Reduction of myocardial damage during acute ischemia

该计划的主要目标是在心肌缺血的动物模型上进行临床前实验，以阐明心肌细胞死亡和随后的CHF发展的机制，并评估不同治疗方式的潜力。最终目的是限制心肌损害的程度，预防或减轻CHF的发展。 促红细胞生成素减轻心肌缺血损伤。 促红细胞生成素(EPO)是一种天然的促红细胞生成剂，近年来被认为是一种具有神经保护作用的潜在抗细胞凋亡因子。我们已经证明，EPO的抗细胞凋亡作用也有心脏保护作用。在大鼠的实验中，我们发现，在冠状动脉永久结扎后立即全身注射重组人促红细胞生成素(3000IU/kg)，可使8周后的心肌梗死面积缩小75%。在心肌梗死后8周内，EPO治疗组大鼠左室重构和功能下降明显减轻，与假手术组比较差异无统计学意义。冠状动脉结扎24小时后，与未处理的大鼠相比，EPO处理的大鼠在心肌危险区域(与梗塞部位直接相邻的区域)测得的心肌细胞凋亡量减少了一半。在随后的实验中，我们证实了单次静脉注射3000IU/kg在冠状动脉结扎后12小时内具有心脏保护作用，但在24小时后就失去了心脏保护作用。如果动物在冠状动脉结扎后立即接受EPO治疗，治疗剂量可以减少到150IU/kg(FDA批准的治疗贫血的剂量)，而不会失去疗效；然而，在这种情况下，治疗窗口也减少到4小时。在另外的实验中，我们发现，与冠状动脉结扎后立即单次注射相比，每天重复注射EPO并没有任何额外的好处。 2.胰高血糖素样肽1(GLP-1)是肠上皮细胞分泌的一种由30个氨基酸组成的肽类激素，由原胰高血糖素分化处理而成。GLP-1是对进食的反应而释放的，在激素离开肠道之前，GLP-1就被二肽基肽酶IV极快地代谢和失活。GLP-1的主要作用是刺激胰岛素的分泌，即作为一种胰岛素激素，并抑制胰升糖素的分泌，从而有助于限制餐后血糖的漂移。它还抑制胃肠运动和分泌，从而起到肠胃泌素和回肠制动机制的作用。GLP-1似乎也是食欲和食物摄入量的生理调节器。由于这些作用，GLP-1或GLP-1受体激动剂目前正在接受2型糖尿病治疗的评估。GLP-1分泌减少可能有助于肥胖的发展，而过度分泌可能是餐后反应性低血糖的原因。GLP-1及其特异性受体在中枢神经系统和心脏中表达。在心脏方面，GLP-1受体的激活改善了血管成形术后患者的心脏血流动力学。以前的研究已经证明了GLP-1对大鼠和小鼠的血压和脉搏的有益影响。GLP-1信号似乎在心脏发育中发挥作用，因为GLP-1R靶向基因缺失的小鼠心脏变大。然而，最近的研究表明，GLP-1可能通过直接介导的方式对心脏功能产生有益影响。 在冠状动脉结扎诱导的大鼠心肌梗死模型上，我们观察了(1)GLP-1R激动剂对心肌缺血损伤的保护作用，以及(2)GLP-1R激动剂能否减轻心肌梗死后的心脏重构。实验一以400 ng/kg/h的微量渗透压泵给4m龄雄性SD大鼠静脉注射GLP-1R激动剂Exendin-4，连续7d后结扎冠状动脉，术后24小时处死大鼠，观察心肌梗死大小。实验二：冠脉结扎后2周开始注入Exendin-4(400 ng/kg/h)，持续8周。 我们发现冠状动脉结扎后24小时，心肌梗死大小不受GLP-1R激动剂的影响。用GLP-1R激动剂治疗8周后，心率略有增加，但体重和尾压没有变化。然而，在心梗后心力衰竭大鼠，GLP-1R激动剂治疗8周后，心肌梗死扩张和心脏重构停止，心功能恶化。

The broad objective of this program is to perform preclinical experimentation on animal models of myocardial ischemia to elucidate the mechanisms of cellular death in the myocardium and development of the subsequent CHF and to evaluate the potential of different therapeutic modalities. The ultimate goal is to limit the extent of myocardial damage and to prevent or attenuate the development of CHF. I. Erythropoietin reduces the myocardial ischemic damage. Erythropoietin (EPO), natural stimulant of erythropoiesis, recently emerged as potential antiapoptotic factor with neuroprotective properties. We have demonstrated that the antiapoptotic effects of EPO also resulted in cardioprotection. In experiments in rats we showed that single systemic administration of recombinant human EPO (3000 IU/kg) immediately after permanent ligation of a coronary artery results in 75% reduction of the size of myocardial infarction eight weeks later. During eight weeks after induction of myocardial infarction, left ventricular remodeling and function decline in EPO treated rats was significantly attenuated and statistically was not different from that in sham operated animals. Twenty four hours after ligation of coronary artery the amount of apoptotic myocytes measured in the myocardial risk area (area immediately adjacent to the infarct site) was reduced by half in the EPO treated rats in comparison to untreated animals. In subsequent experiments we established that a single intravenous dose of 3000 IU/kg is cardioprotective up to 12 hrs after coronary ligation, but it loses its cardioprotective properties after 24 hrs. If animals are treated with EPO immediately after coronary