High Density Lipoprotein Biogenesis and Speciation

高密度脂蛋白生物发生和形态形成

• 批准号: 9321088
• 负责人: Henry J. Pownall
• 金额: $ 58.06万
• 依托单位: METHODIST HOSPITAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-01 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9321088
• 关键词: ATP binding cassette transporter 1; Address; Amino Acids; Animals; Antiatherogenic; Apolipoprotein A-I; Apolipoproteins; Apolipoproteins B; Atherosclerosis; Biogenesis; Biophysics; Cardiovascular Diseases; Cell membrane; Cellular biology; Chemicals; Cholesterol; Circular Dichroism; Competence; Complex; Dependovirus; Disease Outcome; Dissociation; Economics; Esterification; Event; Excretory function; Family; Functional disorder; Gene Delivery; Head; Hepatic; Hepatocyte; High Density Lipoprotein Cholesterol; High Density Lipoprotein therapy; High Density Lipoproteins; Human; Immune; In Vitro; Intervention; Kinetics; LDL Cholesterol Lipoproteins; Lecithin; Lipid Binding; Lipids; Lipoproteins; Liver; Mediating; Membrane; Membrane Lipids; Methods; Modeling; Modification; Molecular; Molecular Conformation; Molecular Target; Mus; Pathway interactions; Patients; Peptide Signal Sequences; Performance; Peripheral; Pharmaceutical Preparations; Phosphatidylcholine-Sterol O-Acyltransferase; Phospholipids; Physiology; Plasma; Play; Precipitation; Process; Production; Proline; Property; Protein Analysis; Proteins; Public Health; Quality of life; Risk Factors; Role; Scientist; Side; Site-Directed Mutagenesis; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Structure; Surface; Tail; Testing; Therapeutic; Thin Layer Chromatography; Time; Tissues; Turbidimetry; Variant; analog; base; cardiovascular risk factor; chemical kinetics; cost; crosslink; design; high density lipoprotein-2; high density lipoprotein-3; improved; in vivo; interest; intrahepatic; lipophilicity; macrophage; mimetics; monolayer; multidisciplinary; novel; particle; peptide B; peptide structure; reverse cholesterol transport; uptake

Cardiovascular disease (CVD) is a public health challenge with a major economic and quality-of-life impact on its victims and their families. Impressive inroads have been made in the management of some CVD lipid risk factors; the statin class of hypolipidemic drugs reduces the number of CVD events by reducing plasma levels of low density lipoprotein-cholesterol. Reduction in CVD via the modification of other lipid-risk factors has been a challenge, especially raising the plasma concentrations of high density lipoproteins-cholesterol (HDL-C), in a cardioprotective way; attempts to reduce CVD via raising HDL-C have not been uniformly successful. Many scientists now appreciate that the mechanistic connections between HDL and atheroprotection are more complex than once thought and that a greater focus on HDL function, rather that HDL-C concentrations alone, is needed. HDL plays a role in reverse cholesterol transport (RCT), the transport of cholesterol in peripheral tissues, including macrophages in the subendothelial space of the arterial wall, to the liver for disposal. The major RCT steps are macrophage cholesterol efflux, HDL-C esterification in plasma, and hepatic disposal, which directs some cholesterol to excretion thereby reducing the total body cholesterol burden. Recent evidence is that not all HDL are the same, i.e., HDL are speciated according to composition, structure, and likely function. Several studies have shown cholesterol efflux to apo B-depleted plasma is lower in CVD patients, independent of HDL-C concentrations. This difference must be due to differences in the “quality” of HDL in patients vs. those without CVD. Some HDL species may be better cholesterol acceptors; apo AI may play a direct role in efflux. Moreover, mimetics based on an optimized apo AI structure may be a promising therapeutic strategy. The aims of this application focus on three aspects of HDL and apo AI structure and/or function— speciated biogenesis of HDL with intact signal peptides (SP); the mechanism of efflux from macrophages giving nascent HDL and microsolubilization as a surrogate for nascent HDL formation, and optimization of apo AI structure for RCT. This background and rationale gave rise to three aims: Aim 1: To test the hypothesis that the biogenesis of HDL containing apos with intact signal peptides (SPs) is unique and speciated, and forms unique HDL species with distinct compositions and functions. Aim 2: To test a novel, hypothetical “pancake” model of phospholipid microsolubilization by apo AI, which yields rHDL, the in vitro analog of nascent HDL produced by hepatocytes and MФ cholesterol efflux. Aim 3: To test the hypothesis that conserved amino acid residues in apo AI are essential to its structure and function. Completion of these aims will reveal new molecular targets for cardioprotective HDL therapies. These aims will be addressed by multidisciplinary in vitro and in vivo approaches that includes chemical kinetics, cell biology, lipid and protein analyses, AAV-mediated gene delivery in vivo, high performance thin-layer chromatography, immune-blotting and precipitation methods, chemical cross-linking, MALDI MS, site-directed mutagenesis, circular dichroism, and kinetic turbidimetry.

