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.

项目总结/摘要 动脉粥样硬化是一种多因素疾病，占发病率和死亡率的主要原因。的 内皮细胞，血管壁的内层，从血管壁的剪切应力（WSS）中传递恒定的和有节奏的壁剪切应力（WSS）。 血液粘度和流动。在动脉分叉的内侧壁，稳定的单向层流（S流） 发展减弱全身炎症反应，而双向和轴向错位流动， 侧壁决定慢性低度炎症反应局部但偏心性质，内皮细胞 重编程和动脉粥样硬化病变的优先形成。我们公正的scRNA序列和基因本体 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一种 抗动脉粥样硬化治疗靶点仍然是难以捉摸的。长期存在的技术挑战之一是 询问细胞信号机制和功能，同时解开关键机制的行动， 病理生理学里程碑的动态变化。全面了解动脉粥样硬化和内皮 可以通过可视化整个斑块的多组图谱来实现动力学。前沿荧光 光学透明斑块中的显微镜检查提供了动脉粥样硬化的定性和定量评价。 然而，荧光成像的当前模式和常规组织的时间要求程序 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， 体素光片荧光显微术结合快速清除斑块可以解决未满足的 挑战在这项资助中，我们将使用新的组织清除方法，先进的图像采集，以阐明 流动敏感机制，从而EndICLT促进动脉粥样硬化。在目标1中，我们将检查 新的二氯甲烷-组织密度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 宽视场和衍射限制的共焦显微镜产生来自散焦照明的干涉， 减少了穿过试样的轴向穿透深度。在目标2中，我们将重点建立一个先进的 LSFM成像平台，随后进行子体素重建（SV-LSFM）和基于机器学习的图像 分割用于以高空间分辨率可伸缩地提取多组学特征。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。在一起， 这些目标将允许范式转变以鉴定动脉粥样硬化的新治疗靶点。这项建议会 让我加深我的研究经验，并提供关键的支持，建立一个坚实的基础， 从事心血管研究。