Induction of HUVEC apoptosis by Sph. (A) Morphological features of HUVEC apoptosis induced by Sph. HUVECs were treated without (a and c) or with (b and d) 20 μmol/L Sph for 4 hours in the presence (a and b) or absence (c and d) of growth factors. Apoptotic cells were detected using Hoechst 33258 staining and visualized with fluorescence microscopy. (B) DNA fragmentation in Sph-treated HUVECs. HUVECs were treated without (lanes a and c) or with (lanes b and d) 20 μmol/L Sph for 4 hours in the presence (a and b) and absence (c and d) of growth factors. Fragmented DNA was isolated and electrophoresed on a 2% agarose gel. DNA was then visualized with ethidium bromide staining. Lane m is a 200-bp DNA ladder.