IMPROVEMENT OF ISCHEMIC TOLERANCE AND COLD PRESERVATION OF AGED MYOCARDIUM BY TRANSFECTION OF HEAT-SHOCK PROTEIN GENE

1. 随着年龄的增长，缺血耐受性降低：分析了12、50和100周龄Fischer 344大鼠离体心脏左心室功能和能量代谢产物的恢复、心肌酶（CPK和LDH）的释放以及缺血-再灌注心律失常。根据langendorff技术，心脏缺血15 ~ 25min，再灌注30min。功能和代谢物的恢复以及酶的释放显示出缺血性耐受性的年龄依赖性下降。老年大鼠心脏缺血-再灌注心律失常也更恶性。在长时间缺血前进行缺血或缺氧预处理对老年心脏没有任何有益作用，甚至会加重心肌损伤。基因转染：hvj -脂质体+质粒法在转染HSP基因的肌细胞中仅检测到30%的热休克蛋白（HSP）。转染后24、48或72小时，该百分比无差异。当阳离子hvj脂质体用量增加时，效率没有显著提高。抗HSP抗体免疫细胞化学分析显示，HSP在内皮细胞中高发，提示阳离子hvj脂质体可能在到达肌细胞之前被困在内皮细胞中。转染HSP基因的心脏在冷停搏和缺血再灌注损伤后的保存：转染HSP基因的心脏在冷停搏10分钟后保存2小时后，其功能和代谢物的恢复以及酶的释放都有改善的趋势，尽管转染HSP基因并没有减少缺血再灌注心律失常。另一方面，心肌缺血25 min和再灌注30 min的功能和代谢产物的恢复、酶的释放、缺血-再灌注心律失常的改善均不显著。离体肌细胞模拟缺血：在转染HSP基因的任意年龄的肌细胞中，模拟缺血（缺氧、pH6.5、乳酸20mM、k12mM） 3min后，细胞内Ca的升高减弱，细胞运动恢复得到改善。少

1. Decrease in ischemic tolerance with aging: Recovery of left ventricular function and energy metabolites, release of myocardial enzymes (CPK and LDH), and ischemia-reperfusion arrhythmias were analyzed in hearts isolated from 12, 50 or 100 week-old Fischer 344 rats. Hearts were subjected to 15 to 25 min of ischemia and 30 min of reperfusion according to the langendorff technique. Recovery of function and metabolites and release of enzymes demonstrated age-dependent decrease in ischemic tolerance. Ischemic-reperfusion arrhythmias were also more malignant in older rat hearts. Ischemic or hypoxic preconditioning performed before prolonged ischemia did not have any beneficial effects or even deteriorated myocardial damage in older hearts.2. Gene transfection: Heat shock protein (HSP) was detected only 30% of myocytes transfected with HSP gene using HVJ-liposome+plasmid methods. This percentage was not different between 24, 48 or 72 hours after transfection. When cationic HVJ-liposome was … More used, efficiency was not improved remarkably. Immunocytochemical analysis with anti-HSP antibody revealed that HSP was detected with high incidence in endothelial cells, which suggested cationic HVJ-liposome might be trapped in the endothelial cells before reaching to myocytes.3. Preservation after cold cardioplegia and iscchemia-reperfusion injury in HSP-transfected heats: Recovery of function and metabolites and release of enzymes in hearts subjected to 10 min cold cardioplegia followed by 2 hours preservation were tended to be improved when hearts of any age were transfected with HSP gene, although ischemia-reperfusion arrhythmias were not decreased by HSP transfection. On the other hand, recovery of function and metabolites, release of enzymes, and ischemia-reperfusion arrhythmias in hearts subjected to 25 min of ischemia and 30 min of reperfusion showed only insignificant improvement.4. Simulated ischemia in isolated myocytes: In myocytes of any age that were transfected with HSP gene, increase in intracellular Ca was attenuated and recovery of cell motion was improved after 3 min of simulated ischemia (hypoxia, pH6.5, lactate 20mM, k12mM). Less