Microfluidic single-cell analysis of cancer exosomes

癌症外泌体的微流控单细胞分析

• 批准号: 8883617
• 负责人: Yong Zeng
• 金额: $ 18.32万
• 依托单位: UNIVERSITY OF KANSAS LAWRENCE
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8883617
• 关键词: AMPA Receptors; Address; Agonist; Amino Acid Neurotransmitters; Animals; Behavior; Biogenic Amines; Biological; Brain; Caliber; Chemicals; Cognitive; Control Animal; Detection; Development; Disease Pathway; Dopamine; Event; Excitatory Amino Acids; Exposure to; Fiber Optics; Generations; Gilles de la Tourette syndrome; Glutamates; Goals; Huntington Disease; Hydrogen Peroxide; Knowledge; Laboratories; Life; Light; Measurement; Measures; Mental Depression; Mercury; Methodology; Methods; Microelectrodes; Mission; Modeling; Molecular Analysis; Movement Disorders; Nature; Nervous System Physiology; Neurobiology; Neuromodulator; Neurotransmitters; Nitric Oxide; Organism; Parkinson Disease; Pathway interactions; Physiologic pulse; Play; Rattus; Research; Research Personnel; Rodent Model; Role; Scanning; Serotonin; Signal Transduction; Slice; Source; System; Techniques; Technology; Testing; Time; Tissues; Ultraviolet Rays; United States National Institutes of Health; Work; addiction; animal tissue; awake; base; brain tissue; burden of illness; carbon fiber; design; disability; empowered; extracellular; gamma-Aminobutyric Acid; human disease; in vivo; innovation; insight; irradiation; millisecond; photoactivation; receptor; research study; response; temporal measurement; tool; uptake

Most eukaryotic cells secrete numerous membrane-derived vesicles of 30-150 nm in size termed exosomes. As an emerging mechanism for cell-to-cell communication, exosomes have been recently found to play important roles in a wide range of biological processes, including cancer development and metastasis. For instance, increasing evidences support the cancer-derived exosomes can reprogram the behavior of recipient cells to promote tumor growth and metastasis. Despite the significance of exosomes, our understanding of their biogenesis, molecular classification, and biological functions remain very limited. One of the challenges is to analyze exosomes released from single cells. Because cells in a tumor are known to be remarkably heterogeneous, single-cell analysis of exosomes is crucial to understanding their pathological roles in cancer. However, current “gold standard” methods can only perform ensemble measurements of exosomes released from a large cell population because of their poor isolation yield, insufficient analysis sensitivity and low throughput. In this proposal, the PI aims to develop for the first time a high-throughput single cell exosome analysis system (SCEAS) capable of probing the secretion and molecular composition of exosomes at the single cell level. The goal will be achieved via two specific aims: 1) develop a microfluidic exosome barcode chip for multiplexed, ultrasensitive immunofluorescence detection of exosomes; and 2) develop and validate a Single Cell Exosome Analysis System (SCEAS) for quantitative phenotyping of exosomes derived from single cancer cells. Success of the work will yield a key tool to enable the studies of heterogeneous exosome release by tumor cells at the single cell level, which would facilitate better understanding of intercellular signaling pathways underlying cancer development, metastasis, and drug resistance.

大多数真核细胞分泌许多大小为30-150 nm的膜衍生囊泡，称为外泌体。作为一种新兴的细胞间通讯机制，外泌体最近被发现在广泛的生物学过程中发挥重要作用，包括癌症的发展和转移。例如，越来越多的证据支持癌症来源的外泌体可以重新编程受体细胞的行为以促进肿瘤生长和转移。尽管外泌体具有重要意义，但我们对其生物起源，分子分类和生物学功能的了解仍然非常有限。挑战之一是分析从单细胞释放的外泌体。由于已知肿瘤中的细胞具有显著的异质性，因此外泌体的单细胞分析对于了解它们在癌症中的病理作用至关重要。然而，目前的“金标准”方法只能对从大细胞群体释放的外泌体进行整体测量，因为它们的分离产率差、分析灵敏度不足和通量低。在这项提案中，PI旨在首次开发一种高通量单细胞外泌体分析系统（SCEAS），能够在单细胞水平上探测外泌体的分泌和分子组成。该目标将通过两个具体目标实现：1）开发用于外泌体的多重、超灵敏免疫荧光检测的微流体外泌体条形码芯片;以及2）开发和验证用于来源于单个癌细胞的外泌体的定量表型的单细胞外泌体分析系统（SCEAS）。这项工作的成功将产生一个关键的工具，使肿瘤细胞在单细胞水平上异质外泌体释放的研究成为可能，这将有助于更好地理解癌症发展、转移和耐药性背后的细胞间信号通路。