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

基本信息

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
项目类别：
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

项目摘要

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.

项目概要高密度脂蛋白 (HDL) 是唯一最强的长寿预测因子，可预防多种疾病疾病，从心血管疾病和神经退行性疾病等慢性疾病到急性感染和败血症死亡率，以及介于两者之间的一切，如果我们能够正确地获得高密度脂蛋白，我们就可以过上长寿、健康的生活。经过 50 多年的研究，HDL 仍然是一个谜，也是改善 HDL 的治疗方法 HDL 的功能已被证明难以捉摸，因为 HDL 具有高度异质性并且难以分离和表征。它们具有胶体、多分子性质，但尺寸非常小（直径< 20 nm），这是进展的一个关键障碍。缺乏同时量化 HDL 颗粒的大小和数量并将其分离的技术它们保持完整并适合各种成分和功能分析。该项目的目的是通过尺寸排除获得有关我们仪器应用的新技术知识色谱法与多个在线静态和动态光学检测器相结合，并测量该技术相对于最先进的分离和物理化学方法的定量优势特别是 HDL 颗粒的表征，该项目将解决量化数量的关键问题。使用非破坏性方法同时测量和分级颗粒大小，使得可以在基于大小的 HDL 子类上评估不同大小的功能和组成目前，研究人员受到没有良好分母的简单问题的限制：唯一的选择是表达我们拥有的 HDL 中重要成分或功能能力的数量基于“浓度”的粗略替代（例如分离组分中的总蛋白质）进行测量。如果样品 A 与 B 中某种蛋白质含量更高（或功能能力更高），则无法区分这是否仅仅是因为样品 A 中有更多的颗粒，或者是否有更多的分子每个颗粒的蛋白质（或更高的功能容量）不同大小的 HDL 具有不同的绝对和相对值。来自少至 2 个主要分子的单个蛋白质、脂质和其他成分的量最小的 HDL 中含有载脂蛋白、载脂蛋白 A-I 和 12 分子胆固醇酯，多达 4-6 个载脂蛋白 A-I 分子和数百个胆固醇酯分子，具有相似的变异性具有多种功能的其他关键成分的浓度，从抗氧化剂到免疫调节、抗蛋白水解等等，因为颗粒大小决定了结合亲和力。受体、清除率，甚至可能还有 HDL 是否可以穿过血脑屏障，知道数量不同尺寸的颗粒，以及它们所携带的货物的每个颗粒的组成对于开发灵敏的、可操作的诊断方法和有针对性的、有效的治疗方法。该项目中开发的技术将深刻地帮助生物医学研究界回答关键问题关于 HDL 功能生物学在广泛的临床和治疗应用中的知识。