker genes. The results of these analyses will provide specific genetic markers for human MP subtypes and genetic treatment targets. Broadly speaking, there are two categories of key marker genes we will functionally investigate: those conserved in human-mouse MP counterparts that (1) have been well-defined in mice, but not previously investigated in their human counterparts; and (2) not well-defined or extensively studied in either species. Third, we will undertake a rigorous functional validation of the key genes identified in human-mouse MP counterparts. This includes (a) in-vivo murine models with selective depletion of specific genes using transgenic and conditional knockout (KO) mice; (b) in vitro model systems for human MPs, including assays for antigen acquisition and processing, cellular interactions, and induction of adaptive immune responses; and (c) create time-lapse videos with cellular-level microscopy for functional and morphological characterization.

项目摘要/摘要 单核吞噬细胞(MP)系统在先天和获得性中都起着基础作用。 豁免权。它包括三大类在小鼠中广泛表征的MPS：(1)巨噬细胞， 包括肺泡巨噬细胞、朗格汉斯细胞和三种不同亚型的间质巨噬细胞； 组织运输单核细胞；和(3)树突状细胞(DC)，分为两种主要类型(DC1和DC2)， 虽然DC2可以进一步细分。除了肺和皮肤独有的AM和LCS外，所有这些MPS， 存在于多个器官中，包括心脏、皮肤、肝脏和肠道。MP子类型显示出明显的 在先天性和适应性免疫期间的分娩，几乎没有功能冗余，这意味着 它们之间的特定相互作用对于针对病毒、细菌和真菌的最佳免疫反应至关重要。 感染。然而，目前在确定和理解这些问题的原因方面存在多个基本差距 MPS在人体器官中的功能限制了我们制定跨疾病预防和治疗策略的能力。 这个项目将研究细胞、基因表达的跨物种和跨组织的同源性。 和功能层面。我们将从多个器官(肺、皮肤和他们的 引流淋巴结)，并使用三种广泛的方法。首先，我们将使用批量RNA测序 (RNA-seq)和单细胞RNA测序(scRNA-seq)，以确定跨物种和跨组织同源性。 Rna-seq提供测序深度(即，整个转录组覆盖)，而scRNA-seq提供 确认批量同源MP亚型并检查先前定义的MP中的异质性 子类型。因此，生物信息学分析将识别同源MP细胞类型的集群并将它们比对 跨物种。第二，对于确定的每个簇，我们将识别跨物种和 组织和特定同源MP亚型所特有的组织，称为标记基因。结果是 这些分析将为人类MP亚型和基因治疗靶点提供特定的遗传标记。 广义地说，我们将从功能上研究两类关键标记基因： 在人-鼠MP对应物中保守，(1)已经在小鼠中很好地定义，但以前没有 在人类对应的物种中进行了研究；以及(2)在两个物种中都没有得到很好的定义或广泛的研究。 第三，我们将对人-鼠MP中确定的关键基因进行严格的功能验证 对口单位。这包括(A)体内特定基因选择性缺失的小鼠模型 转基因和条件性基因敲除(KO)小鼠；(B)人类MPS的体外模型系统，包括检测 (C)抗原获取和处理、细胞相互作用和适应性免疫反应的诱导； 使用细胞级显微镜创建延时视频，以进行功能和形态表征。