Ras pathway inhibition prevents neovascularization by repressing endothelial cell sprouting
Peter D. Westenskow, Toshihide Kurihara, Edith Aguilar, Elizabeth L. Scheppke, Stacey K. Moreno, Carli Wittgrove, Valentina Marchetti, Iacovos P. Michael, Sudarshan Anand, Andras Nagy, David Cheresh, Martin Friedlander
Peter D. Westenskow, Toshihide Kurihara, Edith Aguilar, Elizabeth L. Scheppke, Stacey K. Moreno, Carli Wittgrove, Valentina Marchetti, Iacovos P. Michael, Sudarshan Anand, Andras Nagy, David Cheresh, Martin Friedlander
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Research Article Vascular biology

Ras pathway inhibition prevents neovascularization by repressing endothelial cell sprouting

Abstract

Vascular networks develop from a growing vascular front that responds to VEGF and other guidance cues. Angiogenesis is required for normal tissue function, but, under conditions of stress, inappropriate vascularization can lead to disease. Therefore, inhibition of angiogenic sprouting may prevent neovascularization in patients with blinding neovascular eye diseases, including macular degeneration. VEGF antagonists have therapeutic benefits but also can elicit off-target effects. Here, we found that the Ras pathway, which functions downstream of a wide range of cytokines including VEGF, is active in the growing vascular front of developing and pathological vascular networks. The endogenous Ras inhibitor p120RasGAP was expressed predominately in quiescent VEGF-insensitive endothelial cells and was ectopically downregulated in multiple neovascular models. MicroRNA-132 negatively regulated p120RasGAP expression. Experimental delivery of α-miR-132 to developing mouse eyes disrupted tip cell Ras activity and prevented angiogenic sprouting. This strategy prevented ocular neovascularization in multiple rodent models even more potently than the VEGF antagonist, VEGF-trap. Targeting microRNA-132 as a therapeutic strategy may prove useful for treating multiple neovascular diseases of the eye and for preventing vision loss regardless of the neovascular stimulus.

Authors

Peter D. Westenskow, Toshihide Kurihara, Edith Aguilar, Elizabeth L. Scheppke, Stacey K. Moreno, Carli Wittgrove, Valentina Marchetti, Iacovos P. Michael, Sudarshan Anand, Andras Nagy, David Cheresh, Martin Friedlander

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Figure 2

The Ras pathway is aberrantly active in OIR and Vldlr–/– mice.

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The Ras pathway is aberrantly active in OIR and Vldlr–/– mice. (A) In ...
(A) In P12 OIR mice, p120RasGAP (white) is detected in the retinal vessels of the superficial plexus. (B) p120RasGAP (green) was overlaid with red GS lectin to label the vasculature. (C–F) At P14 and P16, it is barely detectable in neovascular regions. (G and H) At P18, p120RasGAP is abundantly expressed in the neovascular tufts. (I) Tip cells devoid of p120RasGAP are identifiable in the deep plexus in flat-mounted P14 Vldlr–/– retinas. (J) In thick-cut cross-sections, pERK is detected in the tip cell of the developing angioma (arrow) and in cells in the inner nuclear layer, and (K) p120RasGAP is very weakly expressed in the tip cell but is detectable in the endothelial cells of the deep plexus (DP). (L) A merged file is shown in. Scale bars: 50 μm (A–I); 10 μm (J–L).