The development of new blood vessels is a complex process, likely requiring the synergy of multiple angiogenic mediators. This study focuses on the proximal angiogenic response using the platelet as a complex carrier of critical mediators of angiogenesis. Platelet levels are controlled by circulating levels of thrombopoietin (TPO) functioning to activate megakaryocyte differentiation and platelet release through the c-mpl receptor. We hypothesized that TPO gene transfer should enhance correction of experimental ischemia by providing increased levels of platelets and hence platelet-derived mediators of angiogenesis. To evaluate this hypothesis, we dissected the role of the TPO-c-mpl-megakaryocyte-platelet pathway in the angiogenic response using a model of acute hindlimb ischemia of wild-type, TPO^-/-, and c-mpl^-/- mice. The data demonstrate that infusion of platelets will enhance the angiogenic response in wild-type mice and that the endogenous angiogenic response is blunted in TPO^-/- and c-mpl^-/- mice. Consistent with this observation, adenovirus (Ad)-mediated transfer of TPO (AdTPO) enhanced the correction of ischemia in wild-type and TPO^-/-, but not c-mpl^-/-, mice. Local versus systemic administration of AdTPO showed that the effect of TPO gene transfer was systemic, not local, and it could be replaced by gene transfer of VEGF, 1 of the many mediators of angiogenesis carried by the platelets, even in the absence of components in the TPO-c-mpl-megakaryocyte-platelet pathway.