ABSTRACT
OBJECTIVE: Bioluminescence imaging (BLI) is emerging as a cost-effective, high-throughput, noninvasive, and sensitive imaging modality to monitor cell growth and trafficking. We describe the use of dynamic BLI as a noninvasive method of assessing vessel permeability during brain tumor growth. METHODS: With the use of stereotactic technique, 10 firefly luciferase-transfected GL26 mouse glioblastoma multiforme cells were injected into the brains of C57BL/6 mice (n = 80). After intraperitoneal injection of D-luciferin (150 mg/kg), serial dynamic BLI was performed at 1-minute intervals (30 seconds exposure) every 2 to 3 days until death of the animals. The maximum intensity was used as an indirect measurement of tumor growth. The adjusted slope of initial intensity (I90/Im) was used as a proxy to monitor the flow rate of blood into the vascular tree. Using a modified Evans blue perfusion protocol, we calculated the relative permeability of the vascular tree at various time points. RESULTS: Daily maximum intensity correlated strongly with tumor volume. At postinjection day 23, histology and BLI demonstrated an exponential growth of the tumor mass. Slopes were calculated to reflect the flow in the vessels feeding the tumor (adjusted slope = I90/Im). The increase in BLI intensity was correlated with a decrease in adjusted slope, reflecting a decrease in the rate of blood flow as tumor volume increased (y = 93.8e-0.49, R2 = 0.63). Examination of calculated slopes revealed a peak in permeability around postinjection day 20 (n = 42, P < .02 by 1-way analysis of variance) and showed a downward trend in relation to both postinjection day and maximum intensity observed; as angiogenesis progressed, tumor vessel caliber increased dramatically, resulting in sluggish but increased flow. This trend was correlated with Evans blue histology, revealing an increase in Evans blue dye uptake into the tumor, as slope calculated by BLI increases. CONCLUSION: Dynamic BLI is a practical, noninvasive technique that can semiquantitatively monitor changes in vascular permeability and therefore facilitate the study of tumor angiogenesis in animal models of disease.
Subject(s)
Brain Neoplasms/complications , Glioblastoma/complications , Luciferases, Firefly , Neovascularization, Pathologic/diagnosis , Neovascularization, Pathologic/etiology , Animals , Cell Line, Tumor , Diagnostic Imaging/methods , Disease Models, Animal , Dose-Response Relationship, Drug , Evans Blue , Luciferases, Firefly/genetics , Luminescence , Mice , Mice, Inbred C57BL , Neoplasm Transplantation , Platelet Endothelial Cell Adhesion Molecule-1/metabolism , Time Factors , Transfection/methodsABSTRACT
BACKGROUND: Statins are thought to have tumorolytic properties, reducing angiogenesis by inhibiting pro-angiogenic factors and inducing apoptosis of mural pericytes within the tumor vascular tree. MATERIALS AND METHODS: An orthotopic mouse glioblastoma (GL-26) model was used to investigate the effect of simvastatin on glioblastoma vasculature in vivo. GL-26 cells were implanted into the striatum of C5LKa mice treated with either control, low- or high-dose simvastatin. Brains were analyzed for necrotic volume, apoptosis, morphology and pericytic cells within the vascular tree. RESULTS: Low-dose simvastatin increased necrosis and apoptosis compared to both control and high-dose simvastatin groups. High-dose simvastatin increased vessel caliber by reducing pericytic cells along the tumor vessel wall compared to both control and low-dose simvastatin groups. CONCLUSION: Simvastatin has a dual effect on tumorigenesis. At high doses, it may worsen instead of 'normalizing' tumor angio-architecture, albeit low doses affect tumor cell survival by promoting necrosis and apoptosis.
