Mitochondrial DNA Mutations in Skeletal Muscles in Aging

描述（由申请人提供）：本授权是先前申请1 R 03 AG 024640 -01的修订版。这项拨款申请的总体目标是研究线粒体在衰老中的作用，特别是在小鼠骨骼肌中测试线粒体衰老理论。根据这一理论，体细胞线粒体DNA（mtDNA）突变导致电子转移缺陷，增加了破坏性活性氧（ROS）的产生，进而产生进一步的mtDNA突变。这种恶性循环可能导致线粒体功能受损，包括能量产生减少。随之而来的组织退化将导致各种老化表型。有间接证据支持这一理论，并且据报道，一些特定的mtDNA突变在衰老过程中在各种组织中积累。然而，对于衰老过程中线粒体DNA整体突变（所有类型的突变共存于同一细胞中，包括低丰度突变）积累的研究仍然缺乏全面的研究。也许更重要的是，与衰老相关的mtDNA突变的生理后果在很大程度上还不清楚。我们最近开发了一种新的方法，从小鼠骨骼肌mtDNA转移到体外细胞培养系统，并改进了方法来分离和鉴定mtDNA突变。结合已有的技术，我们将通过评估年轻、中年和老年小鼠骨骼肌中的个体mtDNA，研究衰老过程中骨骼肌中总体mtDNA突变的积累，并进一步分离和鉴定与年龄相关的mtDNA突变。该研究的成功将有助于我们深入了解衰老的分子机制以及与衰老相关的人类退行性疾病。

DESCRIPTION (provided by applicant): This grant is a revised version of previous application 1 R03 AG024640-01. The overall goal of this grant application is to examine the role of mitochondria in aging, and, in particular, to test the mitochondrial theory of aging in mouse skeletal muscles. According to this theory, somatic mitochondrial DNA (mtDNA) mutations cause defective electron transfer, increasing the generation of damaging reactive oxygen species (ROS), that, in turn, produce further mtDNA mutations. This vicious cycle presumably results in compromised mitochondrial function including decreased energy production. The ensuing tissue degeneration will lead to various aging phenotypes. There are circumstantial evidences to support this theory, and some specific mtDNA mutations have been reported to accumulate in various tissues during aging. However, there is still no comprehensive study on the overall mtDNA mutation (all kinds of mutations co-exist in the same cell including those with low abundance) accumulation during aging. Perhaps more importantly, the physiological consequences of the aging related mtDNA mutations are largely unclear. We recently developed a novel approach to transfer mtDNA from mouse skeletal muscle to an in vitro cell culture system, and improved methods to isolate and identify mtDNA mutations. Combined with the established technologies, we will investigate the accumulation of overall mtDNA mutations in skeletal muscle during aging by evaluating individual mtDNA from the muscle in young, middle-age and old mice, and further isolate and identify age-associated mtDNA mutations. The success of study will help us to gain insights of molecular mechanism of aging and aging-related human degenerative diseases.