rK5 transduces apoptotic signal via an intrinsic pathway. (A) Mitochondrial membrane depolarization following 24-hour treatment of endothelial cells with rK5 in the presence or absence of VEGF. HUVECs were incubated with the ΔψM-sensitive mitochondrial dye, TMRE (40 nM), for 15 minutes prior to harvesting and immediately analyzed by FACS. The profiles of unstained and etoposide (10 μg/mL)–treated HUVECs are included with each histogram. (B) Western blot analysis of full-length caspase-7 and cleaved caspase-7 in rK5-treated HUVECs. Total cell lystates from rK5-treated HUVECs were collected at different time points and subjected to Western blot analysis. β-actin was used to normalize the amount of loaded proteins. (C) Representative Western blot analysis of cleaved caspase-3 and β-actin in rK5-treated HUVECs. Total cell lystates from rK5-treated HUVECs were collected at different time points and subjected to Western blot analysis. (D) Densitometric analysis was used to quantify the levels of caspase-3 activation. Values were normalized against β-actin and presented as fold increase compared with the basal level (0 hours of treatment). Data are shown as means ± SE.