Oxidative Stress and G6PDH Expression

氧化应激和 G6PDH 表达

• 批准号: 0136127
• 负责人: Susan Stapleton
• 金额: $ 29.06万
• 依托单位: Western Michigan University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-06-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0136127&HistoricalAwards=false
• 关键词: Oxidative; Stress; G6PDH; Expression

Reactive oxygen species (ROS) are produced inside the cell during normal oxidative metabolism. Tight control of this process is crucial for numerous biological events to occur, including enzyme activation, cell cycle regulation and programmed cell death. Abnormal production of ROS can lead to an imbalance in redox homeostasis known as oxidative stress. Often times exposure to environmental agents, including, UV irradiation, ozone, herbicides, pesticides and metals can produce additional ROS that lead to this imbalance.Upon exposure to these agents, this elevated level of ROS can cause damage to proteins, nucleic acids and lipids, as well as, alter physiological responses such as signal transduction and gene expression. Long term consequences of oxidative stress have also been associated with the pathogenesis of a variety of toxicities and diseases. Under some circumstances, cells can utilize an antioxidant defense system in order to cope with the excess ROS produced during oxidative stress. However, the mechanism by which some key components in the antioxidant defense system work is not completely known. For example, NADPH is a critical biological cofactor that helps to maintain a balance between the prooxidant and antioxidant status of the cell. The pentose phosphate pathway through the activities of glucose-6-phosphate dehydrogenase (G6PDH) and 6-phosphogluconate dehydrogenase generates most of the NADPH needed by the cell. G6PDH is the key rate-limiting enzyme in this pathway and thus not only regulates the flow of carbon through this pathway, but also the production of NADPH. Since NADPH is necessary for maintaining the cell's redox balance and G6PDH is important for its production then it seems reasonable to conclude that the redox state of the cell would influence the expression of this important enzyme. Indeed, studies from this laboratory and others have suggested that expression of the gene for G6PDH is regulated by the oxidative status of the cell. However, the mechanism by which this regulation occurs is unknown. Therefore, this research is aimed at testing the hypothesis that the environmental contaminant cadmium, which influences the oxidative state of a cell by decreasing levels of glutathione, regulates the expression of G6PDH through a redox sensitive transcriptional event. Cadmium is generating a great deal of environmental concern due to its accumulation from such industrial practices as production of plastics, materials, batteries, tobacco, electroplating, welding and smelting. Organisms are easily exposed to cadmium due to its relatively high vapor pressure, which accounts for its high solubility in water. The studies will be done in primary rat hepatocytes in culture. The liver is the major target organ for acute inorganic Cd exposure. Additionally, hepatocytes are attractive to use as a model since the metabolic responses to hormones, nutrients or environmental agents consistently mimic those observed in vivo.These studies would represent the first of this kind and add significantly to our understanding of the role of the oxidative state of the cell on the expression of enzymes involved in major biochemical pathways such as the pentose phosphate pathway. These studies would also add valuable information about how a cell protects itself from damage induced from environmental agents such as metals. A team learning environment will facilitate the completion of the work by the PI and students.

活性氧（ROS）是在正常的氧化代谢过程中在细胞内产生的。对这一过程的严格控制对于许多生物事件的发生至关重要，包括酶激活、细胞周期调节和程序性细胞死亡。ROS的异常产生可导致氧化还原稳态的失衡，称为氧化应激。 暴露于环境因素（包括紫外线照射、臭氧、除草剂、杀虫剂和金属）通常会产生额外的ROS，导致这种失衡。暴露于这些因素后，ROS水平升高会导致蛋白质、核酸和脂质的损伤，以及改变信号转导和基因表达等生理反应。 氧化应激的长期后果也与多种毒性和疾病的发病机制有关。 在某些情况下，细胞可以利用抗氧化防御系统来科普氧化应激过程中产生的过量ROS。 然而，抗氧化防御系统中的一些关键成分的工作机制还不完全清楚。 例如，NADPH是一种关键的生物辅因子，有助于维持细胞的促氧化剂和抗氧化剂状态之间的平衡。 戊糖磷酸途径通过葡萄糖-6-磷酸脱氢酶（G6 PDH）和6-磷酸葡萄糖酸脱氢酶的活性产生细胞所需的大部分NADPH。G6 PDH是该途径中的关键限速酶，因此不仅调节通过该途径的碳流，而且还调节NADPH的产生。 由于NADPH是维持细胞氧化还原平衡所必需的，而G6 PDH对其产生很重要，因此似乎可以合理地得出结论，细胞的氧化还原状态会影响这种重要酶的表达。 事实上，该实验室和其他实验室的研究表明，G6 PDH基因的表达受细胞氧化状态的调节。然而，这种调节发生的机制尚不清楚。 因此，本研究的目的是测试的假设，即环境污染物镉，影响细胞的氧化状态，通过降低谷胱甘肽水平，调节G6 PDH的表达，通过氧化还原敏感的转录事件。 由于镉在塑料、材料、电池、烟草、电镀、焊接和冶炼等工业生产中的积累，镉引起了大量的环境问题。 由于镉的蒸气压相对较高，生物体很容易暴露于镉，这是其在水中的高溶解度的原因。 将在培养的原代大鼠肝细胞中进行研究。 肝脏是急性无机镉暴露的主要靶器官。 此外，肝细胞是有吸引力的作为一个模型，因为激素，营养物质或环境因素的代谢反应始终模仿那些在vivo.These研究中观察到的将代表这类的第一个，并添加显着增加我们的理解的作用，细胞的氧化状态对酶的表达参与主要的生化途径，如戊糖磷酸途径。 这些研究还将增加有关细胞如何保护自身免受金属等环境因子损伤的宝贵信息。 团队学习环境将促进PI和学生完成工作。