Cardiovascular Drugs and Therapy 19 33–40 2005 C  2005 Springer Science + Business Media, Inc. Manufactured in The Netherlands BASIC PHARMACOLOGY Erythropoietin Just Before Reperfusion Reduces Both Lethal Arrhythmias and Infarct Size via the Phosphatidylinositol-3 Kinase-Dependent Pathway in Canine Hearts Akio Hirata 1 , Tetsuo Minamino 1 , Hiroshi Asanuma 1 , Shoji Sanada 1 , Masashi Fujita 1 , Osamu Tsukamoto 1 , Masakatsu Wakeno 2 , Masafumi Myoishi 2 , Ken-ichiro Okada 1 , Hidekazu Koyama 1 , Kazuo Komamura 3 , Seiji Takashima 1 , Yoshiro Shinozaki 4 , Hidezo Mori 3 , Hitonobu Tomoike 3 , Masatsugu Hori 1 , and Masafumi Kitakaze 3 1 Department of Internal Medicine and Therapeutics, Osaka University Graduate School of Medicine, Suita, Osaka, Japan; 2 Department of Bioregulatory Medicine, Osaka University Graduate School of Medicine, Suita, Osaka, Japan; 3 Cardiovascular Division of Internal Medicine, National Cardiovascular Center, Suita, Osaka, Japan; 4 Department of Physiological Science, Tokai University School of Medicine, Isehara, Kanagawa, Japan Summary. Although recent studies suggest that erythropoi- etin (EPO) may reduce multiple features of the myocardial ischemia/reperfusion injury, the cellular mechanisms and the clinical implications of EPO-induced cardioprotec- tion are still unclear. Thus, in this study, we clarified dose-dependent effects of EPO administered just before reperfusion on infarct size and the incidence of ventric- ular fibrillation and evaluated the involvement of the phosphatidylinositol-3 (PI3) kinase in the in vivo canine model. The canine left anterior descending coronary artery was occluded for 90 min followed by 6 h of reperfusion. A single intravenous administration of EPO just before reperfusion significantly reduced infarct size (high dose (1,000 IU/kg): 7.7 ± 1.6%, low dose (100 IU/kg): 22.1 ± 2.4%, control: 40.0 ± 3.6%) in a dose-dependent manner. Fur- thermore, the high, but not low, dose of EPO adminis- tered as a single injection significantly reduced the inci- dence of ventricular fibrillation during reperfusion (high dose: 0%, low dose: 40.0%, control: 50.0%). An intracoro- nary administration of a PI3 kinase inhibitor, wortman- nin, blunted the infarct size-limiting and anti-arrhythmic ef- fects of EPO. Low and high doses of EPO equally induced Akt phosphorylation and decreased the equivalent num- ber of TUNEL-positive cells in the ischemic myocardium of dogs. These effects of EPO were abolished by the treatment with wortmannin. In conclusion, EPO administered just be- fore reperfusion reduced infarct size and the incidence of ventricular fibrillation via the PI3 kinase-dependent pathway in canine hearts. EPO administration can be a realistic strategy for the treatment of acute myocardial infarction. Key Words. erythropoietin, myocardial infarction, ventric- ular arrhythmia, phosphatidylinositol-3 kinase, ischemia- reperfusion injury, apoptosis Introduction Recent studies have extended the traditional role of erythropoietin (EPO) from a mediator of erythroid maturation to one that provides protection against apoptotic cell death [1,2]. Recombinant human EPO (rhEPO) has been shown to exert marked protective effects against ischemia/reperfusion injury in rats and rabbits when rhEPO is administered at different time points [3–8]. Indeed, rhEPO reduced myocardial infarct size, enhanced recovery of left ventricular developed pressure, reduced the number of apoptotic cells, and induced the phosphorylation of Akt [3–8]. In these stud- ies, high (1,000–5,000 IU/kg) doses of rhEPO, nearly 10 times higher than that used in anemic patients with chronic renal failure [9], have been applied. Recently, it was reported that phosphatidylinositol-3 (PI3) kinase Address for correspondence: Tetsuo Minamino, MD, PhD, De- partment of Internal Medicine and Therapeutics, Osaka Univer- sity Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan, Tel.: 81-6-6879-3635; Fax: 81-6-6879-3473; E-mail: minamino@medone.med.osaka-u.ac.jp 33