Figure 4
Adenoviral COMP-Ang1 gene or COMP-Ang1 protein transfer induces eNOS phosphorylation and increases cGMP concentration.
(a–c) eNOS phosphorylation and cGMP expression in corpus cavernosum tissue. (a) Western blot analysis demonstrating the relative abundance of phospho-eNOS (p-eNOS, Ser1177) in age-matched control (C) or hypercholesterolemic mice 7 days after intracavernous injection of ad-LacZ (L, 2 × 108 parts/20 μl), BSA (B, 5.88 μg/20 μl), ad-COMP-Ang1 (CA, 2 × 108 parts/20 μl), COMP-Ang1 protein (CP, 5.88 μg/20 μl), or cholesterol diet only (CH). Data are representative of four independent experiments. The ratios indicate the relative ratio of p-eNOS to eNOS as determined by densitometry. The relative ratio of the control group (C) was arbitrarily set equivalent to 1. Kruskal-Wallis test was used for statistical analysis. *p <0.01 vs C, CA and CP groups, #p <0.01 vs CH, L and B groups. (b) Immunohistochemical staining of cavernous tissue using antibodies to PECAM-1 (CD31; red) and p-eNOS (green) in age-matched control or hypercholesterolemic mice 8 weeks after intracavernous injection of each gene or protein. Results were similar for three independent experiments. Scale bar = 100 μm. (c) Cavernous cGMP concentrations. Each bar depicts the mean ± SEM for n = 4 animals per group. One-way ANOVA was used for statistical analysis. *p <0.01 vs C, CA and CP groups, †p <0.01 vs C group, ‡p <0.01 vs C, CA and CP groups, #p <0.01 vs CH, L and B groups. (d–g) eNOS phosphorylation and cGMP expression in HUVECs and MCECs. The cells were incubated with COMP-Ang1 protein (400 ng/ml/d) for three consecutive days and cells were harvested 24 hours after treatment. (d, f) Western blot analysis for p-eNOS and eNOS. Data are the mean ± SEM of three independent experiments. Mann-Whitney U-test was used for statistical analysis. *p <0.01 vs control group. (e, g) cGMP concentrations in HUVECs and MCECs. Each bar represents the mean ± SEM for n = 6 per group. Mann-Whitney U-test was used for statistical analysis. *p <0.05 vs control group.
