Fig. 4

p53 inactivation in NRP-1-mediated EC survival. p53 is involved in NRP-1-mediated EC survival in zebrafish. Fluorescent image of embryos injected with indicated morpholinos and subjected to the antibody stain with anti-GFP antibody and TUNEL assay to detect apoptosis; Insets show a magnification of apoptotic foci in the vessel. The arrows indicate apoptotic EC. i. Control; ii. zNRP-1a/1b MOs (4.5 ng+4.5 ng); iii. zNRP-1a/1b MOs (4.5 ng+4.5 ng)+zp53 MO (9.0 ng); iv. zMdm2 MO (4.5ng); v. zMdm2 MO (4.5 ng)+zp53 MO (9.0 ng). b. NRP-1/EGNP-1-mediated p53 inactivation. EGNP-1 transduced HUVEC or PAEC/NRP-1 were subjected to EGF or VEGF for the indicated times and their lysates were immunoblotted for p53 with β-actin as a loading control. c. p53 is the downstream signaling molecule of PI3K/Akt in EGNP-1-mediated EC survival. HUVEC were transfected with EGNP-1 then treated with Ly294002 or co-transfected with EGNP-1 and Akt(DN). Immunoblotting was performed to determine p53 expression level. d. Inhibition of mitochondrial apoptotic pathway in EGNP-1-mediated EC survival. Immunoblotting was carried out in EGNP-1 transduced HUVEC stimulated with EGF at indicated times to assess the activation of caspase-9 and caspase-3. p53 and caspases were quantified as a ratio to the amount of β-Actin *p<0.001, **p<0.005, ***p<0.01, ****p<0.05 in a Student's t test.