Nakano A, Baines C P, Kim S O, Pelech S L, Downey J M, Cohen M V, Critz S D
Department of Physiology, University of South Alabama, Mobile, AL 36688-0002, USA.
Circ Res. 2000 Feb 4;86(2):144-51. doi: 10.1161/01.res.86.2.144.
Recent studies suggest that p38 mitogen-activated protein kinase (MAPK) may be involved in ischemic preconditioning (PC). To further test this possibility, the regulation of MAPK-activated protein kinase 2 (MAPKAPK2), a kinase immediately downstream from p38 MAPK, and the activity of c-Jun NH(2)-terminal kinase (JNK), a second MAPK, were examined in preconditioned hearts. Isolated, perfused rabbit hearts were subjected to 20 to 30 minutes of global ischemia. Ventricular biopsies before treatment and after 20 minutes of ischemia were homogenized, and the activities of MAPKAPK2 and JNK were evaluated. For the MAPKAPK2 experiments, 7 groups were studied, as follows: control hearts; preconditioned hearts; hearts treated with 500 nmol/L R(-) N(6)-(2-phenylisopropyl) adenosine (PIA), an A(1)-adenosine receptor agonist; preconditioned hearts pretreated with 100 micromol/L 8-(p-sulfophenyl) theophylline (SPT), an adenosine receptor antagonist; preconditioned hearts also treated with SB 203580, a potent inhibitor of p38 MAPK activation; hearts treated with 50 ng/mL anisomycin (a p38 MAPK/JNK activator); and hearts treated with both anisomycin (50 ng/mL) and the tyrosine kinase inhibitor genistein (50 micromol/L). MAPKAPK2 activity was not altered in control hearts after 20 minutes of global ischemia. By contrast, there was a 3.8-fold increase in activity during ischemia in preconditioned hearts. Activation of MAPKAPK2 in preconditioned hearts was blocked by both SPT and SB 203580. MAPKAPK2 activity during ischemia increased 3.5-fold and 3.3-fold in hearts pretreated with PIA or anisomycin, respectively. MAPKAPK2 activation during ischemia in hearts pretreated with anisomycin was blocked by genistein. In separate hearts, anisomycin mimicked the anti-infarct effect of PC, and that protection was abolished by genistein. JNK activity was measured in control and preconditioned hearts. There was a comparable, modest decline in activity during 30 minutes of global ischemia in both groups. As a positive control, a third group of hearts was treated with anisomycin before global ischemia, and in these, JNK activity increased by 290% above baseline. These results confirm that the p38 MAPK/MAPKAPK2 pathway is activated during ischemia only if the heart is in a preconditioned state. These data further support p38 MAPK as an important signaling component in ischemic PC.
近期研究表明，p38丝裂原活化蛋白激酶（MAPK）可能参与了缺血预处理（PC）。为进一步验证这种可能性，我们检测了p38 MAPK下游紧邻的激酶——MAPK活化蛋白激酶2（MAPKAPK2）的调节情况以及另一种MAPK——c-Jun氨基末端激酶（JNK）的活性，检测对象为经过预处理的心脏。将离体灌注的兔心脏进行20至30分钟的全心缺血处理。在处理前及缺血20分钟后采集心室组织活检样本并匀浆，评估MAPKAPK2和JNK的活性。对于MAPKAPK2实验，研究了7组对象，如下：对照心脏；预处理心脏；用500 nmol/L R(-)N(6)-(2-苯异丙基)腺苷（PIA）（一种A(1)-腺苷受体激动剂）处理的心脏；用100 μmol/L 8-(对磺基苯基)茶碱（SPT）（一种腺苷受体拮抗剂）预处理的预处理心脏；也用SB 203580（一种p38 MAPK激活的强效抑制剂）处理的预处理心脏；用50 ng/mL茴香霉素（一种p38 MAPK/JNK激活剂）处理的心脏；以及用茴香霉素（50 ng/mL）和酪氨酸激酶抑制剂染料木黄酮（50 μmol/L）共同处理的心脏。全心缺血20分钟后，对照心脏中的MAPKAPK2活性未改变。相比之下，预处理心脏在缺血期间活性增加了3.8倍。SPT和SB 203580均可阻断预处理心脏中MAPKAPK2的激活。用PIA或茴香霉素预处理的心脏在缺血期间MAPKAPK2活性分别增加了3.5倍和3.3倍。用茴香霉素预处理的心脏在缺血期间MAPKAPK2的激活被染料木黄酮阻断。在另外的心脏实验中，茴香霉素模拟了PC的抗梗死作用，而这种保护作用被染料木黄酮消除。检测了对照心脏和预处理心脏中的JNK活性。两组在全心缺血30分钟期间活性均有类似的适度下降。作为阳性对照，第三组心脏在全心缺血前用茴香霉素处理，在这些心脏中，JNK活性比基线水平增加了290%。这些结果证实，仅当心脏处于预处理状态时，p38 MAPK/MAPKAPK2通路才会在缺血期间被激活。这些数据进一步支持p38 MAPK作为缺血PC中的一个重要信号成分。
