Strategies to enhance skeletal myoblast (SkM) survival after transplantation in the ischemic heart have achieved little success. We posit that preconditioned (PC) SkMs show improved survival and promote repair of the infarcted myocardium via paracrine signaling after transplantation. SkMs from male Fischer-344 rats (rSkMs) were PC for 30 minutes with 200 micromol/L diazoxide. Treatment of PC rSkMs with 100 micromol/L H(2)O(2) for 2 hours resulted in significantly reduced cell injury, as shown by lactate dehydrogenase-release assay, and prevented apoptosis, as demonstrated by cytochrome c translocation, TUNEL, annexin V staining, and preservation of mitochondrial membrane potential. PC rSkMs expressed elevated phospho-Akt (1.85-fold), basic fibroblast growth factor (1.44-fold), hepatocyte growth factor (2.26-fold), and cyclooxygenase-2 (1.33-fold) as compared with non-PC rSkMs. For in vivo studies, female Fischer-344 rats after permanent coronary artery ligation were grouped (n=12/group) to receive 80 microL of basal medium without rSkMs (group 1) or containing 1.5 x 10(6) non-PC (group 2) or PC (group 3) rSkMs. Real-time PCR for sry gene 4 days after transplantation (n=4/group) showed 1.93-fold higher survival of rSkMs in group 3 as compared with group 2. Four weeks later, echocardiography revealed improved indices of left ventricular function, including ejection fraction and fractional shortening in group 3 (P<0.02) as compared with groups 1 and 2. Blood vessel count per surface area (at x400 magnification) was highest in scar and periscar areas in group 3 as compared with the other groups (P<0.05). We conclude that activation of signaling pathways of preconditioning in SkMs promoted their survival by release of paracrine factors to promote angiomyogenesis in the infarcted heart. Transplantation of PC SkMs for heart cell therapy is an innovative approach in the clinical perspective.