Inserte Javier, Garcia-Dorado David, Hernando Victor, Barba Ignasi, Soler-Soler Jordi
Servicio de Cardiologia, Hospital Universitari Vall d'Hebron, Passeig Vall d'Hebron, 119-129, 08035 Barcelona, Spain.
Cardiovasc Res. 2006 May 1;70(2):364-73. doi: 10.1016/j.cardiores.2006.02.017. Epub 2006 Mar 6.
We previously demonstrated that ischemic preconditioning (IPC) attenuates calpain activation during reperfusion. Herein, we tested the hypothesis that enhancement of Na+/K+-ATPase activity during early reperfusion as a result of calpain inhibition is involved in the protection afforded by myocardial IPC.
Intracellular Na+ concentration ([Na+]i) measured using 23Na-magnetic resonance spectroscopy, Na+/K+-ATPase activity, detachment of Na+/K+-ATPase alpha subunits from the membrane cytoskeleton, degradation of fodrin and ankyrin, and calpain activation were analysed in isolated rat hearts reperfused after 60 min of ischemia with or without previous IPC and different treatments aimed to mimic or blunt the effects of IPC.
In non-treated hearts subjected to ischemia (control hearts), reperfusion for 5 min severely reduced Na+/K+-ATPase activity and dissociated alpha1 and alpha2 subunits of Na+/K+-ATPase from the membrane-cytoskeleton complex in parallel with proteolysis of alpha-fodrin and ankyrin-B and calpain activation. IPC accelerated the recovery of [Na+]i, increased Na+/K+-ATPase activity, and prevented dissociation of Na+/K+-ATPase from the membrane-cytoskeleton complex. IPC also prevented alpha-fodrin and ankyrin-B loss and calpain activation, effects that were associated with attenuated lactate dehydrogenase (LDH) release and infarct size and improved contractile recovery. These effects of IPC were reproduced by perfusing the hearts with the calpain inhibitor MDL-28170 and by transient stimulation of cAMP-dependent protein kinase (PKA) with CPT-cAMP, and they were reverted by perfusing with the PKA inhibitor H89.
The results of the present study are consistent with the hypothesis that enhanced recovery of Na+/K+-ATPase activity during reperfusion as a result of attenuated calpain-mediated detachment of the protein from the membrane-cytoskeleton complex contributes to the protection afforded by IPC.
我们之前证明了缺血预处理（IPC）可减轻再灌注期间的钙蛋白酶激活。在此，我们检验了以下假设：钙蛋白酶抑制导致再灌注早期钠钾ATP酶（Na+/K+-ATPase）活性增强，这与心肌IPC所提供的保护作用有关。
使用23Na磁共振波谱法测量细胞内钠浓度（[Na+]i），分析Na+/K+-ATPase活性、Na+/K+-ATPaseα亚基从膜细胞骨架上的脱离、血影蛋白和锚蛋白的降解以及钙蛋白酶激活情况，这些分析是在经历60分钟缺血后再灌注的离体大鼠心脏中进行的，心脏分为有或没有预先IPC处理的组，以及接受不同处理以模拟或减弱IPC作用的组。
在未经处理的缺血心脏（对照心脏）中，再灌注5分钟会严重降低Na+/K+-ATPase活性，并使Na+/K+-ATPase的α1和α2亚基与膜细胞骨架复合物解离，同时伴有α-血影蛋白和锚蛋白-B的蛋白水解以及钙蛋白酶激活。IPC加速了[Na+]i的恢复，增加了Na+/K+-ATPase活性，并防止了Na+/K+-ATPase从膜细胞骨架复合物上解离。IPC还防止了α-血影蛋白和锚蛋白-B的丢失以及钙蛋白酶激活，这些作用与乳酸脱氢酶（LDH）释放减少、梗死面积减小以及收缩功能恢复改善相关。通过用钙蛋白酶抑制剂MDL-28170灌注心脏以及用CPT-cAMP短暂刺激环磷酸腺苷依赖性蛋白激酶（PKA），可重现IPC的这些作用，而用PKA抑制剂H89灌注可逆转这些作用。
本研究结果与以下假设一致：由于钙蛋白酶介导的蛋白质从膜细胞骨架复合物上的脱离减弱，再灌注期间Na+/K+-ATPase活性的增强恢复有助于IPC所提供的保护作用。
