Figure 5

Changes in endothelial cells and vessel morphology following OXi4503 treatment. Mice were treated with a single IP dose of OXi4503 (100mg/kg) at 16 days after tumor induction. Tissues were collected at one hour, 24 hours and five days after treatment. Formalin fixed liver sections were stained with anti-CD34 antibody to visualize tumor vessels. (A), Control tumor, arrow indicates a patent tumor vessel; 1hr OXi4503, arrows indicate endothelial cells rounding up and detaching from the vessel basement membrane; 1hr OXi4503, EC apoptosis, the section was doubly immunostained for CD34 and active caspase-3 (apoptosis marker), to visualise endothelial cells undergoing apoptosis (arrows); 24hrs OXi4503 center, arrow points at a totally occluded tumor vessel; 24hrs OXi4503 periphery, arrow indicates patent tumor vessel; 5 days OXi4503 , center, demonstrating regenerating tumor vessels surrounded by proliferating tumor cells; Single staining magnification scale bar=50μm, double staining magnification scale bar=25μm. (B), Enumeration of vascular endothelial cell apoptosis show significant differences between the tumor center and periphery at one and 24 hours after treatment (*P <0.001); (C), Quantification of tumor vascular changes following OXi4503 treatment. Vascular density decreased significantly in the tumor center (**P<0.0001), but not the periphery (P=0.173) at 24 hours after treatment. Tumor revascularization at day five is significantly higher compared to the untreated control both at tumor center and the periphery (*P=0.001). Results are mean values ± SEM, (n≥5). Black bars = tumor periphery, Grey bars = tumor center.