Development of single-cell metabolomics to dissect cellular heterogeneity ofimmune cells

开发单细胞代谢组学来剖析免疫细胞的细胞异质性

• 批准号: 9278113
• 负责人: Cuong Q Nguyen
• 金额: $ 7.5万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-25 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9278113
• 关键词: Acetyl Coenzyme A; Address; Adenosine Diphosphate; Adenosine Triphosphate; Adenovirus Vector; Amino Acids; Anaerobic Bacteria; Animals; Autoimmune Diseases; Autoimmune Process; Autoimmune Responses; Autoimmunity; B-Lymphocytes; Biogenesis; Biological; Biological Factors; Cell Cycle; Cell Proliferation; Cell Respiration; Cell physiology; Cells; Cellular Metabolic Process; Characteristics; Chronic; Citric Acid Cycle; Complex; Data; Development; Environment; Equilibrium; Exhibits; Fatty Acids; Generations; Genetic; Genomics; Glucose; Glycolysis; Goals; Heterogeneity; Hydrolysis; Image; Immune; Immune response; Immune system; Immunologic Factors; Immunologics; Individual; Individuality; Interleukin-17; Journals; Lacrimal gland structure; Lipids; Mass Spectrum Analysis; Measurement; Measures; Memory; Metabolic; Metabolic Pathway; Metabolism; Methods; Microfluidic Microchips; Microinjections; Mus; NADH; Nicotinamide adenine dinucleotide; Nucleic Acids; Output; Oxidative Phosphorylation; Palmitic Acids; Pathogenicity; Phenotype; Phosphorylation; Population; Population Heterogeneity; Process; Production; Proteomics; Publishing; Recombinants; Resolution; Respiration; Salivary Glands; Signal Transduction; Silicones; Sister; Sjogren' s Syndrome; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Spectrometry, Mass, Secondary Ion; Techniques; Technology; Testing; Th1 Cells; Th2 Cells; Time; base; biological systems; cofactor; design; high throughput analysis; insight; ionization; long chain fatty acid; metabolic profile; metabolome; metabolomics; nano; nano-electrospray; oxidation; progenitor; response; technology development; tool; transcriptomics

ABSTRACT The immune system is comprised of a complex heterogeneity of cells with dynamic interconnectivity eliciting a diverse array of immunological responses. An individual cell progeny originated from a single clone is programmed to function and behave differently than progenitor or sister cells. Genomics of cells convey the genetic “potential”, whereas transcriptomics and proteomics describe the phenotypic cellular characteristics. Metabolomics transmits real-time biological activity and energetics, thereby providing a more accurate depiction of the cellular response. Many analytical technologies measure only the average response and fail to quantitate cell autonomy from a highly heterogeneous population of cells. This average measurement undermines the functional individuality of each cell and skews the interpretation of the whole biological system. Our preliminary data have indicated that individual T helper 17 (Th17) cells produce different levels of IL-17, and Th17 cells isolated from non-autoimmune mice preferentially use the β-oxidation metabolic pathway for energy production. However, very little is known regarding the metabolic pathway that each Th17 cell uses to initiate and sustain the pathogenicity in Sjögren's syndrome (SjS), an autoimmune disease primarily targeting the destruction of lacrimal and salivary glands. As a result, there is pertinent need to examine cellular processes at a single-cell resolution. More importantly, to obtain accurate measurement of functional processes of cellular metabolism, there is an essential need to develop a pragmatic method with high- throughput single-cell metabolomics. To accomplish this goal, we will pursue two major specific aims, Aim 1: Develop a single-cell metabolomics technology that can be applied to profile a single cell's metabolome by integrating single-cell microengraving technique with MALDI-MS, with the hypothesis: Single-cell metabolomics will have sufficient sensitivity to accurately measure metabolome of individual metabolically active cells, Aim 2: Apply single-cell metabolomics technology to compare the metabolome produced by individual Th17 cells in the salivary glands of normal versus SjSs mice, with the hypothesis: Cellular heterogeneity of autoimmune Th17 cells is driven by the changing magnitude of glycolysis versus β-oxidation of metabolic pathway. Results are expected to generate a single-cell metabolomics tool that could be used to measure the metabolomes of individual cells, and provide a proof-of- concept in deciphering the cellular heterogeneity governed by the magnitude of separate metabolic pathways resulting in diverse autoimmune responses. On a more fundamental level, results should establish the proper direction needed to move this approach forward for elucidating the metabolic pathway of highly autoimmunogenic Th17 cells based on their unique metabolite profile or “metabolic signature”.

摘要 免疫系统由具有动态互连性的细胞的复杂异质性组成， 各种各样的免疫反应源自单个克隆的单个细胞后代是 被编程为与祖细胞或姐妹细胞不同的功能和行为。细胞的基因组学传达了 遗传“潜力”，而转录组学和蛋白质组学描述的是表型细胞特征。 代谢组学传输实时生物活性和能量学，从而提供更准确的 描绘细胞反应。许多分析技术只测量平均响应， 来定量来自高度异质的细胞群体的细胞自主性。这个平均值 破坏了每个细胞的功能个体性，扭曲了对整个生物系统的解释。 我们的初步数据表明，单个T辅助17（Th 17）细胞产生不同水平的IL-17， 从非自身免疫小鼠中分离的Th 17细胞优先使用β-氧化代谢途径， 能源生产。然而，关于每种Th 17细胞用于 启动并维持干燥综合征（SjS）的致病性，这是一种主要针对 泪腺和唾液腺的破坏。因此，有必要检查细胞 以单细胞分辨率进行处理。更重要的是，为了获得功能的精确测量， 细胞代谢过程中，有必要开发一种实用的方法， 通量单细胞代谢组学。为了实现这一目标，我们将追求两个主要的具体目标，目标1： 开发一种单细胞代谢组学技术，可以应用于分析单细胞的 通过将单细胞微雕刻技术与MALDI-MS相结合，利用以下假设： 单细胞代谢组学将具有足够的灵敏度来准确地测量代谢物的代谢组。 目标2：应用单细胞代谢组学技术， 正常小鼠和SjSs小鼠唾液腺中单个Th 17细胞产生的代谢组，其中 假设：自身免疫性Th 17细胞的细胞异质性由变化的幅度驱动 糖酵解与代谢途径的β-氧化。预期结果将产生单细胞 代谢组学工具，可用于测量单个细胞的代谢组，并提供 一个解释细胞异质性的概念，细胞异质性由不同的代谢途径的大小决定 导致不同的自身免疫反应。在更根本的层面上，成果应确立适当的 需要将这种方法向前推进，以阐明高度代谢的代谢途径。 自体免疫原性Th 17细胞基于其独特的代谢物谱或“代谢特征”。