Deciphering the role of lipid metabolism pathways in the RPE and their implications for AMD

解读 RPE 中脂质代谢途径的作用及其对 AMD 的影响

• 批准号: 9762116
• 负责人: Venkata Ramana Murthy Chavali
• 金额: $ 20.13万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9762116
• 关键词: Address; Affect; Age related macular degeneration; Aging; Apical; Appearance; Atherosclerosis; Biochemical; Biogenesis; Biological Assay; Blindness; Bruch' s basal membrane structure; CRISPR/Cas technology; Cell Culture Techniques; Cells; Cholesterol; Cholesterol Ester Transfer Proteins; Cholesterol Esters; Cholesterol Homeostasis; Clinical Trials; Critical Pathways; Data; Deposition; Diagnosis; Drusen; Elderly; Ensure; Epithelial; Expression Profiling; Extracellular Matrix; Eye; Fatty Acids; Functional disorder; Future; Gene Expression; Gene Proteins; Genes; Health; High Density Lipoprotein Cholesterol; Homeostasis; Human; Lipids; Lipoproteins; Low-Density Lipoproteins; Maintenance; Measures; Mediating; Metabolism; Molecular; Mutation; Nature; Outcome; Pathology; Pathway interactions; Patients; Pharmacotherapy; Phenotype; Plasma; Predisposition; Protein Overexpression; Protein Secretion; Proteins; Quantitative Trait Loci; Research Proposals; Retina; Retinal; Retinal Degeneration; Retinal Pigments; Role; Structure of retinal pigment epithelium; Testing; Transcript; Triglycerides; United States; United States National Institutes of Health; Variant; Very low density lipoprotein; age effect; choroidal circulation; drug efficacy; fetal; gene function; genome wide association study; hepatic lipase; human fetal retinal pigment epithelial cell; induced pluripotent stem cell; inhibitor/antagonist; insight; knock-down; lipid metabolism; lipid transport; lipoprotein cholesterol; loss of function; overexpression; retinal progenitor cell; stable cell line; therapeutic target; therapy development; tool; transcriptome sequencing; viral gene delivery

PROJECT SUMMARY Age-related macular degeneration (AMD) is the leading cause of blindness among the elderly in the United States. Strong association between variants in the high-density lipoprotein (HDL) cholesterol pathway genes (hepatic lipase [LIPC] and cholesterol ester transfer protein [CETP]) and advanced AMD were found in recent genome-wide association studies. The current proposal aims to understand the mechanism underlying AMD caused by SNPs in LIPC and CETP genes, which are historically known to be associated with lipid biogenesis pathways. The research proposal tests the overall hypothesis that the SNPs in LIPC and CETP exert their effects through modulating ocular lipoprotein pathways that are critical for the maintenance of retinal cholesterol homeostasis, thus causing retinal pathology. In Aim 1, we propose to (i) characterize the complete lipoprotein secretion profile including HL and CETP in the apical and basal conditioned media derived from 2D polarized primary human fetal retinal pigment epithelial (hfRPE) cultures in normal and cholesterol loaded conditions, (ii) study the effect of HL and CETP knockdown or overexpression using CRISPR/Cas9 gene editing and viral gene delivery, respectively in hfRPE cultures, and understand their function in maintenance of lipid homeostatic pathways in the RPE. We will determine the function of HL and CETP in these cultures by performing lipid efflux and influx assays, lipid secretion profile among other assays. In Aim 2, we will establish the effect of AMD-associated SNPs on LIPC and CETP transcript expression by characterizing the presence of eQTLs in these genes. We propose to use patient derived induced pluripotent stem cells (iPSC) cultures harboring AMD-associated SNPs in these genes along with isogenic controls. The iPSCs will be differentiated into well-characterized RPE cultures. These iPSC-RPE cultures will be analyzed to establish and identify any SNPs in LIPC and CETP that can regulate their expression. Outcome of our studies will open new avenues to understand the molecular pathophysiology of AMD and address the importance of HDL cholesterol metabolism pathways in maintaining retinal health. Understanding the function of genes in these pathways will not only allow us to determine potential therapeutic targets to treat AMD but also enable us to use and/or warn against the use of already available drugs for treatment of Atherosclerosis.

项目总结