EFFECTS OF SHORT-AND LONG-TERM FLEX-HET THERAPY ON MITOCHONDRIAL ELECTRON

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 我们最近发表了我们的观察结果，即领先的Flex-Het化合物SHetA2对线粒体密度、膜完整性、细胞色素C释放和ROS生成的快速影响，所有这些都表明了内在的细胞凋亡途径。最近，我们观察到Flex-hets是线粒体电子传递链复合体I的有效的、非竞争性的抑制剂，也是复合体II的抑制剂。由于目前已知的复合体I和II的抑制剂很少，在本建议的第一部分中，我们将进一步研究Flex-Het药物对琥珀酸脱氢酶和完全依赖泛醌的琥珀酸：泛醌氧化还原酶活性的影响。然而，线粒体靶向药物可能是一把双刃剑，ROS本身对细胞具有高度毒性，线粒体可以逐渐积累氧化损伤，导致线粒体崩溃和细胞死亡。在这项建议的第二部分，我们建议评估长期Flex-Het治疗后心脏和肝脏细胞线粒体损伤的可能性。在一项正在进行的结肠癌长期化学预防项目评估ShetA2中，将在细胞、线粒体和亚线粒体颗粒中检测线粒体功能受损的标志，如细胞ATP降低、线粒体谷胱甘肽降低和复合体I活性降低。这些线粒体和亚线粒体颗粒是从动物的冷冻心脏和肝脏组织中分离出来的，这些组织是在正在进行的结肠癌长期化学预防项目中评估ShetA2的。

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. We have recently published our observation of the rapid effects of the lead Flex-Het compound, SHetA2, on mitochondrial density, membrane integrity, Cytochrome C release and ROS generation, all indicative of the intrinsic apoptosis pathway. More recently, we have observed that Flex-Hets are potent, uncompetitive inhibitors of mitochondrial electron transport chain complex I and also inhibitors of complex II. Since few inhibitors of both complex I and II are know at present, in the first part of this proposal we will further examine the effect of Flex-Het drugs on succinate dehydrogenase and full ubiquinone dependent succinate:ubiquinone oxidoreductase activity. However mitochondria targeting drugs may be a two-edged sword, ROS themselves are highly toxic to cells and mitochondria can gradually accumUlate oxidative damage leading to mitochondrial collapse and cell death. In the second part of this proposal we propose to assess the possibility of mitochondrial injury in heart and liver cells following long-term Flex-Het treatment. Markers of impaired mitochondrial function such as decreased cellular ATP, decreased mitochondrial glutathione and decreased complex I activity will be measured in both cells, mitochondria and submitochondrial particles isolated from snap-frozen heart and liver tissue obtained from animals sacrificed following necropsy in an ongoing long-term colon cancer chemoprevention project evaluating ShetA2.