EphB4 mediates resistance to antiangiogenic therapy in experimental glioma

Abstract

Alterations in vascular morphogenesis are hallmarks of antiangiogenesis-resistant tumor vessels. Vascular morphogenesis is regulated by ephrinB2-EphB4 system which may induce different biological effects depending on the oncological and molecular contexts. It was the aim of the current study to characterize the influence of EphB4 on tumor microcirculation after antiangiogenic treatment using different SF126 glioma models. Using an ecotropic transfection system, empty vector (pLXSN) or EphB4 (EphB4 OE ) overexpressing Phoenix-ECO cells were coimplanted with SF126 glioma cells subcutaneously (dorsal skinfold chamber, DSC) and orthotopically (cranial window, CW). Tumor volume was assessed by MRI. Intravital microscopy (IVM) allowed microcirculatory analysis (total {TVD} and functional vessel density {FVD}, diameter {D}, and permeability index {PI}) before and after antiangiogenic treatment (Sunitinib: DSC: 40 mg/kg BW, 6 days; CW: 80 mg/kg BW, 4 days). Immunohistochemistry included Pecam–Desmin, Ki67, TUNEL, and Caspase 3 stainings. EphB4 OE induced large and treatment-resistant tumor vessels ( FVD: Control / Su: 110 ± 23 cm / cm 2 vs. EphB4 OE / Su: 103 ± 42 cm / cm 2 ). Maintenance of pericyte–endothelial cell interactions ( Control: 80 ± 12 vs. Control / Su: 47 ± 26%; EphB4 OE : 88 ± 9 vs. EphB4 OE / Su: 74 ± 25% ) and reduced antiproliferative ( Control: 637 ± 80 vs. Control / Su: 110 ± 22; EphB4 OE : 298 ± 108 vs. EphB4 OE / Su: 213 ± 80 ) and proapoptotic responses ( Control: 196 ± 25 vs. Control / Su: 404 ± 60; EphB4 OE : 183 ± 20 vs. EphB4 OE / Su: 270 ± 66 ) were observed under EphB4 overexpression. EphB4 overexpression leads to vascular resistance by altering vascular morphogenesis, pericyte coverage, and cellular proliferation/apoptosis in experimental SF126 glioma models.