We assessed the effects of protein kinase C epsilon (PKC epsilon) for improving stem cell therapy for acute myocardial infarction (AMI). Primary mesenchymal stem cells (MSCs) were harvested from rat bone marrow. PKC epsilon-overexpressed MSCs and control MSCs were transplanted into infarct border zones in a rat AMI model. MSCs and PKC epsilon distribution and expression of principal proteins involved in PKC epsilon signaling through the stromal cell-derived factor 1 (SDF-1)/CXC chemokine receptor type 4 (CXCR4) axis and the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT) pathway were analyzed by immunofluorescence and western blot 1 day after transplantation. Echocardiographic measurements and histologic studies were performed at 4 weeks after transplantation, and MSC survival, expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), transforming growth factor beta (TGF beta), cardiac troponin I (cTnI), von Willebrand factor (vWF), smooth muscle actin (SMA) and factor VIII and apoptosis in infarct border zones were assessed. Rat heart muscles retained more MSCs and SDF-1, CXCR4, PI3K and phosphorylated AKT increased with PKC epsilon overexpression 1 day after transplantation. MSC survival and VEGF, bFGF, TGF beta, cTnI, vWF, SMA and factor VIII expression increased in animals with PKC epsilon-overexpressed MSCs at 4 weeks after transplantation and cardiac dysfunction and remodeling improved. Infarct size and apoptosis decreased as well. Inhibitory actions of CXCR4 or PI3K partly attenuated the effects of PKC epsilon. Activation of PKC epsilon may improve retention, survival and differentiation of transplanted MSCs in myocardia. Augmentation of PKC epsilon expression may enhance the therapeutic effects of stem cell therapy for AMI.