The role of TWIST1 in smooth muscle cells during atherosclerosis

TWIST1在动脉粥样硬化过程中平滑肌细胞中的作用

• 批准号: 10250965
• 负责人: Robert Wirka
• 金额: $ 16.24万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10250965
• 关键词: Adopted; Affect; ApoE knockout mouse; Apoptosis; Area; Arterial Fatty Streak; Arteries; Atherosclerosis; Automobile Driving; Award; Bayesian Modeling; Binding; Biological Models; Biology; Blood Vessels; Cell Death; Cell Differentiation process; Cell Lineage; Cell physiology; Cells; Cessation of life; Chromatin; Coronary; Coronary Arteriosclerosis; Coronary artery; Data; Data Set; Development; Diabetes Mellitus; Differentiation Antigens; Disease; Fellowship; Fibroblasts; Gene Expression; Genes; Genetic Transcription; Genetic study; Goals; Human; Human Genetics; Hypertension; Immunofluorescence Immunologic; Immunohistochemistry; In Situ; Injury; Knock-out; Knockout Mice; Learning; Lesion; Link; Maps; Medicine; Mentors; Modeling; Molecular; Monitor; Mus; Nature; Paper; Pathway Analysis; Pathway interactions; Phenotype; Plant Roots; Positioning Attribute; Process; Proliferating; Regulation; Research; Research Personnel; Risk Factors; Role; Rupture; Sampling; Serum Response Factor; Shapes; Smooth Muscle Myocytes; TWIST1 gene; Time; Training; Transcriptional Regulation; United States; Variant; Vascular Diseases; Vascular Smooth Muscle; Work; aggressive therapy; blood lipid; career development; causal variant; cell motility; conditional knockout; design; differential expression; disorder risk; genome-wide; helix-loop-helix protein E12; human disease; in vivo; insight; migration; myocardin; novel; promoter; response; single-cell RNA sequencing; skills; targeted treatment; transcription factor

The purpose of this award is to provide Dr. Robert Wirka protected time, allowing him to obtain the training necessary to become an independent investigator using human genetic findings to define novel mechanisms driving coronary artery disease (CAD). Career development activities are designed to strengthen Dr. Wirka's skills in three key areas necessary for achieving this research paradigm: i) Linking disease-associated variation to causal genes; ii) Determining how causal genes affect cellular and vascular biology; and iii) Defining the molecular pathways and networks affected by causal genes. His mentor, Dr. Quertermous, will guide and monitor his progress towards independence. During atherosclerosis, vascular smooth muscle cells (SMCs) de- differentiate, proliferate and migrate into the lesion in a process known as “phenotypic modulation”. In a recent first-author paper in Nature Medicine, Dr. Wirka generated single-cell gene expression datasets from diseased mouse and human arteries and characterized the process of SMC phenotypic modulation in unprecedented detail. In the current proposal, Dr. Wirka has used these data to identify the transcription factor TWIST1 as a potential novel regulator of SMC phenotypic modulation. In pathway analysis of the mouse and human gene expression data, TWIST1 is strongly predicted to promote SMC phenotypic modulation. Consistent with this predicted role, TWIST1 gene expression increases in SMCs during phenotypic modulation in the mouse and human single-cell data, and preliminary studies show that TWIST1 inhibits expression of SMC differentiation markers. Importantly, TWIST1 is also strongly associated with multiple vascular diseases, including CAD, in human genetic studies. The proposed studies will determine the role of TWIST1 in vascular SMCs during disease, and determine the cellular and molecular mechanisms by which this occurs. In Aim 1, SMC- specific conditional Twist1 knockout mice will be used to determine how Twist1 affects SMC phenotype within the atherosclerotic lesion: i) assessment of single-cell gene expression will be used to determine the effect of SMC-specific Twist1 knockout on the ability of SMCs to undergo phenotypic modulation during disease and to map the molecular pathways affected by Twist1, and ii) in situ studies of the diseased artery wall will examine the effect of SMC-specific Twist1 knockout on classical aspects of lesion phenotype and the contribution of SMCs to the lesion. In Aim 2, cultured human coronary artery SMCs (HCASMCs) will be used to determine the cellular and molecular effects of TWIST1: i) the effect of TWIST1 perturbation on SMC phenotypes relevant to atherosclerosis such as differentiation, proliferation, migration, invasion, and cell death will be determined, and ii) the detailed molecular mechanism by which TWIST1 inhibits expression of the SMC differentiation marker ACTA2 will be explored. These studies will determine the cellular and molecular mechanisms by which a CAD- associated gene and potential novel SMC master regulator influences disease risk. Importantly, these studies will also provide Dr. Wirka with the final skills necessary to achieve scientific independence.

该奖项的目的是为 Robert Wirka 博士提供受保护的时间，让他能够获得培训 有必要成为一名独立研究者，利用人类遗传学发现来定义新机制 驾驶冠状动脉疾病 (CAD)。职业发展活动旨在增强 Wirka 博士的能力 实现这一研究范式所需的三个关键领域的技能： i) 将疾病相关变异联系起来 因果基因； ii) 确定因果基因如何影响细胞和血管生物学； iii) 定义 受因果基因影响的分子途径和网络。他的导师 Quertermous 博士将指导并 监测他走向独立的进展。在动脉粥样硬化期间，血管平滑肌细胞 (SMC) 在称为“表型调节”的过程中分化、增殖并迁移到病变中。在最近的一次 Wirka 博士在《自然医学》杂志上发表第一作者论文，从患病者中生成了单细胞基因表达数据集 小鼠和人类动脉并以前所未有的方式表征了 SMC 表型调节过程 细节。在当前的提案中，Wirka 博士使用这些数据将转录因子 TWIST1 识别为 SMC表型调节的潜在新型调节剂。小鼠和人类基因的通路分析 表达数据显示，强烈预测 TWIST1 会促进 SMC 表型调节。与此一致 预测的作用，在小鼠表型调节过程中，TWIST1 基因在 SMC 中的表达增加， 人类单细胞数据，初步研究表明TWIST1抑制SMC分化表达 标记。重要的是，TWIST1 还与多种血管疾病（包括 CAD）密切相关。 人类遗传学研究。拟议的研究将确定 TWIST1 在血管 SMC 中的作用 疾病，并确定发生这种情况的细胞和分子机制。在目标 1 中，SMC- 特定条件 Twist1 敲除小鼠将用于确定 Twist1 如何影响 SMC 表型 动脉粥样硬化病变：i) 单细胞基因表达的评估将用于确定 SMC 特异性 Twist1 敲除可增强 SMC 在疾病期间进行表型调节的能力 绘制受 Twist1 影响的分子途径，并且 ii) 对患病动脉壁进行原位研究将检查 SMC 特异性 Twist1 敲除对病变表型经典方面的影响以及 SMC 到达病变部位。在目标 2 中，将使用培养的人冠状动脉 SMC (HCASMC) 来确定 TWIST1 的细胞和分子效应：i) TWIST1 扰动对 SMC 表型的影响 将确定动脉粥样硬化，例如分化、增殖、迁移、侵袭和细胞死亡，并且 ii) TWIST1抑制SMC分化标志物表达的详细分子机制 将探索 ACTA2。这些研究将确定 CAD 的细胞和分子机制 相关基因和潜在的新型 SMC 主调节因子影响疾病风险。重要的是，这些研究 还将为维尔卡博士提供实现科学独立所需的最终技能。