CARDIOMYOCYTE AND VASCULAR ENDOTHELIAL CELL SIGNALING BY CHOLESTEROL OZONATION

胆固醇臭氧化的心肌细胞和血管内皮细胞信号转导

• 批准号: 7609943
• 负责人: RAO M UPPU
• 金额: $ 12.5万
• 依托单位: LOUISIANA STATE UNIV A&M COL BATON ROUGE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7609943
• 关键词: Acetylcysteine; Alzheimer' s Disease; Antioxidants; Apoptosis; Atherosclerosis; Cardiac Myocytes; Caspase; Cell Death; Cessation of life; Cholesterol; Computer Retrieval of Information on Scientific Projects Database; Dementia; Disruption; Funding; Generations; Glutathione; Grant; Human; Institution; Ketones; Lewy Bodies; Ligands; Membrane Potentials; Mitochondria; Oxidative Stress; Pathway interactions; Permeability; Play; Production; Reactive Oxygen Species; Reporting; Research; Research Personnel; Resources; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Transduction; Source; Transcriptional Activation; Trolox; United States National Institutes of Health; Up-Regulation; Vascular Endothelial Cell; alanylaspartic acid; base; caspase-3; caspase-8; caspase-9; cytochrome c; cytotoxic; in vivo; inhibitor/antagonist; prevent; receptor

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Cholesterol secoaldehyde (3¿-hydroxy-5-oxo-5,6-secocholestan-6-al or ChSeco), a major ozonation product of cholesterol is known to occur in vivo in humans and is associated with atherosclerosis, Alzheimers disease and Lewy body dementia. In the last annual project we reported that ChSeco induced cell death in H9c2 cardiomyocytes that is typical of apoptosis. In this study period, we investigated the downstream signaling mechanisms that culminate in apoptosis. ChSeco caused activation of caspase dependent mechanism with no changes in Bcl-2 and Bax expressions. Quantitative RT-PCR analysis revealed that ChSeco activates mitochondrial (intrinsic) and death receptor (extrinsic) pathways. The mitochondrial pathway was shown to be of rapid onset and it involved disruption of transmembrane potential (¿¿m), formation of permeability transition pore (release of cytochrome c), activation of caspase 9, and up-regulation of Apaf1. The extrinsic pathway involved up-regulation of death receptor and its ligand. The significance of this pathway was also evident based on activation of caspase 8. It was found that a combined activation of intrinsic and extrinsic pathways resulted in activation of effector caspases 3 and 7. Inhibitors of caspases 8 and 9 decreased the activity of caspase 3, and the cell death was partially prevented by a pancaspase inhibitor, Z-Val-Ala-Asp-fluoromethyl ketone. ChSeco caused a rapid increase in the generation of reactive oxygen species (ROS) associated with a decrease in intracellular glutathione. Antioxidants such as N-acetyl-L-cysteine and Trolox significantly reversed ChSeco-apoptosis by inhibiting all the components of either pathway indicating that oxidative stress lies upstream to these pathways and plays a primary role triggering cell death. This shows that ROS production plays a critical role in initiating apoptosis by pathways involving both mitochondria and death receptor in H9c2 cardiomyocytes and that antioxidants play an effective role in combating the cytotoxic effects of ChSeco. Further studies are underway to elucidate the effect of ChSeco in vascular endothelial cells. Also, the effect of other cholesterol ozonation products will be studied in both cardiomyocytes and vascular endothelial cells.

该副本是使用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这是调查员的机构。 已知胆固醇胆固醇（胆固醇3--羟基-5-oxo-5,6-秒六甲醛或Chseco）是胆固醇的主要臭氧化产物，它在人类的体内发生，并且与动脉粥样硬化，阿尔茨海默氏病，阿尔茨海默氏病和lewy的身体凹陷有关。在上一年的年度项目中，我们报告说，Chseco诱导了典型凋亡的H9C2心肌细胞中的细胞死亡。在本研究期间，我们研究了最终凋亡的下游信号传导机制。 CHSECO引起caspase依赖机制的激活，而Bcl-2和Bax表达没有变化。定量RT-PCR分析表明，CHSECO激活了线粒体（内在）和死亡受体（外部）途径。线粒体途径显示出快速发作，并且涉及跨膜电位（``m）的破坏，渗透性过渡孔的形成（细胞色素C的释放），caspase 9的激活和APAF1的上调。外部途径涉及死亡接收器及其配体的上调。该途径的重要性也是基于caspase 8的激活的证据。发现固有和外在途径的组合激活导致效应胱天蛋白酶3和7的激活。caspase 8和9的抑制剂8和9降低了caspases 3的活性，pancaspase抑制剂抑制了calla-apasp抑制细胞死亡的活性。 Chseco导致与细胞内谷胱甘肽降低相关的活性氧（ROS）的产生迅速增加。抗氧化剂（例如N-乙酰L-半胱氨酸和Trolox）通过抑制任何任一途径的所有成分，表明氧化应激位于这些途径上游，并起着触发细胞死亡的主要作用，从而显着逆转了Chseco凋亡。这表明ROS产生在H9C2心肌细胞中涉及线粒体和死亡受体的途径引发凋亡中起着至关重要的作用，并且抗氧化剂在梳理Chseco的细胞毒性作用方面起着有效的作用。正在进行进一步的研究，以阐明Chseco在血管内皮细胞中的影响。同样，其他胆固醇臭氧化产物的作用将在心肌细胞和血管内皮细胞中研究。