FUNCTIONAL HIERARCHY OF REMNANT LIPOPROTEIN RECEPTORS

残余脂蛋白受体的功能层次

• 批准号: 2466751
• 负责人: SERGIO FAZIO
• 金额: $ 24.45万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2002-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2466751
• 关键词: Adenoviridae; antibody; apolipoprotein E; arteriosclerosis; blood lipoprotein biosynthesis; blood lipoprotein metabolism; bone marrow transplantation; clearance rate; disease /disorder model; gene expression; gene targeting; genetically modified animals; hematopoietic stem cells; laboratory mouse; liver cells; low density lipoprotein receptor; macrophage; protein binding; protein degradation; radiotracer; transfection /expression vector

DESCRIPTION (Adapted from applicant's abstract): The focus of this grant is the elucidation of the mechanism by which remnant lipoprotein particles are cleared by hepatic receptors. The Principal Investigator and his colleague, Dr. M. Linton, have pioneered the use of bone marrow transplant to study apo E delivery into genetically altered mice via transplanted macrophages. The critical observation in the preliminary data, which serves as the basis for the grant proposal, is that BMT into apo E deficient mice is capable of fully restoring normal lipid levels, but the same transplantation does not normalize lipids in animals that are deficient in both apo E and the LDL receptor. Apo E deficient animals who are homozygous for the LDL-R have a good response to BMT, but lipid values are higher than in animals with two intact genes encoding the LDL-R. Importantly, the apo E levels in the double ko animals are even higher than those found in normal mice, and data presented in the grant indicates that this apo E is present in the space of Disse in the liver, where lipoprotein capture by hepatic lipoprotein receptors should occur. These findings indicate that the apo E provided by macrophages does not substitute fully for endogenous hepatic apo E in generating a pathway that can metabolize remnant particles. The PI lists three major specific aims, each of which is divided into three subaims. The first aim is designed to assess the roles of the LDL receptor and the LRP receptor in the clearance of remnant lipoproteins using bone marrow transplantation of apo E expressing macrophages to partially re-constitute apo E expression in mice genetically deficient in apo E and the LDL-R. Turnover studies of apo B-48 and apo B-100 remnants will be analyzed before and after transplantation and the fate of lipoprotein associated apo E after internalization by either the LRP or LDL-R will be studied. In Specific Aim 2, the investigators will test their hypothesis that secretion-capture of lipoproteins in the space of Disse is critical to remnant uptake and that this process requires local apo E production by the hepatocyte. This will be accomplished using an adenovirus to express E locally in E deficient animals, before and after BMT. The effect of apo E expression on LRP activity will be assessed by radiolabeled antibodies directed against LRP in vivo, complemented by in vitro studies of apo E enriched B-VLDL binding to fibroblasts deficient in either the LDL-R or the LRP. The third specific aim will focus on the role of apo E and LRP in macrophage foam cell formation and atherosclerosis. BMT into LDL-R /apo E double ko's will permit the assessment of macrophage-specific E expression on atherosclerosis development in animals with persistently elevated lipids. Fetal hepatocyte transplantation is proposed as a method for generating LRP deficient macrophages to determine what activities of wild type macrophages are attributable to LRP. Finally, in vitro studies will be conducted to assess the role of LRP in the binding and degradation of B-VLDL.

描述（改编自申请人摘要）： 这项资助的重点是阐明 残余脂蛋白颗粒被肝脏清除的机制 受体。 首席研究员和他的同事，M。林顿， 率先使用骨髓移植来研究载脂蛋白E的传递 通过移植的巨噬细胞植入基因改变的小鼠体内。 临界 初步数据中的观察结果，作为赠款的基础 建议，是骨髓移植到apo E缺陷小鼠是能够完全 恢复正常的脂质水平，但同样的移植不能 使缺乏载脂蛋白E和低密度脂蛋白的动物的脂质正常化 受体的 对于LDL-R为纯合子的Apo E缺陷动物具有 对BMT的反应良好，但脂质值高于两个 编码LDL-R的完整基因。 更重要的是，apo E水平在 双ko动物甚至高于正常小鼠，数据显示， 在补助金中提出的表明，这种载脂蛋白E存在于空间， Disse在肝脏中，其中脂蛋白被肝脂蛋白捕获 受体应该发生。 这些结果表明，载脂蛋白E提供的， 巨噬细胞不能完全替代内源性肝载脂蛋白E， 生成一条可以代谢残留粒子的路径 PI列出了三个主要的具体目标，每个目标又分为三个 子目标 第一个目的是评估LDL受体的作用 和LRP受体在骨清除残余脂蛋白中的作用 表达apo E的巨噬细胞的骨髓移植， 在apo E基因缺陷的小鼠中重建apo E表达， LDL-R。 将对载脂蛋白B-48和载脂蛋白B-100残留物进行周转研究。 分析移植前后脂蛋白的变化 相关的apo E在被LRP或LDL-R内化后， 研究了 在具体目标2中，研究人员将测试他们的假设 脂蛋白在Disse空间的分泌捕获对于 残留摄取，这一过程需要通过 肝细胞 这将通过使用腺病毒表达E 在BMT之前和之后，局部在E缺乏动物中。 载脂蛋白E的作用 将通过放射性标记抗体评估LRP活性的表达 在体内直接针对LRP，辅之以载脂蛋白E的体外研究 富集的B-VLDL与LDL-R或LDL受体缺陷的成纤维细胞结合， LRP。 第三个具体目标将集中在载脂蛋白E和LRP的作用， 巨噬细胞泡沫细胞形成和动脉粥样硬化。 BMT转化为LDL-R /apo E 双ko’s将允许评估巨噬细胞特异性E表达 对血脂持续升高动物动脉粥样硬化的影响。 胎儿肝细胞移植是产生LRP的一种方法 缺乏巨噬细胞，以确定野生型巨噬细胞的活性 都是由LRP造成的 最后，将进行体外研究， 评估LRP在B-VLDL结合和降解中的作用。