Anti-VEGF-mediated barrier closure

抗 VEGF 介导的屏障闭合

• 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
• 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

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（血管）的急性神经化 内皮生长因子））停止长期暴露于多余VEGF的血管泄漏。反对- VEGF抑制血管渗透性的能力提高和/或维持了患者的生活质量 患有普遍，无法治愈的疾病，例如增殖的糖尿病性视网膜病（PDR），糖尿病性黄斑 水肿（DME）和新血管相关的黄斑变性（NAMD）。这种临床观察 展示抗VEGF的效率，并开始一个问题，即抗VEGF如何平衡病理血管。 我们肯定的是，长期升高的VEGF改变了血管内皮的基因表达 一种放松内皮屏障的方式，反VEGF逆转了这种变化。 目的1。检验抗VEGF在转录上起作用以关闭屏障的假设。 我们的初步数据表明，长期暴露于VEGF改变了许多基因的表达， 包括编码内皮细胞屏障的已知调节剂的。这些发现支持我们 抗VEGF在转录上的作用假设以关闭屏障。该假设将被检验为 跟随。基于RNASEQ的初始相位将比较具有或具有的细胞中的基因表达谱 未用抗VEGF处理，从而确定抗VEGF不同表达的基因（DEG）。 候选人的DEG将根据其已知功能进行优先级，然后对VEGF/ant-的贡献进行贡献。 将确定VEGF介导的对屏障功能的控制。我们将确定抗VEGF取决于 关闭已被VEGF破坏的内皮细胞屏障。 目标2。研究抗VEGF对患者衍生的视网膜内皮细胞的作用机理。 AIM 1将使用健康的成年供体的原代人类永久内皮细胞（HREC）完成。 AIM 2意志 重复AIM 1，除了使用PREC，仍然是从病理血管中分离出的内皮细胞 在PDR患者中发展。这是抗VEGF治疗的靶细胞类型。确定 抗VEGF在这些细胞中的作用机理将提供临床相关的信息。 目标3。确定抗VEGF如何平静患者的病理血管。 在AIM 3中，我们将通过比较新鲜的基因表达特征来了解抗VEGF如何在患者中起作用 幼稚和抗VEGF治疗的患者的病理血管中分离的内皮。 此外，我们将比较所有3个目的的结果，以确定哪些抗VEGF介导的效应 发生在患者中，通过抗VEGF治疗培养的Prec（AIM 2）或HREC（AIM 1）来忠实地建模。 该项目将揭示抗VEGF治疗益处的分子介质。这样的信息将 删除开发生物标志物和抗VEGF替代品的当前障碍，这是需要的 满足患有多种盲目条件（例如DME，PDR和NAMD）的患者的需求。