Investigating the pathogenesis of Moyamoya Disease using patient derived induced pluripotent stem cells

使用患者来源的诱导多能干细胞研究烟雾病的发病机制

基本信息

批准号：
10487543
负责人：
GARY K STEINBERG
金额：
$ 19.68万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-09-15 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10487543
关键词：
3-Dimensional Adult Affect Angiogenic Factor Animal Model Arteries Biological Assay Blood Vessels Brain hemorrhage Bypass Caliber Cell Communication Cell Culture Techniques Cell Differentiation process Cell Proliferation Cell Survival Cell model Cells Cerebral Infarction Cerebrospinal Fluid Cerebrovascular Disorders Cerebrovascular system Characteristics Child Chronic Clinical Coculture Techniques Complex Cues Data Development Disease Disease Progression Disease model Endothelial Cells Environmental Risk Factor Etiology Extracellular Matrix Future Gene Expression Genetic Grant Growth High-Throughput Nucleotide Sequencing Human Hypoxia In Vitro Inflammation Ischemic Stroke Knowledge Lead Length Measures Mediating Mediator of activation protein Methods Modeling Molecular Molecular Target Moyamoya Disease Nervous System Physiology Operative Surgical Procedures Organoids Paralysed Pathogenesis Pathogenicity Pathologic Pathology Pathway Analysis Pathway interactions Patients Pattern Pharmaceutical Preparations Phenotype Platelet-Derived Growth Factor Play Process Proliferating Property Reporting Research Role Sampling Serum Signal Transduction Smooth Muscle Myocytes Stroke Techniques Technology Thinness Transient Ischemic Attack Translating Tube Up-Regulation Vascular Proliferation Vascular Smooth Muscle angiogenesis base cell type cerebral artery clinically relevant functional disability induced pluripotent stem cell insight migration novel therapeutics phenotypic biomarker revascularization surgery transcriptome transcriptome sequencing vascular smooth muscle cell proliferation

项目摘要

PROJECT SUMMARY Moyamoya Disease (MMD) is a rare, chronic cerebrovascular disease that affects the blood vessels of the brain, causing occlusion of major cerebral arteries and formation of fragile vessels in the vicinity. Clinical manifestations of MMD are transient ischemic attacks and cerebral infarctions, often leading to ischemic or hemorrhagic stroke. Invasive revascularization surgery is the only current treatment available. There may be a combination of genetic, circulating and environmental factors involved in the pathogenesis of MMD, however, the molecular mechanisms underlying MMD is largely unknown, mainly due to the lack of established MMD-specific cellular or animal models. In this proposal we aim to understand the pathogenesis of MMD by using MMD patient- derived iPSCs cellular models in combination with functional assays and high throughput sequencing approaches. The main histopathological finding in MMD is the fibro-cellular thickening of the innermost layer of the vessel (intima) which causes narrowing and occlusion of the vessel. This is likely due to an increase in proliferating vascular smooth muscle cells (VSMCs) or endothelial cells (ECs) and extracellular matrix components. Cues from ECs could cause VSMCs to switch to a phenotype that is proliferative and migrates from media to the intima, thus contributing to the thickening of the intima. Thus, we hypothesize that dysregulated signaling between VSMCs and ECs drive MMD pathology. Using MMD patient iPSC-derived ECs and VSMCs, we have established co-culture model and 3D cellular model by generating vascular organoids. Preliminary co-culture data show that both MMD ECs and VSMCs are functionally impaired when compared to healthy controls, with respect to cell proliferation and in vitro angiogenic tube stabilization. In Aim 1, we aim to characterize the functional properties of MMD iPSC-derived ECs and VSMCs in co-cultures by assessing their ability in cell proliferation, migration and tube formation in normal and hypoxic conditions. VSMC phenotype switching will be examined using specific phenotypic markers and contractility assay. We will also characterize vessel structural characteristics using our established vascular organoids generated from MMD iPSCs. In Aim 2, we will use RNA sequencing technology to investigate the transcriptome of VSMCs and ECs and identify potential molecular mediators involved in MMD. Top targets will be validated using quantitative PCR and their expression pattern will be investigated in our cellular models using immunostaining. Our study will elucidate cell-type specific factors that may drive MMD pathology. Vascular organoids from MMD may be an efficient human in vitro MMD model and provide invaluable information on MMD mechanisms. Data from our studies will advance the knowledge in MMD pathogenesis and open up new avenues of research to yield clinically relevant drug-based methods to treat MMD.