ligation, the treatment dose can be reduced up to 150 IU/kg (FDA approved dose for the treatment of anemia) without the loss of effectiveness; however, the therapeutic window in this case is also reduced to 4 hrs. In additional experiments we showed that repeated daily EPO injections do not have any added benefits compared with a single injection immediately after coronary ligation. II. Glucagon-like peptide 1 (GLP-1) is a 30-amino acid peptide hormone produced in the intestinal epithelial endocrine L-cells by differential processing of proglucagon. GLP-1 is released in response to meal intake, extremely rapidly metabolized and inactivated by the enzyme dipeptidyl peptidase IV even before the hormone has left the gut. The main actions of GLP-1 are to stimulate insulin secretion, i.e., to act as an incretin hormone, and to inhibit glucagon secretion, thereby contributing to limit postprandial glucose excursions. It also inhibits gastrointestinal motility and secretion and thus acts as an enterogastrone and part of the ileal brake mechanism. GLP-1 also appears to be a physiological regulator of appetite and food intake. Because of these actions, GLP-1 or GLP-1 receptor agonists are currently being evaluated for the therapy of type 2 diabetes. Decreased secretion of GLP-1 may contribute to the development of obesity, and exaggerated secretion may be responsible for postprandial reactive hypoglycemia. GLP-1 and its specific receptor are expressed in the central nervous system and heart. In the heart, GLP-1 receptor activation improves cardiac hemodynamics in patients following angioplasty. Previous studies have demonstrated the beneficial effects of GLP-1 on blood pressure and pulse in rats and mice. GLP-1 signaling seems to play a role in cardiac development since mice with a targeted gene deletion of the GLP-1r have enlarged hearts. Recent work, however, indicates that GLP-1 may have beneficial effects on cardiac function that are mediated directly. In the rat model of myocardial infarction induced by a permanent ligation of coronary artery we intended to test (1) whether pretreatment with GLP-1R agonist protects myocardium from ischemic damage, and (2) whether treatment with GLP-1R agonist might attenuate the cardiac remodeling after induction of myocardial infarction. In the first experiment, GLP-1R agonist, Exendin-4, was delivered to 4m old male SD rats by mini-osmotic pump at a rate of 400ng/kg/h for 7 days prior to ligation of a coronary artery and rats were euthanized 24 hrs after surgery to assess the MI size. In the second experiment, Exendin-4 was initiated via osmotic pump implantation (400ng/kg/h) 2 weeks after coronary ligation and continued for 8 weeks. We found that 24h after coronary ligation the MI size was not affected by pretreatment with GLP-1R agonist. 8-week treatment with GLP-1R agonist slightly increased the heart rate but did not change body weight and tail blood pressure. However, in post-MI heart failure rats, 8-week treatment with GLP-1R agonist resulted in arrest of the infarct expansion and cardiac remodeling, and deterioration of cardiac function.