Abstract 9965: Low-Intensity Pulsed Ultrasound Stimulation Restored the Neovascularization Capacity of Endothelial Progenitor Cells in Mice

Abstract

Background Endothelial progenitor cells (EPCs) have been used for therapeutic angiogenesis to rescue the tissue with critical ischemia. However, the effects are unsatisfactory. The cause may be use of EPCs with impaired neovascularization capacity caused by risk factors of arteriosclerosis. Ultrasound stimulation provides cells with several biological effects. Accordingly, we assessed the hypothesis that ultrasound stimulation can restore the impaired neovascularization capacity of EPCs. Methods and Results We cultured peripheral blood-derived mononuclear cells obtained from healthy volunteers or patients with >two risk factors of arteriosclerosis for four days to achieve EPCs. During the culture, we daily stimulated the cells with a 1.011 MHz pulsed ultrasound through a gas permeable membrane film from the base of the culture plate. In healthy volunteers-derived EPCs, the low-intensity pulsed ultrasound stimulation (LIPUS) of 100 mW cm -2 intensity and 240 sec exposure time generated the greatest number of EPCs with maintained cell viability and augmented migration capacity in the modified Boyden-chamber assay. LIPUS to cultured EPCs significantly increased the concentration of angiopoietin 2 in the culture medium, suggesting the augmented secretion capacity. LIPUS increased expressions of phosphorylated-Akt and endothelial nitric oxide synthetase in the EPCs on the western blotting with the increased concentration of nitrogen oxides in the medium. The intramuscular injection of the atherosclerotic patients-derived EPCs in the hind-limb ischemia mouse model showed lower ischemic/non-ischemic hind-limb blood flow ratio after 14 days than the injection of healthy volunteers-derived EPCs (0.59±0.02 vs. 0.68±0.03; p<0.05, n=7, each), indicating impaired neovascularization capacity of the atherosclerotic patients-derived EPCs. LIPUS to the atherosclerotic patients-derived EPCs significantly increased the ratio from 0.59±0.02 to 0.7±0.04 (p<0.05, n=7, each), indicating restoration of the impaired neovascularization capacities. Conclusion In conclusion, LIPUS to the atherosclerotic patients-derived EPCs may be a new strategy to enhance clinical effects of therapeutic angiogenesis.