心血管疾病（CVD）是一项公共卫生挑战，对经济和生活质量造成重大影响 对受害者及其家人的影响。在以下方面取得了显著进展： 一些CVD脂质风险因素;他汀类降血脂药物通过以下方式减少CVD事件的数量： 降低低密度脂蛋白-胆固醇的血浆水平。通过对以下物质的改性来减少CVD 其他血脂风险因素一直是一个挑战，特别是提高高密度脂蛋白的血浆浓度， 脂蛋白胆固醇（HDL-C），以保护心脏的方式;试图通过提高HDL-C来减少CVD 并不是所有人都成功的。许多科学家现在意识到， 高密度脂蛋白和动脉粥样硬化保护之间的关系比以前想象的要复杂得多， 需要HDL功能，而不是单独的HDL-C浓度。HDL在逆转胆固醇中起作用 胆固醇转运（RCT），胆固醇在外周组织中的转运，包括巨噬细胞中的巨噬细胞。 动脉壁的内皮下空间，到肝脏处置。RCT的主要步骤是巨噬细胞 胆固醇流出、血浆中HDL-C酯化和肝脏处置，这将一些胆固醇 从而减少全身胆固醇负担。最近的证据表明，并非所有的HDL 相同的，即，HDL是根据组成、结构和可能的功能来分类的。几项研究 已经表明，CVD患者的胆固醇流出到载脂蛋白B耗尽的血浆中较低，与HDL-C无关 浓度的这种差异一定是由于患者HDL的“质量”与 没有CVD。一些HDL种类可能是更好的胆固醇受体;载脂蛋白AI可能在 流出此外，基于优化的载脂蛋白AI结构的模拟物可能是一种有前途的治疗方法， 战略 本申请的目的集中在HDL和apo AI结构和/或功能的三个方面- 具有完整信号肽（SP）的HDL的物种生物合成;巨噬细胞的流出机制 给出新生HDL和微溶解作为新生HDL形成的替代物， 载脂蛋白AI结构的随机对照试验。这一背景和理由产生了三个目标： 目的1：验证含有完整信号肽的HDL的生物来源假说 是独特的和物种化的，并形成具有不同组成和功能的独特HDL种类。 目的2：测试载脂蛋白AI对磷脂微溶解的一种新的、假设的“煎饼”模型， 其产生rHDL，由肝细胞和M β胆固醇产生的新生HDL的体外类似物 流出 目的3：验证载脂蛋白AI的保守氨基酸残基是其结构所必需的假说 和功能 这些目标的完成将揭示心脏保护HDL疗法的新分子靶点。这些 目标将通过多学科的体外和体内方法来解决，包括化学 动力学、细胞生物学、脂质和蛋白质分析、AAV介导的体内基因递送、高性能 薄层层析法、免疫印迹法和沉淀法、化学交联法、MALDI MS、定点诱变、圆二色性和动态比浊法。