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 a 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 number 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 compared with 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 (usual, 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 comparing with a single injection immediately after coronary ligation. II. Therapeutic window of cardioprotective properties of Erythropoietin in experimental ischemic-reperfusion model. While effective doses and the therapeutic window of EPO in the experimental model of permanent ligation of a coronary artery were well established in our previous studies (see I above), most published experimental studies on cardioprotective properties of EPO were based on more a clinically relevant model - temporary myocardial ischemic followed by reperfusion. In those reports, the EPO was injected either preventively, prior to interruption of myocardial circulation, or at the moment of restoration of circulation. Both scenarios are logistically not feasible in a clinical setting, where experimental treatment most likely will be following the interventional revascularization. In the present study we employed a rat ischemic/reperfusion model. The time of ischemia was chosen as 2 hrs - the average time between the onset of the symptoms of heart attack and implementation of clinical intervention to restore circulation in the hospital (data for USA). After 2 hrs of ischemia the circulation was restored. A single dose of EPO, 3000 IU/kg, was injected intraperitoneal either at the moment of restoration of myocardial circulation or 2 hrs later. The size of the ensuing MI, as well as the extent of apoptosis and inflammation in the area at risk were assessed histologically 24 hrs following the onset of ischemia. Results indicated that the effects of EPO applied at the moment of reperfusion were comparable with effects of EPO described for the model of a permanent coronary ligation with respect to MI size and apoptosis: MI size was significantly reduced in EPO-treated compared to untreated animals and the extent of apoptosis was significantly attenuated. However, the EPO injected 2 hrs following reperfusion did not affect either ensuing MI size or the apoptosis in the area at risk. Results, while preliminary at this time, are important for design of future clinical trials.
