Hydrogen peroxide in endothelial function and dysfunction
过氧化氢在内皮功能和功能障碍中的作用
基本信息
- 批准号:10320952
- 负责人:
- 金额:$ 44.22万
- 依托单位:
- 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:2021-01-01 至 2024-12-31
- 项目状态:已结题
- 来源:
- 关键词:AlanineAmino AcidsAnimal ModelAntioxidantsBiosensorBlood PressureBlood VesselsCardiomyopathiesCardiovascular DiseasesCatalysisCaveolaeCell NucleusCell Surface ReceptorsCellsClinical TrialsCountryCytosolD-Amino Acid DehydrogenaseDevelopmentDiabetes MellitusDiseaseEndothelial CellsEndotheliumExperimental ModelsFunctional disorderGenerationsGenetic TranscriptionGoalsHeartHeart failureHomeostasisHumanHydrogen PeroxideHypertensionImageIn VitroIntracellular MembranesInvestigationLeadMetabolismModelingMolecularMorbidity - disease rateNF-kappa BNOS3 geneNitric OxideNitric Oxide Signaling PathwayOxidantsOxidation-ReductionOxidative StressP2Y2 receptorPathologicPathway interactionsPharmaceutical PreparationsPhenotypePhosphorylationPhysiologicalPhysiologyPost-Translational Protein ProcessingPreparationProteinsPublic HealthPurinoceptorReactive Oxygen SpeciesRecombinantsResearchRoleSignal PathwaySignal TransductionTechnologyTestingTranscriptTransgenic MiceVascular DiseasesVascular EndotheliumWorkYeastsblood pressure controlcellular imagingcofactorendothelial dysfunctionexperimental studyhemodynamicsimaging approachin vivo Modelmortalitynew therapeutic targetnovelprogramsresponseshear stresstime usetranscriptome
项目摘要
The proposed studies will use new biosensors and novel chemogenetic approaches to identify the molecular
mechanisms whereby reactive oxygen species (ROS) regulate nitric oxide (NO) signaling pathways in the
vascular endothelium. The proposed studies build on recent work in which we used chemogenetics to develop
a new animal model of cardiomyopathy caused by oxidative stress. Here we plan to expand this chemogenetic
approach to develop a new experimental program to study endothelial dysfunction and hypertension.
Many studies have implicated oxidative stress in endothelial dysfunction and hypertension, yet the underlying
molecular mechanisms remain incompletely understood. Low levels of the stable ROS hydrogen peroxide (H2O2)
modulate NO-dependent physiological responses, while higher ROS levels are associated with hypertension.
The proposed experiments exploit recent advances in chemogenetic and biosensor technologies to identify the
mechanisms underlying the transition from physiological H2O2 signaling to the development of hypertension and
other vascular disease states associated with pathological oxidative stress. We will pursue multispectral imaging
experiments that will simultaneously analyze H2O2, NO and Ca2+ using highly selective and sensitive HyPer7,
geNOp, and GECO biosensors. These studies will establish the mechanisms whereby purinergic P2Y2 receptors
modulate H2O2-, Ca2+-, and NO-dependent endothelial responses that control blood pressure.
Hemodynamic shear stress leads to eNOS activation and to increases in endothelial ROS that can promote
both physiological as well as pathophysiological responses. We found that physiological laminar shear stress
preferentially increases H2O2 in the endothelial cell nucleus, while pathological oscillatory shear stress increases
H2O2 more in the cell cytosol. We used a chemogenetic approach to generate H2O2 in endothelial cells, using
novel recombinant constructs expressing a yeast D-amino acid oxidase (DAAO) that robustly produces H2O2.
The recombinant yeast DAAO is quiescent since vascular cells contain L- but not D-amino acids. H2O2 can be
generated by adding D-alanine to cells expressing recombinant DAAO. Our studies showed that H2O2 generated
in the endothelial cell nucleus activates Nrf2-modulated transcripts, whereas generation of H2O2 in the cytosol
principally increases NF-kB-dependent transcripts. These differential transcriptional responses establish a
causal role for H2O2 and provide a strong connection between chemogenetic approaches and endothelial
pathophysiology. Studying in vitro, ex vivo, and in vivo models, we propose to extend these studies from cultured
human endothelial cells (Aim 1) to the investigation of arterial preparations and transgenic mice expressing
DAAO in the endothelium (Aim 2). This experimental program may lead to the development of a new
“chemogenetic” animal model of hypertension. These studies will use powerful new cell imaging approaches to
test the hypothesis that perturbations in intracellular H2O2 metabolism modulate endothelial responses both in
the normal vasculature and in hypertension, and in other vascular disease states caused by oxidative stress.
