Mechanism of Glutathione Conjugate Dependent Toxicity

谷胱甘肽缀合物依赖性毒性机制

• 批准号: 6805315
• 负责人: Arthur Joseph Cooper
• 金额: $ 35.55万
• 依托单位: WINIFRED MASTERSON BURKE MED RES INST
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6805315
• 关键词: SDS polyacrylamide gel electrophoresis; alkenes; bioenergetics; brain; chemical conjugate; cytotoxicity; environmental exposure; environmental toxicology; enzyme activity; fluorocarbon polymers; glutathione; halogenation; kidney; laboratory rat; liquid chromatography mass spectrometry; liver; membrane potentials; mitochondria; toxin metabolism; transaminases

DESCRIPTION (provided by applicant): A significant portion of the US population is exposed to toxic, halogenated alkenes (e.g., trichloroethylene, tetrachloroethylene and tetrafluoroethylene) in the workplace and environment. These compounds are metabolized in part to halogenated cysteine S-conjugates, which are thought to be the major toxicants. S-(1,1,2,2-Tetrafluoroethyl)-L-cysteine (TFEC), the cysteine S-conjugate of tetrafluoroethylene, is chosen here as a representative toxic, halogenated cysteine S-conjugate. Toxic, halogenated cysteine S-conjugates are converted to pyruvate, ammonia and a reactive (thioacylating) fragment by cysteine S-conjugate ?-lyases. In vivo, the kidney and, to some extent, liver and brain, are susceptible. Previously, we showed that (i) a high-Mr cysteine S-conjugate ?-lyase in rat kidney co-purifies with mitochondrial HSP70 and protein disulfide isomerase, and contains mitochondrial aspartate aminotransferase (mitAspAT) (ii) several aminotransferases [mitochondrial branched-chain aminotransferase (BCATm), cytosolic branched-chain aminotransferase, alanine-glyoxylate aminotransferase II, mitAspAT] possess cysteine S-conjugate ?-lyase activity, but are themselves inactivated during turnover (syncatalytic inactivation), and (iii) exposure of PC12 cells and astrocytes in culture to TFEC causes selective loss of key mitochondrial enzymes of energy metabolism, including mitAspAT and ?-ketoglutarate dehydrogenase complex (KGDHC). Others have shown that KGDHC and branched-chain keto acid dehydrogenase complex (BCKAD) are targets of TFEC in rat kidney cells in vivo, and that halogenated cysteine S-conjugates are metabolic poisons of isolated kidney and liver mitochondria. The PI and coworkers have suggested that KGDHC and BCKAD are sensitive to inactivation due to toxicant channeling involving mitAspAT and BCATm, respectively. The overall goal of the present proposal is to determine the mechanism by which mitochondrial metabolism is poisoned by TFEC/TFEC thioacylating fragment. Accordingly, our aims are to determine: a) the effects of TFEC on respiration, Ca 2+ homeostasis, membrane potential and swelling in isolated rat liver, brain and kidney mitochondria, and correlate such pathological changes with loss of key mitochondrial enzyme activities, b) the mechanism whereby mitAspAT and BCATm are syncatalytically inactivated by TFEC, and c) the mechanism of toxicant (TFEC thioacylating fragment) transfer (channeling) from mitAspAT to KGDHC and from BCATm to BCKAD. The findings should elucidate the link between exposure to certain halogenated xenobiotics and impaired energy metabolism, and may suggest a means to minimize the toxic effects in heavily exposed individuals.

描述（由申请人提供）：在工作场所和环境中，大部分美国人口暴露于有毒，卤代烷烃（例如三氯乙烯，四氯乙烯和四氟乙烯）。这些化合物的一部分是用卤代半胱氨酸S-偶联物代谢的，后者被认为是主要有毒物质。 S-（1,1,2,2-二氟乙基）-L-半胱氨酸（TFEC）是四氟乙烯的半胱氨酸S-偶联物，在这里被选为代表性的有毒，卤代的卤代卤代半胱氨酸S-偶联物。有毒，卤代半胱氨酸S-偶联物被转化为丙酮酸，氨和通过半胱氨酸S-偶联物的反应性（硫代）片段？在体内，肾脏和肝脏和大脑在某种程度上很容易受到影响。以前，我们表明（i）大鼠肾脏中的高MR半胱氨酸s-偶联酶与线粒体HSP70和蛋白二硫化物异构酶，并包含线粒体天冬氨酸氨基植物氨基转移酶（mitaspat）（mitaspat）（MITASPAT）（MITASPAT）（II）分支氨基酸酶[minotial-氨基酸酶）[MITASPATS（II） （BCATM），胞质分支链氨基转移酶，丙氨酸 - 甘氧基氨基转移酶II，mitaspat]具有半胱氨酸S-偶联物的活性？ - 但在失流期间灭活时是否会灭活（pcatalytive灭绝）和（III）的composities和（III），以及（iii II II II II II II II II），以供PC12的供应量表，以供培养物，以供PC12的供应量表能量代谢的线粒体酶，包括mitaspat和？ - 酮戊二酸脱氢酶复合物（KGDHC）。其他人则表明，kGDHC和分支链酮酸脱氢酶复合物（BCKAD）是体内大鼠肾细胞中TFEC的靶标，而卤代半胱氨酸S-偶联物是分离的肾脏和肝脏软骨病的代谢性毒药。 PI及其同事建议KGDHC和BCKAD对分别涉及Mitaspat和BCATM的毒物通道引起的失活敏感。本提案的总体目标是确定线粒体代谢被TFEC/TFEC硫酰基片段中毒的机制。 Accordingly, our aims are to determine: a) the effects of TFEC on respiration, Ca 2+ homeostasis, membrane potential and swelling in isolated rat liver, brain and kidney mitochondria, and correlate such pathological changes with loss of key mitochondrial enzyme activities, b) the mechanism whereby mitAspAT and BCATm are syncatalytically inactivated by TFEC, and c) the mechanism of有毒物质（TFEC硫酰化片段）从Mitaspat到KGDHC以及从​​BCATM到BCKAD的转移。这些发现应阐明暴露于某些卤代异种生物学和能量代谢受损之间的联系，并可能提出一种方法来最大程度地减少暴露的个体的毒性作用。