Smith D R, Stone D, Darley-Usmar V M
Enzymology Group, Wellcome Research Laboratories, Beckenham, Kent, UK.
Free Radic Res. 1996 Mar;24(3):159-66. doi: 10.3109/10715769609088013.
An increase in mitochondrial matrix free calcium has been shown to occur during oxygen and substrate deprivation of the perfused heart which predisposes to calcium overload and inhibition of mitochondrial function on reoxygenation. In the current study we have assessed the effect of substrate free hypoxia on mitochondrial oxygen consumption and ATP synthesis in isolated rat cardiomyocytes. Myocytes were subjected to 40 min of substrate-free hypoxia and the oxygen consumption measured together with the effects on ATP and PCr synthesis. After hypoxia myocytes showed a fall in ATP to 10% of the control value. Within 5 sec of reoxygenation the ATP level recovered to a new steady state level of 30% of the original value. The rate of oxygen consumption of hypoxic/reoxygenated cells was 3-4 fold higher than that of cells maintained under normoxic controls but in the presence of oligomycin the difference was only 1.5-fold higher, indicating a greater requirement for mitochondrial synthesis of ATP following reoxygenation. Reoxygenation in the absence of extracellular Ca2+ resulted in a lower rate of oxygen consumption (50% of the rate measured in the presence of 1 mM-Ca2+) but did not affect the steady state concentration of ATP attained 5 min after reoxygenation. These results support the idea that the increased O2 consumption of myocytes following hypoxia/reoxygenation is due to an increased demand for ATP synthesis by mitochondria and is a response to the NA+ and Ca2+ loading of the cells which occurs under these conditions. This increased demand is likely to result in a greater generation of free radicals such as superoxide by the respiratory chain which could impair cellular function over the long term.
灌注心脏在缺氧和缺乏底物时，线粒体基质游离钙会增加，这会导致钙超载，并在再灌注时抑制线粒体功能。在本研究中，我们评估了无底物缺氧对分离的大鼠心肌细胞线粒体氧消耗和ATP合成的影响。将心肌细胞置于无底物缺氧环境40分钟，测量其氧消耗以及对ATP和磷酸肌酸（PCr）合成的影响。缺氧后，心肌细胞的ATP水平降至对照值的10%。再灌注后5秒内，ATP水平恢复到原始值30%的新稳态水平。缺氧/再灌注细胞的氧消耗率比常氧对照下的细胞高3 - 4倍，但在存在寡霉素的情况下，差异仅为高1.5倍，这表明再灌注后线粒体合成ATP的需求更大。在无细胞外Ca2+的情况下进行再灌注，导致氧消耗率降低（为存在1 mM Ca2+时测量速率的50%），但不影响再灌注5分钟后达到的ATP稳态浓度。这些结果支持以下观点：缺氧/再灌注后心肌细胞耗氧量增加是由于线粒体对ATP合成的需求增加，并且是对这些条件下细胞内Na+和Ca2+负荷增加的一种反应。这种增加的需求可能会导致呼吸链产生更多的自由基，如超氧阴离子，长期来看可能会损害细胞功能。
