Regulation of VEGF-Induced Vascular Permeability and VEGFR2 Signaling by Thrombospondin-1
X Zhang, S Parangi, J Lawler. University of Chicago Medical Center, Chicago; Massachusetts General Hospital, Boston; Beth Israel Deaconess Medical Center, Boston
Background: VEGF is a well-established stimulator of vascular permeability and angiogenesis. VEGF-induced vascular hyperpermeability is an important characteristic of many disease states. Thrombospondin-1 (TSP-1) is a potent angiogenic inhibitor. Therapeutics based on the antiangiogenic domain of TSP-1, designated the three TSP-1 type 1 repeats (3TSR), have shown promising antiangiogenic and anti-tumor efficacy. We performed this study to characterize the regulatory effects of TSP-1 on VEGF-induced vascular permeability and VEGF receptor 2 (VEGFR2) signaling.
Design: Basal-level and VEGF-induced permeability were evaluated in TSP-1-null mice and 3TSR-treated FVB mice using the Miles assay. Lung extracts from TSP-1-null mice and 3TSR-treated mice with or without VEGF iv injection were analyzed for VEGFR2 phosphorylation at tyrosine 1173. Wild-type and/or buffer-treated mice were used as control in the above experiments. VEGFR2 phosphorylation, including time course and dose response, was further characterized in vitro using 3TSR-treated human dermal microvascular endothelial cells (HDMEC) and vascular endothelial cells isolated from TSP-1-null mice.
Results: Systemic treatment of wild-type mice with 3TSR significantly decreased VEGF-induced permeability (p<0.01). VEGF-stimulated VEGFR2 phosphorylation was also significantly decreased in lung extracts from 3TSR-treated mice (p=0.03). Moreover, 3TSR significantly decreased VEGF-stimulated VEGFR2 phosphorylation in HDMECs in culture. We further performed the Miles assay in wild-type and TSP-1-null mice. Basal levels of permeability in multiple organs were equivalent in the two strains. Surprisingly, VEGF-induced permeability was significantly decreased in TSP-1-null mice (p<0.03) as compared to the wild-type control mice. In addition, systemic treatment of TSP-1-null and wild-type mice with VEGF produced lower levels of VEGFR2 phosphorylation in lung extracts of TSP-1-null mice. Vascular endothelial cells from TSP-1-null mice also showed significantly decreased VEGFR2 phosphorylation upon VEGF treatment. Whereas the magnitude of the response to VEGF is reduced in TSP-1-null endothelial cells as compared to their wild-type counterparts, the time course and dose response were comparable.
Conclusions: VEGF-induced vascular permeability and VEGFR2 phosphorylation display a biphasic response to TSP-1 concentration in tissues and isolated endothelial cells.
Tuesday, March 10, 2009 1:00 PM
Poster Session IV # 204, Tuesday Afternoon