Mechanism of Lysosome Functional Impairment in Cholesterol-engorged Foam Cells

胆固醇充沛的泡沫细胞溶酶体功能损伤的机制

• 批准号: 7888326
• 负责人: WALTER G JEROME
• 金额: $ 30.7万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-08 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7888326
• 关键词: Accounting; Acidity; Acids; Affect; Age; Alzheimer' s Disease; Arterial Fatty Streak; Atherosclerosis; Cells; Cholesterol; Cholesterol Esters; Cholesterol Homeostasis; Data; Development; Disease; Electron Spin Resonance Spectroscopy; Exhibits; Failure; Fluorescence; Foam Cells; Functional disorder; Genetic; Hydrolysis; Incubated; Indium; Lesion; Link; Lipids; Lipoproteins; Lysosome Proton; Lysosomes; Measures; Mediating; Membrane; Metabolism; Modeling; Participant; Pathologic; Phase; Play; Process; Property; Proton Pump; Protons; Pump; Relative (related person); Research; Role; Sphingolipids; Staging; Sterols; Testing; Triglycerides; Work; atherogenesis; base; effective therapy; enzyme activity; functional disability; in vivo; laurdan; lysosome membrane; macrophage; oxidation; oxidized lipid; particle; physical state; prevent; public health relevance; trafficking

DESCRIPTION (provided by applicant): Progressive decreases in lysosomal function have been linked to a number of diseases. One cause of decreased function is build-up of indigestible material within lysosomes, which can produce dysfunction. This link is clearest in genetic lysosomal storage disorders but is also apparent with other diseases. The occurrence of lysosomal malfunction increases with age and can be the result of secondary effects creating imbalances in cell homeostatic mechanisms and metabolism. Both innate and acquired lysosomal storage disorders almost always have pathologic consequences and are often fatal. Recently, there has been a growing understanding of the importance of lysosomes in diseases of cholesterol metabolism, such as Niemann-Pick type C, Alzheimer's and most notably atherosclerosis. Lysosomal sterol accumulation in macrophage foam cells is a ubiquitous but not well-understood aspect of atherosclerosis. Macrophages in culture incubated with atherogenic particles also exhibit an accumulation of both free and esterified cholesterol in lysosomes. Our studies show that, initially, free cholesterol accumulates as a result of normal lysosomal hydrolysis of lipoprotein-derived cholesteryl ester but the generated free cholesterol remains trapped in the lysosomes. Subsequent to the free cholesterol accumulation, cholesteryl ester hydrolysis is inhibited and so undegraded lipoprotein cholesteryl ester also accumulates in the lysosomes. Our preliminary data suggests that the inhibition of hydrolysis is mediated, at least in part, through the induction of an inability of lysosomes to maintain an acidic pH. The proposed studies systematically investigate the mechanism by which sterol accumulation inhibits the lysosome's ability to maintain a low pH and tests the hypothesis that free cholesterol, at least partially, mediates the inhibition. In aim 1, cells and isolated lysosomes are used to determine the ability of specific lipids to 1) inhibit pumping of protons into lysosomes, 2) alter the synthesis, stability and trafficking of lysosome proton pumps, 3) increase leakiness of lysosome membranes and 4) whether hydrolysis of other substrates is affected. The lipid content of lysosomes will be altered by incubation with lipoprotein or other lipid-containing particles similar to those found in atherosclerotic lesions. Aim 2 directly modulates sterol, triglyceride, and sphingolipid content of lysosomes ex-vivo to determine if any or all of these affect the lysosomes ability to maintain acidity. Aim 3 explores whether the inability of lysosomes to maintain an active pH is related to cholesterol-mediated alterations in membrane order. Finally, aim 4 investigates whether intra-lysosomal oxidation of lipid explains the trapping of free cholesterol within lysosomes and/or the subsequent inhibition of CE hydrolysis. PUBLIC HEALTH RELEVANCE: Establishing the mechanism and consequences of lysosomal sterol accumulation in foam cells will increase our understanding of atherosclerosis lesion development. The research also has broader implications for other sterol-modulated diseases such as Niemann-Pick and Alzheimer's.

描述（由申请人提供）：溶酶体功能的逐渐减少与多种疾病有关。功能降低的原因之一是溶酶体内的不可消化材料积累，这会产生功能障碍。该链接在遗传溶酶体储存障碍中最明显，但在其他疾病中也很明显。溶酶体故障的发生随着年龄的增长而增加，可能是次要作用导致细胞稳态机制和代谢失衡的结果。先天和获得的溶酶体储存障碍几乎总是具有病理性的后果，而且通常是致命的。最近，人们对溶酶体在胆固醇代谢疾病中的重要性越来越多，例如Niemann-Pick型C，阿尔茨海默氏症和最著名的动脉粥样硬化。巨噬细胞泡沫细胞中的溶酶体固醇积累是无处不在但不是动脉粥样硬化的方面。与动脉粥样硬化颗粒一起孵育的培养物中的巨噬细胞也表现出溶酶体中游离和酯化的胆固醇的积累。我们的研究表明，最初，游离胆固醇是由于脂蛋白衍生的胆固醇酯的正常溶酶体水解而累积的，但产生的游离胆固醇仍然被困在溶酶体中。在游离胆固醇积聚之后，胆固醇酯的水解受到抑制，因此未果脂蛋白胆固醇酯也积聚在溶酶体中。我们的初步数据表明，至少部分地介导了水解抑制是通过诱导溶酶体维持酸性pH的。拟议的研究系统地研究了固醇积累抑制溶酶体保持低pH值的能力的机制，并检验了游离胆固醇至少部分地介导抑制作用的假设。在AIM 1中，使用细胞和孤立的溶酶体来确定特异性脂质对1）抑制质子将质子抑制为溶酶体的能力，2）改变溶酶体质子泵的合成，稳定性和运输性，3）3）增加溶酶体膜的渗漏，以及其他肌酸液体溶解是否受影响。溶酶体的脂质含量将通过与脂蛋白或其他类似于动脉粥样硬化病变中发现的脂肪蛋白或其他含脂质的颗粒孵育来改变。 AIM 2直接调节溶酶体的固醇，甘油三酸酯和鞘脂含量，以确定是否有任何或全部影响溶酶体维持酸度的能力。 AIM 3探讨了溶酶体维持活性pH的无能与胆固醇介导的膜顺序变化有关。最后，AIM 4研究了脂质内溶酶体内氧化是否解释了自由胆固醇在溶酶体内的捕获和/或随后抑制CE水解。公共卫生相关性：建立泡沫细胞中溶酶体固醇积累的机制和后果将增加我们对动脉粥样硬化病变发展的理解。这项研究对其他固醇调节的疾病（例如尼曼·佩克和阿尔茨海默氏病）具有更广泛的影响。