Development of a Novel Technology for Preparative Fractionation and Characterization of Lipoprotein Particles

脂蛋白颗粒制备分级分离和表征新技术的开发

• 批准号: 10708003
• 负责人: Angela M Zivkovic
• 金额: $ 30.36万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-25 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10708003
• 关键词: Activities of Daily Living; Acute Disease; Address; Affinity; Age; Antioxidants; Apolipoprotein A-I; Apolipoproteins; Behavior; Binding; Biological; Biology; Biomedical Research; Cardiovascular Diseases; Characteristics; Chemicals; Cholesterol Esters; Chronic; Chronic Disease; Clinical; Communities; Complement; Complex; Coupled; Data; Development; Diagnostic; Diameter; Disease; Ethnic Origin; Excision; Exclusion; Exposure to; Fractionation; Glucose; Health; Heterogeneity; High Density Lipoproteins; High Pressure Liquid Chromatography; In Vitro; Individual; Knowledge; Lipids; Lipoprotein (a); Lipoproteins; Longevity; Mass Spectrum Analysis; Measurement; Measures; Mediator; Metabolic Clearance Rate; Methods; Mission; Modification; Molecular; Molecular Sieve Chromatography; Names; National Institute of General Medical Sciences; Nature; Nerve Degeneration; Nuclear Magnetic Resonance; Optics; Oxidants; Particle Size; Performance; Physiological; Plasma; Polysaccharides; Protein Analysis; Proteins; Publishing; Refractive Indices; Research; Research Personnel; Sampling; Sepsis; Series; Signal Transduction; Structure; System; Technology; Therapeutic; Transmission Electron Microscopy; Validation; Woman; acute infection; blood-brain barrier crossing; detector; experimental study; gel electrophoresis; glycation; glycosylation; immunoregulation; improved; innovation; instrument; instrumentation; light scattering; men; mortality; new technology; oxidation; particle; receptor; reconstitution; theories; therapeutically effective

Project Summary High-density lipoproteins (HDL) are the single strongest predictors of longevity and protect against a wide array of diseases, from chronic conditions like cardiovascular disease and neurodegeneration, to acute infection and sepsis mortality, and everything in between. If we can get HDL right, we can live long, healthy lives. Yet despite over 50 years of research, HDL have remained an enigma and therapeutic approaches for improving HDL function have proven elusive. HDL are highly heterogeneous and difficult to isolate and characterize because of their colloidal, multi-molecular nature yet very small size (< 20 nm in diameter). A critical barrier to progress is the lack of technologies to simultaneously quantify the size and number of HDL particles, and isolate them such that they remain intact and amenable for a variety of both compositional and functional analyses. The objectives of this project are to gain new technological knowledge on the application of our instrument using size exclusion chromatography coupled with multiple inline static and dynamic optical detectors, and to measure the quantitative advantages of this technology over state-of-the-art approaches for the isolation and physicochemical characterization of HDL particles. In particular, this project will solve the critical problem of quantifying the number of particles using a non-destructive approach that simultaneously measures and fractionates the particles by size, making it possible to evaluate the function and composition of different size-based HDL subclasses on a per particle basis. Currently, researchers are limited by the simple problem of not having a good denominator: the only option is to express the amount of an important constituent or functional capacity in the HDL we have measured based on a rough substitute for “concentration” (e.g. total protein in the isolated fraction). Therefore, if there is more of a certain protein (or higher functional capacity) in sample A vs. B, there is no way to distinguish whether that is simply because sample A has more particles in it or whether there are more molecules of that protein (or higher functional capacity) per particle. Different sizes of HDL carry different absolute and relative amounts of individual proteins, lipids, and other components, from as few as 2 molecules of the main apolipoprotein, apolipoprotein A-I, and 12 molecules of cholesteryl ester in the smallest HDL, to as many as 4-6 molecules of apolipoprotein A-I and hundreds of molecules of cholesteryl ester, with similar variability in the concentrations of other critical components that confer dozens of functions, from antioxidant, to immunomodulatory, to anti-proteolytic to name a few. And because particle size determines binding affinity to receptors, clearance rates, and likely even whether HDL can cross the blood-brain-barrier, knowing the number of particles of different sizes, and also the per particle composition of the cargo they carry is critical to the development of sensitive, actionable diagnostics, and targeted, effective therapeutics. Thus, the technology developed in this project will profoundly enable the biomedical research community to answer critical questions about HDL functional biology across a broad array of clinical and therapeutic applications.

项目摘要 高密度脂蛋白（HDL）是寿命的单一强烈预测指标，并预防较宽的阵列 疾病，从心血管疾病和神经变性等慢性病，到急性感染和 败血症死亡率以及两者之间的一切。如果我们能做到HDL，我们可以长寿，健康的生活。目的地 超过50年的研究，HDL仍然是改善HDL的谜和治疗方法 功能已被证明难以捉摸。 HDL高度异质，由于 它们的胶体，多分子性质但尺寸很小（直径<20 nm）。进步的关键障碍是 缺乏简单地量化HDL颗粒的大小和数量的技术，并将其隔离 它们保持完整且适合各种组成和功能分析。目标 该项目的旨在获得有关使用尺寸排除的仪器应用的新技术知识 色谱与多个内联静态和动态光学检测器相结合，并测量 这项技术的定量优势比最先进的隔离和物理方法 HDL颗粒的表征。特别是，该项目将解决量化数字的关键问题 使用非破坏性方法的颗粒，该方法简单地测量并通过 大小，可以评估不同基于大小的HDL子类的功能和组成 根据粒子。目前，研究人员受到没有良好分母的简单问题的限制： 唯一的选择是在HDL中表达重要的宪法或功能能力的数量 根据“浓度”的粗糙替代品测量（例如，在分离的部分中的总蛋白质）。所以， 如果样品A与B中有更多的蛋白质（或更高的功能能力），则无法区分 是否仅仅是因为样品a中有更多的粒子，或者是否有更多的分子 每个粒子的蛋白质（或较高功能能力）。不同尺寸的HDL具有不同的绝对和相对 单个蛋白质，脂质和其他成分的量，来自主要的2个分子 载脂蛋白，载脂蛋白A-I和最小的HDL中胆固醇酯的12个分子，至4-6 载脂蛋白A-I的分子和数百个胆固醇酯的分子，在该分子中具有相似的变异性 从抗氧化剂到数十个功能的其他关键组成部分的浓度 免疫调节，以抗蛋白质水解为例。并且因为粒径决定了与 受体，清除率，甚至HDL是否可以越过血脑屏障，知道数量 不同尺寸的颗粒以及它们携带的货物的每颗粒组成对 开发敏感，可操作的诊断和有针对性的有效疗法。那，技术 在该项目中开发的将深刻地使生物医学研究界能够回答关键问题 关于在广泛的临床和治疗应用中的HDL功能生物学。