Das D K, Maulik N, Sato M, Ray P S
Department of Surgery, University of Connecticut School of Medicine, Farmington 06030-1110, USA.
Mol Cell Biochem. 1999 Jun;196(1-2):59-67.
Ischemic preconditioning has been shown to trigger a signaling pathway by potentiating tyrosine kinase phosphorylation leading to the activation of p38 MAP kinase and MAPKAP kinase 2. Recently, the nuclear transcription factor, NFkappaB, was found to play a role in the signaling process. Since NFkappaB is a target of oxygen free radicals, we hypothesized that reactive oxygen species might play a role in the signaling process. To test this hypothesis, isolated rat hearts were perfused in the absence or presence of either dimethyl thiourea (DMTU), a OH* radical scavenger, or SN 50 peptide, a NFkappaB blocker. Hearts were then subjected to ischemic preconditioning by four repeated episodes of 5 min ischemia each followed by 10 min reperfusion. All hearts were then made globally ischemic for 30 min followed by 2 h of reperfusion. The results of our study demonstrated enhanced tyrosine kinase phosphorylation during ischemic preconditioning which was blocked by DMTU. DMTU also inhibited preconditioning mediated increased phosphorylation of p38 MAP kinase and MAPKAP kinase 2 activity. However, DMTU had no effect on the translocation and activation of protein kinase C (PKC) resulting from preconditioning. Preconditioning reduced myocardial infarct size as expected. This cardioprotective effect of preconditioning was abolished by both DMTU and SN 50. Preconditioning resulted in the nuclear translocation and activation of NFkappaB. Increased NFkappaB binding was blocked by both DMTU and SN 50. The results of this study demonstrate that reactive oxygen species play a crucial role in signal transduction mediated by preconditioning. This signaling process appears to be potentiated by tyrosine kinase phosphorylation resulting in the activation of p38 MAP kinase and MAPKAP kinase 2 leading to the activation of NFkappaB suggesting a role of oxygen free radicals as second messenger. Free radical signaling seems to be independent of PKC although PKC is activated during preconditioning process suggesting the role of two separate signaling pathways in ischemic preconditioning.
缺血预处理已被证明可通过增强酪氨酸激酶磷酸化来触发信号通路，从而导致p38丝裂原活化蛋白激酶（MAP激酶）和MAPKAP激酶2的激活。最近，发现核转录因子NFκB在该信号传导过程中起作用。由于NFκB是氧自由基的靶点，我们推测活性氧可能在信号传导过程中起作用。为了验证这一假设，将离体大鼠心脏在不存在或存在二甲基硫脲（DMTU，一种OH*自由基清除剂）或SN 50肽（一种NFκB阻滞剂）的情况下进行灌注。然后通过四次重复的5分钟缺血发作，每次发作后再灌注10分钟，对心脏进行缺血预处理。随后所有心脏进行30分钟的全心缺血，然后再灌注2小时。我们的研究结果表明，缺血预处理期间酪氨酸激酶磷酸化增强，而DMTU可阻断这种增强。DMTU还抑制预处理介导的p38 MAP激酶磷酸化增加和MAPKAP激酶2活性增加。然而，DMTU对预处理导致的蛋白激酶C（PKC）转位和激活没有影响。预处理如预期的那样减小了心肌梗死面积。DMTU和SN 50均消除了预处理的这种心脏保护作用。预处理导致NFκB的核转位和激活。DMTU和SN 50均阻断了NFκB结合增加。本研究结果表明，活性氧在预处理介导的信号转导中起关键作用。该信号传导过程似乎通过酪氨酸激酶磷酸化得到增强，导致p38 MAP激酶和MAPKAP激酶2激活，进而导致NFκB激活，提示氧自由基作为第二信使的作用。自由基信号传导似乎独立于PKC，尽管PKC在预处理过程中被激活，这表明在缺血预处理中有两条独立的信号传导途径发挥作用。
