Stimulation of DNA synthesis and suppression of apoptosis by Sph-1-P. (Left panel) Incorporation of [³H]thymidine into HUVEC DNA. HUVECs were treated with the indicated concentrations of Sph-1-P for 22 hours in the presence of 10% FCS, followed by incubation in the presence of [³H]thymidine for 2 hours. DNA-associated radioactivity was measured using a liquid scintillation counter. Columns and error bars represent the mean ± SD (n = 3). (Right panel) Inhibition of apoptosis induced by HUVEC growth factor deprivation. HUVECs were treated with indicated concentrations of Sph-1-P for 4 hours in the absence of growth factors. Apoptotic cells were detected using Hoechst 33258 staining and visualized with fluorescence microscopy. The protection percentage from apoptosis was calculated as ([apoptotic cells in the absence of growth factors] − [apoptotic cells in the presence of Sph-1-P])/ ([apoptotic cells in the absence of growth factors] − [apoptotic cells in the presence of growth factors]) × 100.