Eat-Me Signaling Cascade in the Apoptotic Cell Engulfment

凋亡细胞吞噬中的“吃我”信号级联

• 批准号: 8581365
• 负责人: Joseph C Ayoob
• 金额: $ 22.23万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-03 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8581365
• 关键词: ATP-Binding Cassette Transporters; Adult; Algorithms; Apoptosis; Apoptotic; Atherosclerosis; Autoimmune Diseases; Autoimmune Process; Autoimmunity; Axon; Binding Proteins; Biological Assay; Biological Models; Caenorhabditis elegans; Candidate Disease Gene; Caspase; Categories; Cell Culture Techniques; Cell physiology; Cells; Critical Pathways; Data; Development; Disease; Double-Stranded RNA; Drosophila genus; Drug Design; Eating; Embryo; Ensure; Excision; Family; Family member; Gene Expression; Genes; Genetic; Genome; Hemocytes; Homologous Gene; Human; Image; Image Analysis; Imagery; Immune response; Inflammation; Inflammatory; Injury; Insecta; Knowledge; Lead; Libraries; Longitudinal Studies; Lung diseases; Lupus; Malignant Neoplasms; Mammals; Measurement; Measures; Microinjections; Microscopy; Modeling; Nerve Degeneration; Neurodegenerative Disorders; Neuroglia; Neurons; Orthologous Gene; Pathway interactions; Phagocytes; Phenotype; Phosphatidylserines; Prevention; Process; Proteins; RNA Interference; Reaction; Recruitment Activity; Reporter; Research; Respiration Disorders; Rheumatoid Arthritis; Role; Signal Transduction; Site; Surface; System; Systemic Lupus Erythematosus; Testing; Time; annexin A5; base; cell type; fly; gene function; genome-wide; high throughput analysis; human disease; in vivo; injured; insight; macrophage; medical schools; neuronal cell body; novel; parallel processing; prevent; public health relevance; research study; respiratory; response; screening; success; therapy design; tool; uptake

DESCRIPTION (provided by applicant): Our understanding of efferocytosis or how the cell corpses are removed has been largely impeded by lack of amenable systems to study this process. This problem has been particularly intractable from the perspective of the dying cell. We recently developed a pH-based readout of efferocytosis that allows for reliable visualization and quantitative measurements of images from Drosophila cell culture and in vivo development. Our novel cell- based assay enables high-throughput analysis to identify genes necessary to produce "eat-me" signals in apoptotic cells. In addition to pursuing a candidate gene-approach, we will implement high-throughput RNAi in Drosophila cells. By integrating several powerful tools including the whole-genome RNAi libraries, high- throughput high-content imaging, computational image analysis, the extensive preexisting genetic RNAi toolkit in flies, classical Drosophila genetics, and a vast amount of pre-compiled gene expression data, we will identify novel genes involved in efferocytosis. Additional assays will help us place the identified genes into broad categories such as pro-apoptotic genes or genes required for phosphatidylserine exposure. To assess the function of gene candidates in vivo, we will measure the corpse load in the macrophages of RNAi-injected Drosophila embryos. An analysis in the parallel process of the removal of injured neurons by glia in adult flies with genetically driven RNAi in neurons will enable us to identify the most broadly-functional signals. The proposed research will lead to identification of genes that promote efferocytosis and will form the basis of our long-term studies in flies and vertebrate systems to better understand efferocytosis and apoptosis in human diseases and to modulate the underlying pathways. The knowledge gained in our studies could be applied to increase the "eat-me" signaling in diseases where efferocytic activity is diminished, such as cancer, atherosclerosis, a number of respiratory diseases, and autoimmune disorders that include systemic lupus erythematous and rheumatoid arthritis. Our understanding of efferocytosis is also important for disorders that include neurodegeneration, where this process may be up-regulated.

描述（由申请人提供）：我们对红细胞增多症或细胞尸体如何被去除的理解在很大程度上受到缺乏研究这一过程的适用系统的阻碍。从垂死细胞的角度来看，这个问题特别棘手。我们最近开发了一种基于pH值的红细胞增多症的读数，可以对果蝇细胞培养和体内发育的图像进行可靠的可视化和定量测量。我们的新的基于细胞的测定使得高通量分析能够鉴定在凋亡细胞中产生“eat-me”信号所必需的基因。除了追求候选基因的方法，我们将在果蝇细胞中实施高通量RNAi。通过整合几个强大的工具，包括全基因组RNAi文库，高通量高内容成像，计算图像分析，广泛的预先存在的遗传RNAi工具包在苍蝇，经典的果蝇遗传学，和大量的预编基因表达数据，我们将确定新的基因参与红细胞增多症。额外的分析将帮助我们将确定的基因分为广泛的类别，如促凋亡基因或磷脂酰丝氨酸暴露所需的基因。为了评估候选基因在体内的功能，我们将测量RNAi注射的果蝇胚胎的巨噬细胞中的尸体负荷。分析成年果蝇中神经胶质细胞去除受损神经元的平行过程，神经元中的遗传驱动RNAi将使我们能够识别最广泛的功能信号。这项拟议中的研究将导致识别促进红细胞增多症的基因，并将成为我们长期研究的基础。 在苍蝇和脊椎动物系统中，以更好地了解人类疾病中的红细胞增多症和细胞凋亡，并调节潜在的途径。在我们的研究中获得的知识可以被应用于增加在肿瘤细胞活性降低的疾病中的“吃我”信号传导，例如癌症、动脉粥样硬化、许多呼吸系统疾病和自身免疫性疾病，包括系统性红斑狼疮和类风湿性关节炎。我们对红细胞增多症的理解对于包括神经变性在内的疾病也很重要，在这些疾病中，这一过程可能被上调。