Weinbrenner C, Liu G S, Cohen M V, Downey J M
Department of Physiology, University of South Alabama, Mobile, AL 36688, USA.
J Mol Cell Cardiol. 1997 Sep;29(9):2383-91. doi: 10.1006/jmcc.1997.0473.
p38 mitogen-activated protein kinase (MAPK) is known to be activated after exposure to endotoxin, osmotic and environmental stress, and, most recently, during ischemia/reperfusion. We investigated whether ischemic preconditioning also causes phosphorylation of the activation sites on p38 MAPK. Three groups of isolated rabbit hearts were studied. Control hearts experienced 30 min of ischemia only. The second group was preconditioned with 5 min of global ischemia and 10 min of reperfusion. Group 3 was also ischemically preconditioned, but in the presence of 100 microM 8-(p-sulfophenyl)theophylline (SPT). Transmural left ventricular biopsies were taken before and during the long ischemic period. Western blot analysis with either p38 MAPK or phospho-specific p38 MAPK (Tyr-182) antibodies showed a decreased phosphorylation during ischemia in non-preconditioned hearts, but phosphorylation was enhanced several fold after 10 and 20 min of ischemia in preconditioned hearts. Furthermore, when protection from ischemic preconditioning was blocked by SPT, increased phosphorylation of p38 MAPK during ischemia was not present. Therefore the phosphorylation of p38 MAPK at tyrosine 182, which is required for the kinase's activation, occurred during ischemia only when protection from preconditioning was evident. In a second study, changes in osmotic fragility were measured during simulated ischemia in rabbit cardiomyocytes. Reduced fragility in ischemically preconditioned myocytes could be completely abolished by the specific p38 MAPK inhibitor SB-203580. In contrast, anisomycin, an activator of p38 MAPK and JUN kinase pathways, was found to be as protective as ischemic preconditioning. We conclude that p38 MAPK phosphorylation correlates with preconditioning's protection, and that its activation may be an important step in the signal transduction cascade of ischemic preconditioning.
已知p38丝裂原活化蛋白激酶（MAPK）在暴露于内毒素、渗透压和环境应激后被激活，最近发现其在缺血/再灌注过程中也会被激活。我们研究了缺血预处理是否也会导致p38 MAPK激活位点的磷酸化。研究了三组离体兔心脏。对照组心脏仅经历30分钟的缺血。第二组用5分钟的全心缺血和10分钟的再灌注进行预处理。第三组也进行了缺血预处理，但同时存在100微摩尔的8-（对磺基苯基）茶碱（SPT）。在长时间缺血期之前和期间采集透壁左心室活检组织。用p38 MAPK或磷酸化特异性p38 MAPK（Tyr-182）抗体进行的蛋白质印迹分析显示，未预处理的心脏在缺血期间磷酸化减少，但预处理的心脏在缺血10分钟和20分钟后磷酸化增强了几倍。此外，当SPT阻断缺血预处理的保护作用时，缺血期间p38 MAPK的磷酸化增加就不存在了。因此，激酶激活所需的p38 MAPK酪氨酸182位点的磷酸化仅在缺血期间出现，且此时预处理的保护作用明显。在第二项研究中，测量了兔心肌细胞模拟缺血期间的渗透脆性变化。特异性p38 MAPK抑制剂SB-203580可完全消除缺血预处理心肌细胞中降低的脆性。相反，发现p38 MAPK和JUN激酶途径的激活剂茴香霉素与缺血预处理具有相同的保护作用。我们得出结论，p38 MAPK磷酸化与预处理的保护作用相关，其激活可能是缺血预处理信号转导级联中的一个重要步骤。
