Anti-VEGF-mediated barrier closure

抗 VEGF 介导的屏障闭合

基本信息

批准号：
10474415
负责人：
Andrius Kazlauskas
金额：
$ 37.34万
依托单位：
UNIVERSITY OF ILLINOIS AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-30 至 2025-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10474415
关键词：
Acute Address Adult Basic Science Biological Markers Blood Vessels Candidate Disease Gene Cells Chronic Clinical Data Diabetes Mellitus Disease Endothelial Cells Endothelium Enzyme-Linked Immunosorbent Assay Exposure to Extravasation Exudative age-related macular degeneration Fibrosis Gene Expression Gene Expression Profile Genes Genetic Transcription Glucose Goals Hour Human Immunofluorescence Immunologic In Vitro Invaded Learning Mediating Mediator of activation protein Modeling Molecular Monitor Operative Surgical Procedures Ophthalmologist Pathologic Patient Care Patients Permeability Phase Process Quality of life Quantitative Reverse Transcriptase PCR Research Proposals Retina Retinal Detachment Risk System Testing Therapeutic Traction Vascular Endothelial Growth Factors Vision Vitrectomy Work base bevacizumab cell type clinically relevant design diabetic differential expression end stage disease experimental study fluorescein isothiocyanate dextran improved in vitro Model macular edema member proliferative diabetic retinopathy recruit sight restoration standard of care surgery outcome tissue culture transcriptome sequencing

项目摘要

This project’s long-term goal is to elucidate how anti-VEGF (i.e. acute neutralization of VEGF (vascular endothelial growth factor)) stops leakage of blood vessels that are chronically exposed to excess VEGF. Anti- VEGF’s ability to curb blood vessel permeability has improved and/or sustained the quality of life of patients afflicted with pervasive, incurable diseases such as proliferative diabetic retinopathy (PDR), diabetic macular edema (DME) and neovascular age-related macular degeneration (nAMD). Such clinical observations demonstrate anti-VEGF’s efficacy, and beg the question of how anti-VEGF calms pathological blood vessels. We posit that chronically elevated VEGF alters gene expression in the endothelium of blood vessels in a way that relaxes the endothelial barrier, and that anti-VEGF reverses such changes. Aim 1. Test the hypothesis that anti-VEGF acts transcriptionally to close the barrier. Our preliminary data reveal that prolonged exposure to VEGF changes the expression of many genes, including those that encode known regulators of the endothelial cell barrier. These discoveries support our work hypothesis that anti-VEGF acts transcriptionally to close the barrier. This hypothesis will be tested as follows. The initial, RNAseq-based phase will compare the gene expression profile in cells that have or have not been treated with anti-VEGF and thereby identify anti-VEGF differentially expressed genes (DEGs). Candidates DEGs will be prioritized based on their known function, and then their contribution to VEGF-/anti- VEGF-mediated control of barrier function will be determined. We will identify genes that anti-VEGF depends on to close the endothelial cell barrier that has been breached with VEGF. Aim 2. Investigate the mechanism of action of anti-VEGF on patient-derived retinal endothelial cells. Aim 1 will be done with primary human retinal endothelial cells (HRECs) from a healthy adult donor. Aim 2 will be a repeat of aim 1, except using PRECs, retinal endothelial cells isolated from pathological blood vessels that develop in patients with PDR. This is the target cell type of anti-VEGF therapy. Determining the mechanism of action of anti-VEGF in these cells will provide clinically relevant information. Aim 3. Determine how anti-VEGF calms pathological blood vessels in patients. In aim 3 we will learn how anti-VEGF acts in patients by comparing the gene expression profile in freshly isolated endothelium from pathological blood vessels of treatment naïve and anti-VEGF-treated patients. In addition, we will compare the results from all 3 aims to determine which of the anti-VEGF-mediated effects that occur in patients are faithfully modeled by anti-VEGF treatment of cultured PRECs (aim 2) or HRECs (aim 1). This project will unveil the molecular mediators of anti-VEGF’s therapeutic benefit. Such information will remove current roadblocks to developing biomarkers and alternatives to anti-VEGF, which are needed to address the needs of patients afflicted with a variety of blinding conditions such as DME, PDR and nAMD.

该项目的长期目标是阐明抗 VEGF（即 VEGF 的急性中和（血管内皮生长因子））可阻止长期暴露于过量 VEGF 的血管渗漏。 VEGF 抑制血管通透性的能力改善和/或维持了患者的生活质量患有普遍的、无法治愈的疾病，例如增殖性糖尿病视网膜病变（PDR）、糖尿病黄斑水肿（DME）和新生血管性年龄相关性黄斑变性（nAMD）等临床观察。证明抗 VEGF 的功效，并提出抗 VEGF 如何镇静病理血管的问题。我们假设长期升高的 VEGF 会改变血管内皮的基因表达一种放松内皮屏障的方法，抗 VEGF 药物可以逆转这种变化。目标 1. 检验抗 VEGF 通过转录作用关闭屏障的假设。我们的初步数据表明，长期暴露于 VEGF 会改变许多基因的表达，包括那些编码已知内皮细胞屏障调节因子的基因。这些发现支持了我们的研究。抗 VEGF 通过转录作用来关闭屏障的工作假设该假设将被检验。接下来，基于 RNAseq 的初始阶段将比较具有或具有的细胞中的基因表达谱。未接受抗 VEGF 治疗，从而鉴定抗 VEGF 差异表达基因 (DEG)。候选 DEG 将根据其已知功能进行优先排序，然后根据其对 VEGF-/抗- 我们将确定 VEGF 介导的屏障功能控制，并确定抗 VEGF 依赖的基因。关闭被 VEGF 破坏的内皮细胞屏障。目标 2. 研究抗 VEGF 对患者来源的视网膜内皮细胞的作用机制。目标 1 将使用来自健康成年捐赠者的原代人视网膜内皮细胞 (HREC) 来完成。重复目标 1，但使用 PREC，即从病理血管中分离出的视网膜内皮细胞这是确定抗 VEGF 治疗的靶细胞类型。抗VEGF在这些细胞中的作用机制将提供临床相关信息。目标 3. 确定抗 VEGF 如何平息患者的病理血管。在目标 3 中，我们将通过比较新鲜的基因表达谱来了解抗 VEGF 在患者中的作用。从未经治疗和接受抗 VEGF 治疗的患者的病理血管中分离出内皮。此外，我们将比较所有 3 个目的的结果，以确定哪些抗 VEGF 介导的作用是通过对培养的 PREC（目标 2）或 HREC（目标 1）进行抗 VEGF 治疗，忠实地模拟了患者中发生的情况。该项目将揭示抗 VEGF 治疗益处的分子介质。消除当前开发生物标志物和抗 VEGF 替代品的障碍，这是满足受 DME、PDR 和 nAMD 等各种致盲病症影响的患者的需求。