Molecular mechanisms underlying myocardial hypertrophy induced by ischemia : correlation between changes in intracellular ion concentration and those in expression of immediate-early genes.

最近的研究表明，在包括心肌细胞在内的许多类型的细胞中，即早基因（IEG）和/或蛋白激酶级联在恢复或适应各种应激或生长刺激中起着重要作用。为了研究心肌细胞如何从短暂缺血中恢复，我们采用体外缺血模型之一的鸡胚（10日龄）心室肌细胞代谢抑制（MI）模型来观察c-jun mRNA的诱导和丝裂原活化蛋白（MAP）激酶的活性。我们研究了使用1 mM氰化钠和20 mM 2-脱氧-d-葡萄糖进行心肌梗死期间以及心肌梗死恢复期间c-jun mRNA的相对丰富度（通过激光密度测定）保持不变（RNA印迹分析），MAP激酶活性（凝胶激酶测定）和[Ca^<2+>] i的水平。但在30,60,90,120 min的恢复阶段显著增加（分别是心肌梗死前的3.0,4.7,2.4,1.9倍）。当细胞用100 mM的H7（一种有效的蛋白激酶C抑制剂）预处理时，在恢复阶段的60分钟，编码C -jun的mRNA的诱导被明显抑制（在没有H-7的情况下，在恢复阶段的60分钟，C -jun mRNA的水平降低了95%）。用2 mM EGTA预处理心肌细胞完全抑制心肌梗死期间[Ca^<2+>] i的增加，[Ca^<2+>] i保持在非常低的水平（心肌梗死30分钟时184<正负>26 nM,n=3）。与未使用EGTA的患者相比，EGTA预处理使心肌梗死30分钟后恢复60分钟时c-jun mRNA水平降低42% （p<0.01, n=3）。MAP激酶活性在心肌梗死期间也没有变化，但在恢复期的5、10、15分钟显著增加（分别比心肌梗死前增加3.0倍、4.1倍、3.4倍）。因此，这些数据表明，IEG c-jun和MAP激酶可能在心肌细胞短暂缺血后的恢复中发挥重要作用。这种C -jun表达的增加可能需要蛋白激酶C的激活，并且在一定程度上需要细胞内Ca^<2+>。少

Recent studies have revealed that in many types of cells including cardiac myocytes immediate-early genes (IEG) and/or protein kinase cascade play important roles in recovering or adapting to various stresses or growth stimuli. In order to investigate how cardiac myocytes recover from a brief period of ischemia, we used a metabolic inhibition (MI) model, one of in vitro ischemia models, of chick embryo (10-day-old) ventricular myocytes to examine the induction of c-jun mRNA and the activity of mitogen-activated protein (MAP) kinase. We studied the levels of c-jun mRNA (RNA blot analysis), MAP kinase activity (in-gel kinase assay) and [Ca^<2+>] i (measured with indo-1) during MI using 1 mM sodium cyanide and 20 mM 2-deoxy-d-glucose and also during the recovery from MI.The relative abundance of c-jun mRNA (determined by laser densitometry) remained unchanged during MI of 30 min, but increased significantly during the recovery phase at 30,60,90,120 min (3.0,4.7,2.4,1.9-fold induction as c … More ompared with the pre-MI controls, respectively). When the cells were pretreated with 100 mM of H7, one of the potent protein kinase C inhibitors, the induction of mRNA encoding c-jun at 60 min. during the recovery phase was markedly suppressed (95% reduction of the levelos of c-jun mRNA at 60 min during the recovery without H-7). Pretreatment of the cardiac myocytes with 2 mM EGTA completely inhibited the increase in [Ca^<2+>] i during MI and [Ca^<2+>] i was kept at very lower level (184<plus-minus>26 nM at 30 min. during MI,n=3). This EGTA-pretreatment reduced the levels of c-jun mRNA by 42% (p<0.01, n=3) at 60 min. during recovery from MI of 30 min. as compared with those without EGTA.MAP kinase activity was also unchanged during MI,but significantly increased at 5,10,15 min during the recovery phase (3.0,4.1,3.4-fold increase as compared with the pre-MI controls, respecitively). Thus, these data suggest that IEG c-jun as well as MAP kinase may play significant roles in recovery of cardiac myocytes from a brief period of ischemia. This augmentation of c-jun expression may need the activation of protein kinase C,and to some extent, intracellular Ca^<2+>. Less