ENDOCYTOSIS AND METABOLISM OF LIPOSOMES

脂质体的内吞作用和代谢

基本信息

批准号：
3297801
负责人：
KENNETH J LONGMUIR
金额：
$ 12.29万
依托单位：
UNIVERSITY OF CALIFORNIA-IRVINE
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-07-01 至 1992-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3297801
关键词：
fluorescence microscopy fluorescent dye /probe intracellular transport lipid biosynthesis lipids liposomes pinocytosis radiotracer

项目摘要

This research will study the intracellular transport and metabolism of liposomes that become cell-associated by endocytosis. Fluorescent and isotopically-labeled lipids will be prepared which have properties similar to lipids with normal fatty acids chains. Unlike most fluorescent lipids now available, these probes will not undergo appreciable rates of monomer transfer between bilayers ("lipid transfer"). Lipids probes will be incorporated into liposomes and incubated with mammalian cells in monolayer culture. Control experiments will be carried out to establish that most liposomal lipid becomes cell-associated by endocytosis, and not by lipid transfer, fusion, or absorption to the cell surface. After endocytosis, the intracellular metabolism and reutilization of lipid will be followed. Intracellular location will be determined by fluorescence microscopy and correlated with the conversion of fluorescent molecules to products of lipid biosynthesis. The metabolism of normal liposomes will be compared to the fate of pH-sensitive liposomes, which fuse with endosomal membranes prior to delivery to lysosomes. Cells will be treated with amphiphilic cations that are known to cause phospholipidosis (abnormal accumulation of lipid in lysosomes). The proposed experiments will evaluate the effect of these agents on 1) rate of uptake of liposomes, 2) rate of breakdown and release of lipid from endocytic compartments, 3) alterations in the reutilization of lysosomal products for lipid biosynthesis. It is a goal of this research to develop a convenient in vitro system for screening the effect of pharmacologic agents on lysosomal lipid metabolism.

这项研究将研究细胞内运输和脂质体的代谢，这些脂质体与细胞相关内吞作用。荧光和同位素标记的脂质将是制备具有类似于正常脂肪的脂质的特性酸链。与现在可用的大多数荧光脂质不同，这些探针不会经历可观的单体转移率在双层之间（“脂质转移”）。脂质探针将掺入脂质体中并孵育与单层培养物中的哺乳动物细胞。控制实验将执行以确定大多数脂质体脂质变为通过内吞作用与细胞相关，而不是通过脂质转移，融合或吸收到细胞表面。内吞作用后细胞内代谢和脂质的再生将是紧随其后。细胞内位置将由荧光显微镜并与荧光分子与脂质生物合成产物。这正常脂质体的代谢将与命运进行比较 pH敏感脂质体，与内体膜融合在输送到溶酶体之前。细胞将用已知已知的两亲阳离子处理引起磷脂病（脂质异常积累溶酶体）。提出的实验将评估这些药物对1）脂质体的摄取速率，2）内吞室中脂质的分解和释放，3）脂质溶酶体产品的再利用改变了脂质生物合成。这项研究的目标是发展方便体外系统用于筛选药理学剂的作用关于溶酶体脂质代谢。