Department of Pharmacology and Physiology, UMDNJ-New Jersey Medical School, Newark, NJ 07103, USA.
Circ Res. 2010 Mar 5;106(4):730-8. doi: 10.1161/CIRCRESAHA.109.206631. Epub 2010 Jan 7.
Uncoupling protein (UCP)2 is a mitochondrial inner membrane protein that is expressed in mammalian myocardium under normal conditions and upregulated in pathological states such as heart failure. UCP2 is thought to protect cardiomyocytes against oxidative stress by dissipating the mitochondrial proton gradient and mitochondrial membrane potential (DeltaPsi(m)), thereby reducing mitochondrial reactive oxygen species generation. However, in apparent conflict with its uncoupling role, UCP2 has also been proposed to be essential for mitochondrial Ca(2+) uptake, which could have a protective action by stimulating mitochondrial ATP production.
The goal of this study was to better understand the role of myocardial UCP2 by examining the effects of UCP2 on bioenergetics, Ca(2+) homeostasis, and excitation-contraction coupling in neonatal cardiomyocytes.
Adenoviral-mediated expression of UCP2 caused a mild depression of DeltaPsi(m) and increased the basal rate of oxygen consumption but did not affect total cellular ATP levels. Mitochondrial Ca(2+) uptake was examined in permeabilized cells loaded with the mitochondria-selective Ca(2+) probe, rhod-2. UCP2 overexpression markedly inhibited mitochondrial Ca(2+) uptake. Pretreatment with the UCP2-specific inhibitor genipin largely reversed the effects UCP2 expression on mitochondrial Ca(2+) handling, bioenergetics, and oxygen utilization. Electrically evoked cytosolic Ca(2+) transients and spontaneous cytosolic Ca(2+) sparks were examined using fluo-based probes and confocal microscopy in line scan mode. UCP2 overexpression significantly prolonged the decay phase of Ca(2+) transients in electrically paced cells, increased Ca(2+) spark activity and increased the probability that Ca(2+) sparks propagated into Ca(2+) waves. This dysregulation results from a loss of the ability of mitochondria to suppress local Ca(2+)-induced Ca(2+) release activity of the sarcoplasmic reticulum.
Increases in UCP2 expression that lower DeltaPsi(m) and contribute to protection against oxidative stress, also have deleterious effects on beat-to-beat Ca(2+) handling and excitation-contraction coupling, which may contribute to the progression of heart disease.
解偶联蛋白 (UCP)2 是一种线粒体内膜蛋白，在正常情况下在哺乳动物心肌中表达，并在心力衰竭等病理状态下上调。UCP2 被认为通过耗散线粒体质子梯度和线粒体膜电位 (DeltaPsi(m)) 来保护心肌细胞免受氧化应激，从而减少线粒体活性氧的产生。然而，与解偶联作用明显矛盾的是，UCP2 也被认为对线粒体 Ca(2+)摄取至关重要，通过刺激线粒体 ATP 产生，这种摄取可能具有保护作用。
本研究的目的是通过研究 UCP2 对新生心肌细胞生物能学、Ca(2+)稳态和兴奋-收缩偶联的影响，更好地了解心肌 UCP2 的作用。
腺病毒介导的 UCP2 表达导致 DeltaPsi(m) 轻度降低并增加基础耗氧率，但不影响总细胞 ATP 水平。在用线粒体选择性 Ca(2+)探针 rhod-2 负载的通透细胞中检查线粒体 Ca(2+)摄取。UCP2 过表达显着抑制线粒体 Ca(2+)摄取。用 UCP2 特异性抑制剂 genipin 预处理在很大程度上逆转了 UCP2 表达对线粒体 Ca(2+)处理、生物能学和氧利用的影响。使用基于荧光的探针和共聚焦显微镜在线扫描模式检查电诱发的胞质 Ca(2+)瞬变和自发的胞质 Ca(2+)火花。UCP2 过表达显着延长电刺激细胞中 Ca(2+) 瞬变的衰减相，增加 Ca(2+) 火花活性并增加 Ca(2+)火花传播为 Ca(2+)波的概率。这种失调是由于线粒体丧失了抑制肌浆网局部 Ca(2+)-诱导 Ca(2+)释放活性的能力所致。
增加 UCP2 表达会降低 DeltaPsi(m)，有助于抵抗氧化应激，但也会对心脏跳动的 Ca(2+) 处理和兴奋-收缩偶联产生有害影响，这可能导致心脏病的进展。
