Macrophage-specific function of GWAS CAD-associated LIPA alleles in atherosclerosis

GWAS CAD 相关 LIPA 等位基因在动脉粥样硬化中的巨噬细胞特异性功能

基本信息

批准号：
9765372
负责人：
Hanrui Zhang
金额：
$ 24.44万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-01 至 2021-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9765372
关键词：
Acceleration Acid Lipase Address Alleles Apolipoprotein E Arterial Fatty Streak Atherosclerosis Automobile Driving Bone Marrow Transplantation Cell Communication Cholesterol Cholesterol Ester Storage Disease Cholesterol Esters Cleaved cell Clustered Regularly Interspaced Short Palindromic Repeats Competence Coronary Arteriosclerosis Crystal Formation Crystallization Data Endothelial Cells Enzymes Foam Cells Functional disorder Genes Genotype Hepatic Human Hydrolase Hydrolysis Hyperlipidemia Inflammasome Inflammation Knock-in Lesion Lipase Lipids Low-Density Lipoproteins Mediating Messenger RNA Modification Mus Oxidative Stress Phase Phenotype Plasma Proton Pump Research Role Smooth Muscle Myocytes Subfamily lentivirinae Susceptibility Gene System Therapeutic Toxic effect Translational Research Triglycerides Vascular Smooth Muscle atherogenesis causal model extracellular gain of function genome wide association study hepatic lipase in vivo induced pluripotent stem cell loss of function mutation mRNA Expression macrophage migration monocyte mouse model novel overexpression prevent promoter risk variant uptake vacuolar H+-ATPase vascular smooth muscle cell proliferation

项目摘要

DESCRIPTION (provided by applicant): Lysosomal acid lipase (LAL), encoded by the LIPA gene, is the key lysosomal hydrolase that cleaves cholesteryl esters (CE) and triglycerides (TG). Loss-of-function (LOF) mutations in LIPA result in cholesteryl ester storage disease (CESD), which manifests with hyperlipidemia, hepatic and macrophage CE accumulation, and atherosclerosis. Recently, genome-wide association studies (GWASs) identified LIPA as a novel locus for coronary artery disease (CAD). Surprisingly, LIPA CAD-GWAS risk alleles do not associate with altered plasma lipids or hepatic LIPA mRNA levels but actually relate to higher monocyte LIPA mRNA expression. Our preliminary data also reveal a coincident increase in both LIPA mRNA and LAL enzymatic activity in monocyte-derived macrophages (HMDM) of CAD risk allele carriers, suggesting that monocyte/macrophage-specific gain-of-function (GOF) of LIPA may explain the GWAS CAD risk alleles. LIPA mRNA and LAL activity were markedly induced upon HMDM differentiation and mature HMDM secrets LAL. Extracellular LAL is abundant in the neointima of advanced human atherosclerotic lesions where LAL can remain enzymatically active in the lesion acidic microenvironment. Thus, macrophage LIPA GOF may aggravate atherosclerosis through accelerating intracellular LAL-induced lysosomal free cholesterol (FC) toxicity and extracellular LAL actions on low-density lipoprotein (LDL). My working hypotheses are that LIPA CAD risk alleles encode for macrophage-specific LIPA GOF, and therefore CAD risk alleles or macrophage LIPA overexpression will (i) increase intracellular LAL activity resulting in a shift toward greater lysosomal CE hydrolysis and FC accumulation and thus accelerate macrophage lysosomal dysfunction during modified-LDL loading; (ii) increase macrophage LAL secretion and extracellular LAL-mediated LDL modification driving atherogenic phenotypes in vascular smooth muscle cells (VSMC) and endothelial cells (EC); and (iii) macrophage LIPA overexpression will accelerate atherogenesis in ApoE-/- and Ldlr-/- mice. These hypotheses will be addressed in primary HMDM of specific LIPA genotype, through causal modeling in a novel human iPSC- differentiated macrophage (IPSDM) system utilizing CRISPR/Cas knock-in of risk alleles, in HMDM with lentivirus-mediated LIPA overexpression (Lenti-LIPA), and in murine models with macrophage-specific Lipa overexpression by lentivirus-infected bone marrow (BM) transplantation driven by macrophage-specific promoter. These will be accomplished by pursuing the following three aims: Aim 1. K99 Phase: Determine the effects of LIPA CAD risk alleles and LIPA overexpression on intracellular macrophage LIPA expression, LAL activity, and phenotype of human macrophages. Aim 2. R00 Phase: Determine the effects of LIPA CAD risk alleles and LIPA overexpression on macrophage LAL secretion, extracellular LAL-modification of LDL and on atherogenic phenotypes in EC and VSMC. Aim 3. R00 Phase: Determine if macrophage Lipa overexpression accelerates atherosclerosis in ApoE-/- and Ldlr-/- mice.

