Integration of advanced imaging and multiOMICs to elucidate pro-atherogenic effects of endothelial-to-Immune cell-like transition (EndICLT)

整合先进成像和多组学技术来阐明内皮细胞向免疫细胞样转变的促动脉粥样硬化效应 (EndICLT)

• 批准号: 10606258
• 负责人: Kyung In Baek
• 金额: $ 7.43万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10606258
• 关键词: 3-Dimensional; Accounting; Address; Antiatherogenic; Arterial Fatty Streak; Arteries; Atherosclerosis; Atlases; Attenuated; Biochemical Reaction; Biological; Biological Markers; Blood Viscosity; Blood flow; CD44 gene; CDH5 gene; Cardiovascular system; Cause of Death; Cell Reprogramming; Cells; Cellular Assay; Chromatin; Chronic; Common carotid artery; Complementary RNA; Confocal Microscopy; Cues; Data; Detection; Disease; Endothelial Cells; Endothelium; Environment; Fluorescence; Fluorescence Microscopy; Fluorescent in Situ Hybridization; Foundations; Genes; Genetic Transcription; Goals; Grant; Hydrogels; Image; Immune; Inflammation; Inflammatory Response; Isotopes; Label; Lateral; Left; Ligation; Light; Lighting; Link; Machine Learning; Macrophage; Maintenance; Maps; Medial; Mediating; Mesenchymal; Methods; Methylene Chloride; Modality; Modeling; Morbidity - disease rate; Morphology; Mus; Nature; Noise; Nucleic Acids; Ontology; Optics; PECAM1 gene; Pathway interactions; Penetration; Periodicity; Pharmaceutical Preparations; Phenotype; Procedures; Process; Proliferating; Proteins; Qualitative Evaluations; Quantitative Evaluations; RNA; RNA Sequences; Reaction; Research; Resolution; Role; Signal Transduction; Smooth Muscle Myocytes; Specimen; Stents; Structure; Techniques; Testing; Therapeutic; Time; Tissues; Touch sensation; Transposase; VWF gene; Vascular Cell Adhesion Molecule-1; Visualization; atherogenesis; career; cell type; chromophore; endothelial stem cell; epigenomics; experience; fluorescence imaging; genome-wide; hypercholesterolemia; imaging Segmentation; imaging approach; imaging platform; in vivo; insight; mortality; mouse model; multidisciplinary; multiple omics; new therapeutic target; novel; novel strategies; preservation; prevent; reconstruction; recruit; shear stress; single-cell RNA sequencing; spatiotemporal; systemic inflammatory response; therapeutic target

Project Summary/Abstract Atherosclerosis is a multifactorial disease accounting for a leading cause of morbidity and mortality. The endothelium, the inner lining of vessel walls, transduces constant and rhythmic wall shear stress (WSS) from blood viscosity and flow. At the medial wall of arterial bifurcation, stable unidirectional laminar flow (S-flow) develops attenuates systemic inflammatory responses, whereas bidirectional and axially misaligned flow in the lateral wall determine focal but eccentric nature of chronic low-grade inflammatory responses, endothelial cell reprogramming and preferential formation of atherosclerotic lesion. Our unbiased scRNA seq and gene ontology analysis suggested that D-flow induces endothelial-to-immune cell like-transition (EndICLT) and pro-atherogenic pathways. However, the pathophysiological significance of EndICLT in vivo and whether it could serve as an anti-atherogenic therapeutic target is yet remains elusive. One of the longstanding technical challenges is interrogating cell signaling machinery and function simultaneously to unravel key mechanisms in action and dynamic changes in pathophysiological milestones. Comprehensive insights into atherosclerosis and endothelial dynamics can be achieved by visualizing multiomic atlas throughout the plaque. Cutting-edge fluorescence microscopy in optically cleared plaque provides qualitative and quantitative evaluations of atherosclerosis. However, current modalities of fluorescence imaging and time demanding procedures of conventional tissue clearing techniques limits high throughput imaging with high spatial resolution. The advent of multi-scale sub- voxel light-sheet fluorescence microscopy combined with a rapid clearing of plaques may address unmet challenges. In this grant, we will use novel approaches of tissue clearing, advanced image acquisition to elucidate flow-sensitive mechanisms, whereby EndICLT promotes atherosclerosis. In Aim1, we will examine whether the novel dichloromethane - histodenz gradient medium clearing technique has potential for rapid extraction of multiomic information in optically cleared plaque. Conventional fluorescence microscopy techniques including wide-field and diffraction-limited confocal microscopy creates interference from out-of-focus illumination and reduced axial penetration depth across the specimen. In Aim 2, we will focus on establishing an advanced imaging platform of LSFM followed by sub-voxel reconstruction (SV-LSFM) and machine-learning based image segmentation for scalable extraction of multiomic features in high spatial resolution. Finally, in Aim 3, we will integrate Aim 1 and 2 to explore key underlying mechanisms in the EndICLT-dependent atherogenesis. Together, these aims will allow a paradigm shift to identify novel therapeutic targets of atherosclerosis. This proposal will allow me to deepen my research experiences and provide a critical support to build a strong foundation for a career in cardiovascular research.

项目概要/摘要 动脉粥样硬化是一种多因素疾病，是发病和死亡的主要原因。这 内皮细胞是血管壁的内层，从 血液粘度和流量。动脉分叉处内壁，稳定的单向层流（S-flow） 发展减弱全身炎症反应，而双向和轴向错位流动 外侧壁决定慢性低度炎症反应的局灶性但古怪的性质，内皮细胞 动脉粥样硬化病变的重编程和优先形成。我们公正的 scRNA seq 和基因本体论 分析表明，D-flow 诱导内皮细胞向免疫细胞样转变 (EndICLT) 和促动脉粥样硬化 途径。然而，EndICLT 在体内的病理生理学意义以及它是否可以作为 抗动脉粥样硬化的治疗目标仍然​​难以捉摸。长期存在的技术挑战之一是 同时询问细胞信号传导机制和功能，以揭示行动中的关键机制和 病理生理里程碑的动态变化。对动脉粥样硬化和内皮细胞的全面见解 动态可以通过可视化整个斑块的多组图谱来实现。尖端荧光 光学透明斑块的显微镜检查可以对动脉粥样硬化进行定性和定量评估。 然而，当前的荧光成像方式和传统组织的耗时程序 清除技术限制了具有高空间分辨率的高通量成像。多尺度子系统的出现 体素光片荧光显微镜结合快速清除斑块可能会解决未满足的问题 挑战。在这笔资助中，我们将使用组织透明化、先进图像采集的新方法来阐明 血流敏感机制，EndICLT 促进动脉粥样硬化。在目标 1 中，我们将检查是否 新型二氯甲烷-histodenz梯度介质澄清技术具有快速提取 光学透明斑块中的多组学信息。传统的荧光显微镜技术包括 宽视场和衍射极限共焦显微镜会产生离焦照明的干扰， 减少了穿过样本的轴向穿透深度。在目标2中，我们将专注于建立先进的 LSFM 成像平台，随后进行亚体素重建 (SV-LSFM) 和基于机器学习的图像 用于高空间分辨率下多组学特征的可扩展提取的分割。最后，在目标 3 中，我们将 整合目标 1 和 2，探索 EndICLT 依赖性动脉粥样硬化形成的关键潜在机制。一起， 这些目标将实现范式转变，以确定动脉粥样硬化的新治疗靶点。该提案将 让我加深我的研究经验，并为建立坚实的基础提供关键支持 从事心血管研究的职业。