Malis C D, Bonventre J V
J Biol Chem. 1986 Oct 25;261(30):14201-8.
With a variety of forms of ischemic and toxic tissue injury, cellular accumulation of Ca2+ and generation of oxygen free radicals may have adverse effects upon cellular and, in particular, mitochondrial membranes. Damage to mitochondria, resulting in impaired ATP synthesis and diminished activity of cellular energy-dependent processes, could contribute to cell death. In order to model, in vitro, conditions present post-ischemia or during toxin exposure, the interactions between Ca2+ and oxygen free radicals on isolated renal mitochondria were characterized. The oxygen free radicals were generated by hypoxanthine and xanthine oxidase to simulate in vitro one of the sources of oxygen free radicals in the early post-ischemic period in vivo. With site I substrates, pyruvate and malate, Ca2+ pretreatment, followed by exposure to oxygen free radicals, resulted in an inhibition of electron transport chain function and complete uncoupling of oxidative phosphorylation. These effects were partially mitigated by dibucaine, a phospholipase A2 inhibitor. With the site II substrate, succinate, the electron transport chain defect was not manifest and respiration remained partially coupled. The electron transport chain defect produced by Ca2+ and oxygen free radicals was localized to NADH CoQ reductase. Calcium and oxygen free radicals reduced mitochondrial ATPase activity by 55% and adenine nucleotide translocase activity by 65%. By contrast oxygen free radicals alone reduced ATPase activity by 32% and had no deleterious effects on translocase activity. Dibucaine partially prevented the Ca2+-dependent reduction in ATPase activity and totally prevented the Ca2+-dependent translocase damage observed in the presence of oxygen free radicals. These findings indicate that calcium potentiates oxygen free radical injury to mitochondria. The Ca2+-induced potentiation of oxygen free radical injury likely is due in part to activation of phospholipase A2. This detrimental interaction associated with Ca2+ uptake by mitochondria and exposure of the mitochondria to oxygen free radicals may explain the enhanced cellular injury observed during post-ischemic reperfusion.
在多种形式的缺血性和中毒性组织损伤中，细胞内Ca2+的蓄积和氧自由基的产生可能会对细胞尤其是线粒体膜产生不利影响。线粒体受损会导致ATP合成受损以及细胞能量依赖过程的活性降低，这可能会导致细胞死亡。为了在体外模拟缺血后或毒素暴露期间的情况，对分离的肾线粒体上Ca2+与氧自由基之间的相互作用进行了表征。通过次黄嘌呤和黄嘌呤氧化酶产生氧自由基，以模拟体内缺血后早期氧自由基的来源之一。使用位点I底物丙酮酸和苹果酸时，先用Ca2+预处理，然后暴露于氧自由基，会导致电子传递链功能受到抑制以及氧化磷酸化完全解偶联。这些效应被磷脂酶A2抑制剂丁卡因部分减轻。使用位点II底物琥珀酸时，未表现出电子传递链缺陷，呼吸仍部分偶联。由Ca2+和氧自由基产生的电子传递链缺陷定位于NADH辅酶Q还原酶。钙和氧自由基使线粒体ATP酶活性降低55%，腺嘌呤核苷酸转位酶活性降低65%。相比之下，单独的氧自由基使ATP酶活性降低32%，对转位酶活性没有有害影响。丁卡因部分阻止了Ca2+依赖性的ATP酶活性降低，并完全阻止了在有氧自由基存在下观察到的Ca2+依赖性转位酶损伤。这些发现表明钙会增强氧自由基对线粒体的损伤。Ca2+诱导的氧自由基损伤增强可能部分归因于磷脂酶A2的激活。这种与线粒体摄取Ca2+以及线粒体暴露于氧自由基相关的有害相互作用可能解释了缺血后再灌注期间观察到的细胞损伤增强现象。
