Nanoelectrode arrays for study of the molecular mechanisms, triggers, and inhibit

用于研究分子机制、触发和抑制的纳米电极阵列

• 批准号: 7778178
• 负责人: PETER J. BURKE
• 金额: $ 24.32万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-15 至 2013-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7778178
• 关键词: Action Potentials; Address; Adenine Nucleotides; Apoptosis; Apoptosis Inhibitor; Apoptosis Regulator; Apoptotic; Binding; Biochemical; Caliber; Cancer Biology; Carbon Nanotubes; Cell Death; Cells; Charge; Cholesterol; Cytosol; Defect; Diamide; Down-Regulation; Electrodes; Functional disorder; Generations; Genes; Glucose; Heme; Hepatocyte; Hexokinase 2; Individual; Induction of Apoptosis; Infusion procedures; Inner mitochondrial membrane; Ions; Kinetics; Lipids; Maintenance; Measures; Membrane; Membrane Potentials; Metabolism; Metals; Microfluidics; Mitochondria; Mitochondrial DNA; Mitochondrial Proteins; Mitochondrial Proton-Translocating ATPases; Mitochondrial Swelling; Molecular; Monitor; Mutation; Neurons; Oxidative Stress; Oxygen; Peroxides; Phase; Phospholipids; Potential Energy; Primary carcinoma of the liver cells; Process; Production; Protein Family; Protein Isoforms; Regulation; Repression; Resolution; Signal Transduction; Signal Transduction Pathway; Source; Surface; Swelling; System; Technology; Testing; Variant; Vesicle; aerobic glycolysis; cancer cell; cell killing; complex IV; coproporphyrinogen III; cytochrome c oxidase; falls; functional status; high throughput technology; inhibitor/antagonist; macromolecule; miniaturize; mitochondrial dysfunction; mitochondrial membrane; mitochondrial permeability transition pore; nano; nanowire; oligomycin sensitivity-conferring protein; response; seal; sensor; single walled carbon nanotube; theories; tumorigenesis; uptake

DESCRIPTION (provided by applicant): Mitochondria are the central regulator of apoptosis, a process initiated by the activation of the mitochondrial permeability transition pore (mtPTP), an aggregate of several mitochondrial proteins. When this pore opens, the critical membrane polarization of the mitochondrial inner membrane disappears and ions equilibrate between the matrix and cytosol resulting in mitochondrial swelling. This leads to release of the contents of the mitochondrial intermembrane space into the cell cytosol, including a number of cell death promoting factors killing the cell. The mtPTP can be activated by uptake of excessive Ca++; increased oxidative stress; decreased mitochondrial membrane potential, and reduced ADP and ATP. It is generally agreed upon that repression of apoptosis is one of the fundamental steps in tumorigenesis. Cancer cells acquire unresponsiveness to apoptosis facilitating signals, thus enabling uncontrolled proliferation. For this reason, the induction of apoptosis is one of the modes of actions of chemotherapeutic compounds. In order to allow further high throughput studies of the biochemical facilitators and inhibitors, of apoptosis, and to determine if changes in individual mitochondrial membrane potential P are important to cellular metabolism, we need to develop a system to monitor P in individual mitochondria. To accomplish this objective, we propose to extend studies that have monitored the action potentials in neurons using an array of parallel electrodes to which the mitochondria are adhered. Our thesis is that a nanoelectrode technology can be developed to capacitively measure membrane potential across the mitochondrial inner membrane phospholipid bilayer without actually penetrating the membrane. We propose to develop nano-electrical transduction sensor arrays with sufficiently high spatial and temporal resolution to monitor the charge changes on the surface of a mitochondrion sized lipid vesicle and the individual mitoplast. With this technology, we will then interrogate the regulation of P in normal and cancer cells. Several key features on mitochondrial metabolism are now recognized as important to the alteration of cancer cell mitochondrial function: changes in the Akt signal transduction pathway, induction of hexokinase II, alteration an adenine nucleotide translocator (ANT) isoform expression, down regulation of the SOC2 cytochrome c oxidase (complex IV, COX) assembly factor, mutation in mitochondrial DNA (mtDNA) genes, and modulation of the mitochondrial permeability transition pore (mtPTP) and its interaction with the pro- and anti- apoptotic Bcl2 family proteins. While all of these are important factors in the alteration of cancer cell metabolism, they still fall short of explaining the near universal alterations in mitochondrial function observed in cancer cells. A high throughput technology to monitor P in mitochondria will allow further studies of these issues in cancer biology.

描述（由申请人提供）：线粒体是细胞凋亡的中心调节因子，细胞凋亡是一个由线粒体通透性转换孔（mtPTP）（几种线粒体蛋白质的聚集体）激活引发的过程。当该孔打开时，线粒体内膜的临界膜极化消失，并且离子在基质和胞质溶胶之间平衡，导致线粒体肿胀。这导致线粒体膜间隙的内容物释放到细胞胞质溶胶中，包括许多杀死细胞的细胞死亡促进因子。线粒体PTP可通过摄取过量的Ca++、增加的氧化应激、降低的线粒体膜电位以及减少的ADP和ATP来激活。细胞凋亡的抑制是肿瘤发生的基本步骤之一。癌细胞获得对细胞凋亡促进信号的无反应性，从而使不受控制的增殖成为可能。因此，诱导细胞凋亡是化疗化合物的作用模式之一。 为了使进一步的高通量研究的生化促进剂和抑制剂，细胞凋亡，并确定如果在个别线粒体膜电位P的变化是重要的细胞代谢，我们需要开发一个系统来监测P在个别线粒体。为了实现这一目标，我们建议扩展研究，监测神经元中的动作电位，使用一个阵列的平行电极的线粒体粘附。我们的论文是，纳米电极技术可以开发电容测量跨线粒体内膜磷脂双分子层的膜电位，而无需实际穿透膜。我们建议开发具有足够高的空间和时间分辨率的纳米电转导传感器阵列，以监测的表面上的电荷变化的脂质囊泡和单个的有丝分裂体。有了这项技术，我们将询问正常和癌细胞中P的调节。 线粒体代谢的几个关键特征现在被认为对癌细胞线粒体功能的改变很重要：Akt信号转导通路的变化，己糖激酶II的诱导，腺嘌呤核苷酸转运体（ANT）亚型表达的改变，SOC 2细胞色素c氧化酶的下调（复合物IV，考克斯）装配因子，线粒体DNA（mtDNA）基因突变，以及线粒体通透性转换孔（mtPTP）的调节及其与促凋亡和抗凋亡Bcl 2家族蛋白的相互作用。虽然所有这些都是改变癌细胞代谢的重要因素，但它们仍然无法解释在癌细胞中观察到的线粒体功能的几乎普遍改变。一种高通量的技术来监测P在线粒体中将允许进一步研究这些问题在癌症生物学。