Figure 3.

Microvessel density analysis.

Density of microvascular profiles was assessed automatically as described previously [31] using an AxioCam MRc connected to an AxioPhot microscope (Zeiss). Microvessel density (MVD) was defined as the percentage of microvessel area/total tumor area. All image acquisition and processing was performed using custom written macros in KS400 image analysis software (Zeiss). Sunitinib treatment resulted in significant decrease in % of microvessels in NU12 tumors (A) but no change in % of microvessels in SK-RC-52 tumors (B). Triple immunofluorescence staining of NU12 control (C) and sunitinib treated tumor (D) and SK-RC-52 control (E) and sunitinib treated tumor (F). Low proliferation of NU12 tumor cells (nuclei in blue as visualized by DRAQ5) as visualized by Ki67 staining (red) is observed, both in controls (C) as well as in sunitinib treated tumors (D). Please note decrease of endothelium (9 F1 staining) in sunitinib treated NU12 tumor (green). In SK-RC-52, Triple immunofluorescence staining revealed high proliferation of SK-RC-52 tumor cells, both in controls (E) as well as in sunitinib treated tumors (F) Necrosis was minimal. ****P < .001.