拟议的研究将使用新的生物传感器和新的化学遗传学方法来识别分子
活性氧 (ROS) 调节一氧化氮 (NO) 信号通路的机制
血管内皮。拟议的研究建立在我们最近使用化学遗传学开发的工作的基础上
氧化应激引起的心肌病的新动物模型。在这里,我们计划扩展这种化学遗传学
方法开发一个新的实验计划来研究内皮功能障碍和高血压。
许多研究表明氧化应激与内皮功能障碍和高血压有关,但潜在的原因
分子机制仍不完全清楚。低水平的稳定 ROS 过氧化氢 (H2O2)
调节 NO 依赖性生理反应,而较高的 ROS 水平与高血压相关。
拟议的实验利用化学遗传学和生物传感器技术的最新进展来识别
从生理性 H2O2 信号转导到高血压发展的潜在机制
与病理性氧化应激相关的其他血管疾病状态。我们将追求多光谱成像
使用高选择性和灵敏的 HyPer7 同时分析 H2O2、NO 和 Ca2+ 的实验,
geNOp 和 GECO 生物传感器。这些研究将建立嘌呤能 P2Y2 受体的机制
调节 H2O2-、Ca2+- 和 NO 依赖性内皮反应来控制血压。
血流动力学剪切应力导致 eNOS 激活和内皮 ROS 增加,从而促进
生理反应和病理生理反应。我们发现生理层流剪应力
优先增加内皮细胞核中的 H2O2,同时病理性振荡剪切应力增加
H2O2较多存在于细胞质中。我们使用化学遗传学方法在内皮细胞中产生 H2O2,使用
表达酵母 D-氨基酸氧化酶 (DAAO) 的新型重组结构,可强力产生 H2O2。
重组酵母 DAAO 是静止的,因为维管细胞含有 L-氨基酸但不含 D-氨基酸。 H2O2 可以
通过向表达重组 DAAO 的细胞中添加 D-丙氨酸而产生。我们的研究表明 H2O2 产生
内皮细胞核中的 Nrf2 激活 Nrf2 调节的转录本,而细胞质中 H2O2 的产生
主要增加 NF-kB 依赖性转录本。这些差异转录反应建立了
H2O2 的因果作用并提供化学遗传学方法和内皮细胞之间的紧密联系
病理生理学。通过研究体外、离体和体内模型,我们建议将这些研究扩展到培养
人内皮细胞(目标 1)用于动脉制剂和转基因小鼠表达的研究
内皮细胞中的 DAAO(目标 2)。这个实验计划可能会导致新的开发
高血压的“化学遗传学”动物模型。这些研究将使用强大的新细胞成像方法
检验细胞内 H2O2 代谢的扰动调节内皮反应的假设
正常脉管系统和高血压以及氧化应激引起的其他血管疾病状态。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Thomas Michel其他文献
Thomas Michel的其他文献
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{{ truncateString('Thomas Michel', 18)}}的其他基金
Dynamic tissue-specific modulation of redox stress using chemogenetics
利用化学遗传学对氧化还原应激进行动态组织特异性调节
- 批准号:
10393690 - 财政年份:2021
- 资助金额:
$ 44.22万 - 项目类别:
Dynamic tissue-specific modulation of redox stress using chemogenetics
利用化学遗传学对氧化还原应激进行动态组织特异性调节
- 批准号:
10214064 - 财政年份:2021
- 资助金额:
$ 44.22万 - 项目类别:
Hydrogen peroxide in endothelial function and dysfunction
过氧化氢在内皮功能和功能障碍中的作用
- 批准号:
10543765 - 财政年份:2021
- 资助金额:
$ 44.22万 - 项目类别:
Chemogenetic approaches to define the roles of redox dysfunction in the cardiomyopathy of aging
化学遗传学方法确定氧化还原功能障碍在衰老心肌病中的作用
- 批准号:
9922852 - 财政年份:2019
- 资助金额:
$ 44.22万 - 项目类别:
REDOX REGULATION OF eNOS SIGNALING PATHWAYS IN VASCULAR ENDOTHELIUM
血管内皮细胞 eNOS 信号通路的氧化还原调节
- 批准号:
8250446 - 财政年份:2011
- 资助金额:
$ 44.22万 - 项目类别:
ANIMAL MODELS OF REDOX METABOLISM AND ARTERIAL DYSFUNCTION
氧化还原代谢和动脉功能障碍的动物模型
- 批准号:
8250450 - 财政年份:2011
- 资助金额:
$ 44.22万 - 项目类别:
REDOX REGULATION OF eNOS SIGNALING PATHWAYS IN VASCULAR ENDOTHELIUM
血管内皮细胞 eNOS 信号通路的氧化还原调节
- 批准号:
7975784 - 财政年份:2010
- 资助金额:
$ 44.22万 - 项目类别:
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