BackgroundGlucocorticoid-mediated inhibition of angiogenesis is important in physiology, pathophysiology and therapy. However, the mechanisms through which glucocorticoids inhibit growth of new blood vessels have not been established. This study addresses the hypothesis that physiological levels of glucocorticoids inhibit angiogenesis by directly preventing tube formation by endothelial cells.Methodology/Principal FindingsCultured human umbilical vein (HUVEC) and aortic (HAoEC) endothelial cells were used to determine the influence of glucocorticoids on tube-like structure (TLS) formation, and on cellular proliferation (5-bromo-2′-deoxyuridine (BrdU) incorporation), viability (ATP production) and migration (Boyden chambers). Dexamethasone or cortisol (at physiological concentrations) inhibited both basal and prostaglandin F2α (PGF2α)-induced and vascular endothelial growth factor (VEGF) stimulated TLS formation in endothelial cells (ECs) cultured on Matrigel, effects which were blocked with the glucocorticoid receptor antagonist RU38486. Glucocorticoids had no effect on EC viability, migration or proliferation. Time-lapse imaging showed that cortisol blocked VEGF-stimulated cytoskeletal reorganisation and initialisation of tube formation. Real time PCR suggested that increased expression of thrombospodin-1 contributed to glucocorticoid-mediated inhibition of TLS formation.Conclusions/SignificanceWe conclude that glucocorticoids interact directly with glucocorticoid receptors on vascular ECs to inhibit TLS formation. This action, which was conserved in ECs from two distinct vascular territories, was due to alterations in cell morphology rather than inhibition of EC viability, migration or proliferation and may be mediated in part by induction of thrombospodin-1. These findings provide important insights into the anti-angiogenic action of endogenous glucocorticoids in health and disease.

BackgroundRecent clinical trials of VEGF inhibitors have shown promise in the treatment of recurrent glioblastomas (GBM). However, the survival benefit is usually short-lived as tumors escape anti-VEGF therapies. Here we tested the hypothesis that Platelet Derived Growth Factor-C (PDGF-C), an isoform of the PDGF family, affects GBM progression independent of VEGF pathway and hinders anti-VEGF therapy.Principal FindingsWe first showed that PDGF-C is present in human GBMs. Then, we overexpressed or downregulated PDGF-C in a human GBM cell line, U87MG, and grew them in cranial windows in nude mice to assess vessel structure and function using intravital microscopy. PDGF-C overexpressing tumors had smaller vessel diameters and lower vascular permeability compared to the parental or siRNA-transfected tumors. Furthermore, vessels in PDGF-C overexpressing tumors had more extensive coverage with NG2 positive perivascular cells and a thicker collagen IV basement membrane than the controls. Treatment with DC101, an anti-VEGFR-2 antibody, induced decreases in vessel density in the parental tumors, but had no effect on the PDGF-C overexpressing tumors.ConclusionThese results suggest that PDGF-C plays an important role in glioma vessel maturation and stabilization, and that it can attenuate the response to anti-VEGF therapy, potentially contributing to escape from vascular normalization.

Endothelial cells of blood vessels in tumors may be thin, fragile, and defective in barrier function. We found previously that the endothelium of vessels in human colon carcinoma xenografts in mice is a mosaic structure. Approximately 85% of tumor vessels have uniform CD31 and/or CD105 immunoreactivity, but the remainder have focal regions that lack these common endothelial markers. The present study assessed the ultrastructure of the vessel lining and the integrity of the basement membrane in these regions. Using immunolabeling and confocal microscopy, we identified blood vessels that lacked CD31 and CD105 immunoreactivity and then analyzed the ultrastructure of these vessels by transmission electron microscopy. Eleven percent of vessels in orthotopic tumors and 24% of vessels in ectopic tumors had defects in CD31 and CD105 staining measuring on average 10.8 Mm (range, 1-41.2 Mm). Ultrastructural studies identified endothelial cells at 92% of CD31-and CD105-negative sites in orthotopic tumors and 70% of the sites in ectopic tumors. Thus, most regions of tumor vessels that lack CD31 and CD105 immunoreactivity represent attenuated endothelial cells with abnormal expression of endothelial cell markers, but some are gaps between endothelial cells. More than 80% of the defects lacked immunoreactivity for multiple basement membrane proteins. (Cancer Res 2005; 65(13): 5740-9)