LYSOSOMES AND ATHEROSCLEROTIC FOAM CELL GENESIS

性状（改编自研究者摘要）：溶酶体脂质 在动脉粥样硬化中，积聚是一种普遍存在但知之甚少的方面， 人和易感动物这种积累的重要性建议 通过其与发展中重要事件的空间和时间联系， 从脂肪条纹到斑块的病变。此外，溶酶体隔离 病变细胞中的胆固醇似乎被截留， 在细胞质储存中。鸽和THP 1巨噬细胞的延长孵育 （人细胞系）与氧化（ox-）LDL产生的积累， (FC)溶酶体中的酯化胆固醇（CE）与病变相似。 形态学和生物化学研究表明， 溶酶体FC之后是溶酶体CE积累。这表明 最初，脂蛋白CE被水解成FC，但随后， 水解受到损害。我们假设，溶酶体FC积累，产生 通过抑制FC从溶酶体中移出，是损伤的介质 水解。本提案将检验这一假设，以确定 溶酶体蓄积机制。利用生物化学和 结构工具溶酶体FC蓄积对溶酶体CE的影响 将探索FC从溶酶体中水解和退出。治疗人类 乙酰化（ac-）LDL的细胞不产生溶酶体蓄积，即使在 胆固醇水平相当的细胞。细胞反应的差异 ox-或ac-LDL负荷将用于识别潜在的重要因素 因为溶酶体充血小鼠巨噬细胞未表现出溶酶体负荷 ox-LDL或ac-LDL。根据这一观察，可疑因素， 主要是FC，将检测诱导溶酶体负荷的能力， 小鼠细胞。最后，特定的溶酶体因子，如鞘磷脂含量 或酶运输将被检查捕获FC的能力和/或 抑制水解。这将决定这些疑似 因子在培养条件下介导溶酶体脂质积累。的 定量显微与生化相结合的分析方法 数据旨在提供对细胞后果的更广泛分析 具体操作。目标的完成将大大扩大 了解溶酶体FC和CE蓄积，并提供信息， 开始确定这种积聚在动脉粥样硬化形成中的作用。

DESCRIPTION (Adapted from Investigator's abstract): Lysosomal lipid accumulation is a ubiquitous but poorly understood aspect of atherosclerosis in man and susceptible animals. The importance of this accumulation is suggested by its spatial and temporal association with important events in development of lesions from fatty streaks to plaques. Moreover, lysosomally sequestered cholesterol in lesion cells appears trapped and less available for efflux than that in cytoplasmic stores. Prolonged incubation of pigeon and THP1 macrophages (a human cell line) with oxidized (ox-) LDL produces accumulation of both free (FC) and esterified cholesterol (CE) in lysosomes similar to that in lesions. Morphological and biochemical studies show that an initial build up of lysosomal FC is followed by lysosomal CE accumulation. This suggests that initially, lipoprotein CE is hydrolyzed to FC but that subsequent to this, hydrolysis is impaired. We hypothesize that lysosomal FC accumulation, produced by inhibition of FC movement out of lysosomes, is a mediator of the impairment of hydrolysis. This proposal will test this hypothesis in order to define mechanism(s) of lysosomal accumulation. Using a combination of biochemical and structural tools the effects of lysosomal FC accumulation on lysosomal CE hydrolysis and exit of FC from lysosomes will be explored. Treatment of human cells with acetylated (ac-)LDL does not produce lysosomal accumulation, even in cells with equivalent cholesterol levels. Differences in responses of cells to ox- or ac-LDL loading will be used to identify potentially important factors for lysosomal engorgement. Mouse macrophages do not exhibit lysosomal loading with either ox- or ac-LDL. Based on this observation, suspected factors, principally FC, will be tested for the ability to induce lysosomal loading in mouse cells. Finally, specific lysosomal factors, such as sphingomyelin content or enzyme trafficking will be examined for the ability to trap FC and/or inhibit hydrolysis. This will determine the potential of these suspected factors under culture conditions to mediate lysosomal lipid accumulation. The analytical approach of integrating quantitative microscopic and biochemical data is designed to provide a broader analysis of the cellular consequences of specific manipulations. Completion of the aims will significantly expand understanding of lysosomal FC and CE accumulation and provide information to begin determining the role of this accumulation in atherogenesis.