We investigated bone marrow mononuclear cells (BM-MNCs) and peripheral blood mononuclear cells (PB-MNCs) for therapeutic angiogenesis in the ischemic hindlimb. BM-MNCs were isolated and injected into ischemic skeletal muscles in mice. Laser Doppler and histological evaluation were performed after the surgical procedure. Fifteen patients suffering from critical lower limb ischemia received subcutaneous injections of recombinant human granulocyte colony-stimulating factor (G-CSF) to mobilize progenitor cells, and PB-MNCs were harvested and transplanted directly into the ischemic limb. Endothelial cells derived from BM-MNCs were plated, then induced to form three-dimensional networks by invading a Matrigel. Four weeks after implantation of BM-MNCs, laser Doppler analysis showed that the blood flow ratio was significantly increased (0.67 +/- 0.02 vs. 0.44 +/- 0.02). Alkaline phosphatase and immunohistochemical analyses showed that capillary density was significantly increased (95.25 +/- 0.07% vs. 39.6 +/- 0.04%). Two months after implantation of PB-MNCs, in both subgroups, ankle-brachial index values, walking distance, pain scale, and transcutaneous oxygen pressure (TcO(2)) were significantly improved (p < 0.005). A total of six of 15 limb ulcers of transplanted patients were healed after cell transplantation. BM-MNC implantation was able to induce functional angiogenesis in mice ischemic hindlimb. This clinical trial shows that G-CSF-based PB-MNC transplantation is a feasible treatment for the ischemic hindlimb.