Role of JNK and p38MAPK signaling pathways in attenuation of myocardial ischemia-reperfusion injury by morphine postconditioning: an in vitro experiment

Objective To evaluate the role of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK) signaling pathways in attenuation of myocardial ischemia-reperfusion (I/R) injury by morphine postconditioning. Methods Healthy adult male Sprague-Dawley rats, weighing 180-240 g, were used in the study.Their hearts were excised and retrogradely perfused in a Langendorff apparatus with Krebs-Ringer (K-R) buffer saturated with 95% O2 -5% O2 at 37 ℃.After 15 min of equilibration, 52 isolated hearts were divided into 4 groups (n=13 each) using a random number table: control group (group C), I/R group, morphine postconditioning group (group MP), and morphine postconditioning plus anisomycin group (group MP+ A). The hearts were continuously perfused with K-R buffer for 105 min in group C. In group I/R, the hearts were subjected to 45 min of global ischemia by stopping perfusion with K-R buffer, followed by 60 min of reperfusion by restoration of perfusion with K-R buffer.In group MP, the hearts were subjected to 45 min of global ischemia, followed by 10 min of reperfusion with K-R buffer containing 3.0 μmol/L morphine and then by 50 min of reperfusion with K-R buffer.In group MP+ A, the hearts were subjected to 45 min of global ischemia, followed by 10 min of reperfusion with K-R buffer containing 3.0 μmol/L morphine and 1.0 μmol/L anisomycin (an activator of JNK and p38MAPK) and then by 50 min of reperfusion with K-R buffer.At 60 min of reperfusion, 8 hearts in each group were selected for measurement of the myocardial infarction and amount of creatine kinase-MB (CK-MB) released from the myocardium, and the myocardial infarct size was calculated.At 20 min of reperfusion, 5 hearts in each group were selected to detect the expression of phosphorylated JNK (p-JNK), phosphorylated p38MAPK (p-p38MAPK) and cytochrome c (Cyt c) in myocardial tissues (by Western blot) and content of nicotinamide adenine dinucleotide (NAD+ ) in myocardial tissues (by spectrophotometry). Results Compared to group C, the myocardial infarct size and amount of CK-MB released from the myocardium were significantly increased, the expression of p-JNK, p-p38MAPK and Cyt c was significantly up-regulated, and the content of NAD+ was significantly decreased in I/R, MP and MP+ A groups (P<0.05). Compared to group I/R, the myocardial infarct size and amount of CK-MB released from the myocardium were significantly decreased in MP and MP+ A groups, and the expression of p-JNK, p-p38MAPK and Cyt c was significantly down-regulated, and the content of NAD+ was significantly increased in group MP (P<0.05). Compared to group MP, the myocardial infarct size and amount of CK-MB released from the myocardium were significantly increased, the expression of p-JNK, p-p38MAPK and Cyt c was significantly up-regulated, and the content of NAD+ was significantly decreased in group MP+ A (P<0.05). Conclusion The mechanism by which morphine postconditioning attenuates myocardial I/R injury is related to inhibition of activation of JNK and p38MAPK signaling pathways in rats. Key words: JNK mitogen-activated protein kinases; p38 mitogen-activated protein kinases; Signal transduction; Morphine; Ischemic postconditioning; Reperfusion injury