Claflin Dennis R, Jackson Malcolm J, Brooks Susan V
Department of Biomedical Engineering, University of Michigan Ann Arbor, MI, USA ; Department of Surgery, Section of Plastic Surgery, University of Michigan Ann Arbor, MI, USA.
Department of Musculoskeletal Biology, Institute of Ageing and Chronic Disease, University of Liverpool Liverpool, UK.
Front Physiol. 2015 Feb 4;6:21. doi: 10.3389/fphys.2015.00021. eCollection 2015.
Compromised mitochondrial respiratory function is associated with advancing age. Damage due to an increase in reactive oxygen species (ROS) with age is thought to contribute to the mitochondrial deficits. The coenzyme nicotinamide adenine dinucleotide in its reduced (NADH) and oxidized (NAD(+)) forms plays an essential role in the cyclic sequence of reactions that result in the regeneration of ATP by oxidative phosphorylation in mitochondria. Monitoring mitochondrial NADH/NAD(+) redox status during recovery from an episode of high energy demand thus allows assessment of mitochondrial function. NADH fluoresces when excited with ultraviolet light in the UV-A band and NAD(+) does not, allowing NADH/NAD(+) to be monitored in real time using fluorescence microscopy. Our goal was to assess mitochondrial function by monitoring the NADH fluorescence response following a brief period of high energy demand in muscle from adult and old wild-type mice. This was accomplished by isolating whole lumbrical muscles from the hind paws of 7- and 28-month-old mice and making simultaneous measurements of force and NADH fluorescence responses during and after a 5 s maximum isometric contraction. All muscles exhibited fluorescence oscillations that were qualitatively similar and consisted of a brief transient increase followed by a longer transient period of reduced fluorescence and, finally, an increase that included an overshoot before recovering to resting level. Compared with the adult mice, muscles from the 28 mo mice exhibited a delayed peak during the first fluorescence transient and an attenuated recovery following the second transient. These findings indicate an impaired mitochondrial capacity to maintain NADH/NAD(+) redox homeostasis during contractile activity in skeletal muscles of old mice.
线粒体呼吸功能受损与衰老进程相关。随着年龄增长，活性氧（ROS）增加所导致的损伤被认为是造成线粒体功能缺陷的原因。辅酶烟酰胺腺嘌呤二核苷酸的还原形式（NADH）和氧化形式（NAD(+)）在线粒体中通过氧化磷酸化导致ATP再生的一系列循环反应中起着至关重要的作用。因此，在从高能量需求状态恢复过程中监测线粒体NADH/NAD(+)氧化还原状态，可以评估线粒体功能。当用UV-A波段的紫外光激发时，NADH会发出荧光，而NAD(+)则不会，这使得可以使用荧光显微镜实时监测NADH/NAD(+)。我们的目标是通过监测成年和老年野生型小鼠肌肉在短暂高能量需求后的NADH荧光反应来评估线粒体功能。这是通过从7个月和28个月大的小鼠后爪分离完整的蚓状肌，并在5秒最大等长收缩期间及之后同时测量力量和NADH荧光反应来实现的。所有肌肉都表现出定性相似的荧光振荡，包括短暂的瞬时增加，随后是较长时间的荧光降低的瞬时阶段，最后是增加，包括在恢复到静息水平之前的一个峰值。与成年小鼠相比，28月龄小鼠的肌肉在第一次荧光瞬变期间峰值延迟，在第二次瞬变后恢复减弱。这些发现表明，老年小鼠骨骼肌在收缩活动期间，线粒体维持NADH/NAD(+)氧化还原稳态的能力受损。