描述（由应用提供）：由LIPA基因编码的溶酶体酸脂肪酶（LAL）是裂解胆固醇酯（CE）和甘油三酸酯（TG）的关键溶酶体水解酶。 LIPA的功能丧失（LOF）突变导致胆固醇酯储存疾病（CESD），其表现为高脂血症，肝和巨噬细胞CE的积累以及动脉粥样硬化。最近，全基因组关联研究（GWASS）将LIPA鉴定为冠状动脉疾病（CAD）的新型基因座。令人惊讶的是，LIPA CAD-GWAS风险等位基因与血浆脂质变化或肝LIPA mRNA水平不同，但实际上与较高的单核细胞LIPA mRNA表达相关。我们的初步数据还揭示了CAD风险等位基因载体的单核细胞衍生巨噬细胞（HMDM）中LIPA mRNA和LAL酶活性的同时增加，这表明LIPA的单核细胞/巨噬细胞特异性功能获得（GOF）可能会解释GWAS CAD CAD CAD CAD CAD CAD CAD REALES。 HMDM分化和成熟的HMDM秘密LAL明显诱导LIPA mRNA和LAL活性。在晚期人动脉粥样硬化病变的新内膜中，细胞外LAL很丰富，其中LAL可以在病变酸性微环境中保持酶活性。巨噬细胞Lipa GOF可能会通过加速的细胞内LAL诱导的溶酶体游离胆固醇（FC）毒性和低密度脂蛋白（LDL）上的细胞外LAL作用加剧动脉粥样硬化。 My working hypotheses are that LIPA CAD risk alleles encode for macrophage-specific LIPA GOF, and therefore CAD risk alleles or macrophage LIPA overexpression will (i) increase intracellular LAL activity resulting in a shift towards greater lysosomal CE hydrolysis and FC acrylic and thus acrylic lysosomal dysfunction during modified-LDL loading; （ii）增加巨噬细胞LAL分泌和细胞外LAL介导的LDL修饰，驱动血管平滑肌细胞（VSMC）和内皮细胞中的动脉粥样硬化表型（EC）；（iii）巨噬细胞LIPA过表达将在ApoE - / - 和LDLR - / - 小鼠中丙烯酸动脉粥样硬化。这些假设将通过特定LIPA基因型的初级HMDM来解决，通过利用CRISPR/CAS敲入的新型人IPSC分化巨噬细胞（IPSDM）系统的因果建模，在HMDM中，HMDM与LINTIVIRUS介导的LIPA Expressive（Lipi-Lipa）和Mishage Mishage Millage模型（MIRMIRINE MIL MIL）中的HMDM中受巨噬细胞特异性启动子驱动的慢病毒感染的骨髓（BM）移植。这些将通过追求以下三个目的来实现：目标1。K99阶段：确定LIPA CAD风险等位基因和LIPA过表达对人类巨噬细胞的细胞内巨噬细胞LIPA表达，LAL活性和表型的影响。 AIM2。R00相：确定LIPA CAD风险等位基因和LIPA过表达对巨噬细胞分泌，LDL的细胞外LAL-修饰以及EC和VSMC中动脉粥样硬化表型的影响。 AIM3。R00相：确定巨噬细胞Lipa过表达是否会加速APOE - / - 和LDLR - / - 小鼠中的动脉粥样硬化。