项目概要烟雾病 (MMD) 是一种罕见的慢性脑血管疾病，会影响大脑血管，导致主要脑动脉闭塞并在附近形成脆弱血管。临床表现 MMD 的主要症状是短暂性脑缺血发作和脑梗塞，通常导致缺血性或出血性中风。侵入性血运重建手术是目前唯一可用的治疗方法。可能是遗传因素的结合，循环和环境因素参与MMD的发病机制，然而，分子机制潜在的 MMD 在很大程度上是未知的，主要是由于缺乏已建立的 MMD 特异性细胞或动物模型。在本提案中，我们的目标是通过使用 MMD 患者来源的 iPSC 来了解 MMD 的发病机制细胞模型与功能测定和高通量测序方法相结合。 MMD 的主要组织病理学发现是血管最内层的纤维细胞增厚（内膜）导致血管变窄和闭塞。这可能是由于增殖的增加血管平滑肌细胞（VSMC）或内皮细胞（EC）和细胞外基质成分。提示来自 EC 的细胞可能会导致 VSMC 转变为增殖性表型并从培养基迁移到内膜，从而促进内膜增厚。因此，我们假设信号传导失调 VSMC 和 EC 之间驱动 MMD 病理。使用 MMD 患者 iPSC 衍生的 EC 和 VSMC，我们有通过生成血管类器官建立了共培养模型和3D细胞模型。初步共培养数据显示，与健康对照相比，MMD EC 和 VSMC 均出现功能受损，关于细胞增殖和体外血管生成管稳定。在目标 1 中，我们的目标是描述通过评估 MMD iPSC 衍生的 EC 和 VSMC 在细胞中的能力在共培养中的功能特性正常和低氧条件下的增殖、迁移和管形成。 VSMC表型转换将使用特定表型标记和收缩性测定进行检查。我们还将表征船舶结构使用我们建立的从 MMD iPSC 生成的血管类器官来分析特征。在目标 2 中，我们将使用 RNA 测序技术研究 VSMC 和 EC 的转录组并鉴定潜在的分子参与 MMD 的调解者。主要目标将使用定量 PCR 及其表达模式进行验证将使用免疫染色在我们的细胞模型中进行研究。我们的研究将阐明细胞类型的特异性因素这可能会导致 MMD 病理。 MMD 血管类器官可能是一种有效的人类体外 MMD 模型并提供有关 MMD 机制的宝贵信息。我们的研究数据将推进知识的发展 MMD 发病机制并开辟新的研究途径，以产生临床相关的基于药物的方法治疗MMD。

项目成果

期刊论文数量（2）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Revascularization improves vascular hemodynamics - a study assessing cerebrovascular reserve and transit time in Moyamoya patients using MRI.

血运重建可改善血管血流动力学——一项使用 MRI 评估烟雾病患者脑血管储备和转运时间的研究。

DOI：
发表时间：
2023-11
期刊：
影响因子：
0
作者：
Zhao, Moss Y;Armindo, Rui Duarte;Gauden, Andrew J;Yim, Benjamin;Tong, Elizabeth;Moseley, Michael;Steinberg, Gary K;Zaharchuk, Greg
通讯作者：
Zaharchuk, Greg

Measuring Quantitative Cerebral Blood Flow in Healthy Children: A Systematic Review of Neuroimaging Techniques.

测量健康儿童的定量脑血流量：神经影像技术的系统回顾。

DOI：
发表时间：
2024-01
期刊：
影响因子：
0
作者：
Zhao, Moss Y;Tong, Elizabeth;Duarte Armindo, Rui;Woodward, Amanda;Yeom, Kristen W;Moseley, Michael E;Zaharchuk, Greg
通讯作者：
Zaharchuk, Greg