Fig. 2: EphB4 is required for PTEN knockout prostate cancer initiation.
From: EphB4-ephrin-B2 are targets in castration resistant prostate cancer
a Quantitative RT-PCR of EPHB4, EPHB1, EPHB2, and EPHB3 RNA levels were measured in wild-type mice (both EPHB4 and PTEN are wild-type, blue), CPPL mice (CPPL mice with wild-type EPHB4, red), and EPHB4 homo-CPPL mice (CPPL mice with homozygous deletion of EPHB4, purple) prostate tissues. Gene expression was normalized against β-actin. Error bars represent standard deviation (s.d.). These experiments were repeated at least three times and similar results were obtained. P-value was calculated using two-tailed, unpaired Student’s t-test. *P < 0.05; **P < 0.002. b BLI of luciferase expression in live mice imaging over 3, 5, and 7 months of age. Left panel shows 3 out of 14 CPPL mice with wild-type EPHB4 (EPHB4+/+;CPPL). Right panel shows 3 out of 29 CPPL mice with homozygous EPHB4 (EPHB4–/–;CPPL). Color bars indicate intensity of luciferase signal. Blue denotes weak signal; Green and yellow denote intermediate signal; Red denotes strong signal. c Quantitative measurement of total BLI signal intensity of 14 EPHB4+/+; CPPL mice and 29 EPHB4–/–;CPPL mice. Blue: 3 months old; red: 5 months; green: 7 months. BLI intensity is labeled on the top of each bar. Error bars represent s.d. d Western blot of EphB4 was performed on tumors harvested from the prostate gland of mice. β-actin was probed to ensure equal loading. These experiments were repeated at least three times and similar results were obtained. e H&E stain of prostate glands from wild-type, EPHB4+/+;CPPL and EPHB4-/-;CPPL mice under magnifications of 100×, 200× and 400×, respectively. f IHC stain of EphB4, PTEN, pAKT, pS6 and Ki67 in prostate tissues. Scale bar = 50 µm. Signal intensities shown on right panel were quantified based on positive staining pixel dots using ImageJ (NIH) software. All error bars represent s.d. WT: wild-type both EPHB4 and PTEN; EPHB4+/+;CPPL: EPHB4 wild-type PTEN-null mice; EPHB4–/–;CPPL: EPHB4-homozygote PTEN-null mice. All experiments were repeated at least three times and similar results were obtained.
