Figure 6.

Effects of N(G)-nitro-l-arginine methyl ester (L-NAME) on mother vessels (MV). A–C: Minipumps delivering L-NAME (134 mg/kg per day) were implanted in nude mice 5 days after ear injection with adenovirus expressing vascular endothelial growth factor (VEGF)-A (Ad-VEGF-A¹⁶⁴). Twenty-four hours later ears were harvested for histologic examination. Note MV bridging (arrows) and collapse to form glomeruloid microvascular proliferations (GMP), accompanied by occasional cell death (asterisks). Pink shading demarcates vascular lumens. D: Minipumps delivering control phosphate-buffered saline (PBS) (C) or 134 mg/kg per day of L-NAME (L) or D-NAME (D) were implanted s.c. in nude mice whose ears had been injected with Ad-VEGF-A¹⁶⁴ 5, 11, 26, or 59 days earlier. Twenty-four hours after minipump implant mice were injected i.v. with Evans blue dye (EBD) and were harvested 10 minutes later to measure vascular volumes. E: Tumors in nude mouse ears 13 days after injection of 10⁶ B16F10 melanoma cells. Mice were then left untreated, injected with aflibercept i.p., or received s.c. minipumps delivering 134 mg/kg per day L-NAME. Twenty-four hours later, mice were injected i.v. with EBD, and entire tumors were harvested with punch biopsies to measure plasma volumes. n = 10 to 39 mice per group (D); n = 8 mice per group (E). ^∗P < 0.05, ^∗∗P < 0.01 (Kruskal-Wallis multiple comparisons test) versus control (D) or versus untreated (E). Scale bars: 20 μm (A and B); 10 μm (C).