Angiopoietin-2 differentially regulates angiogenesis through TIE2 and integrin signaling
Moritz Felcht, … , Sergij Goerdt, Hellmut G. Augustin
Moritz Felcht, … , Sergij Goerdt, Hellmut G. Augustin
Published May 15, 2012
Citation Information: J Clin Invest. 2012;122(6):1991-2005. https://doi.org/10.1172/JCI58832.
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Research Article Vascular biology

Angiopoietin-2 differentially regulates angiogenesis through TIE2 and integrin signaling

Abstract

Angiopoietin-2 (ANG-2) is a key regulator of angiogenesis that exerts context-dependent effects on ECs. ANG-2 binds the endothelial-specific receptor tyrosine kinase 2 (TIE2) and acts as a negative regulator of ANG-1/TIE2 signaling during angiogenesis, thereby controlling the responsiveness of ECs to exogenous cytokines. Recent data from tumors indicate that under certain conditions ANG-2 can also promote angiogenesis. However, the molecular mechanisms of dual ANG-2 functions are poorly understood. Here, we identify a model for the opposing roles of ANG-2 in angiogenesis. We found that angiogenesis-activated endothelium harbored a subpopulation of TIE2-negative ECs (TIE2lo). TIE2 expression was downregulated in angiogenic ECs, which abundantly expressed several integrins. ANG-2 bound to these integrins in TIE2lo ECs, subsequently inducing, in a TIE2-independent manner, phosphorylation of the integrin adaptor protein FAK, resulting in RAC1 activation, migration, and sprouting angiogenesis. Correspondingly, in vivo ANG-2 blockade interfered with integrin signaling and inhibited FAK phosphorylation and sprouting angiogenesis of TIE2lo ECs. These data establish a contextual model whereby differential TIE2 and integrin expression, binding, and activation control the role of ANG-2 in angiogenesis. The results of this study have immediate translational implications for the therapeutic exploitation of angiopoietin signaling.

Authors

Moritz Felcht, Robert Luck, Alexander Schering, Philipp Seidel, Kshitij Srivastava, Junhao Hu, Arne Bartol, Yvonne Kienast, Christiane Vettel, Elias K. Loos, Simone Kutschera, Susanne Bartels, Sila Appak, Eva Besemfelder, Dorothee Terhardt, Emmanouil Chavakis, Thomas Wieland, Christian Klein, Markus Thomas, Akiyoshi Uemura, Sergij Goerdt, Hellmut G. Augustin

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Figure 1

ANG-2 blockage inhibits angiogenesis by interfering with remodeling of the stalk and phalanx cell vasculature as well as by inhibiting the sprouting tip cell EC phenotype.

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ANG-2 blockage inhibits angiogenesis by interfering with remodeling of t...
(A) Newborn mice were injected intraperitoneally with 30 μg/pup neutralizing ANG-2 antibody (red bars throughout) or IgG control antibody (blue bars) at P1, P3, and P5. Mice were sacrificed at day 6, and the enucleated eyes were processed for retinal whole mount analysis and staining with FITC-labeled lectin (scale bars: 400 μm). (B–I) Quantitative analysis of the vasculature with Fiji analysis (see Supplemental Experimental Procedures) characterizing the total vessel area (scale bar: 400 μm) (B and C), the vessel density (D), the non-vascularized area (defined as an area that was more than 40 μm away from the next vessel [green]) (scale bars: 400 μm) (E and F), the junctional branch points per retina (G), the vessel segments per retina (H), and histogram of the branch length distribution (I). For B–H, *P < 0.05. (J–L) Higher-resolution images at the edge of the vascular plexus (red arrows, tip cells; yellow arrow, filopodia) demonstrating the pronounced tip cell phenotype–inhibiting effect of the ANG-2 blocking antibody with reduced total number of tip cells/retina (K) and fewer filopodia/tip cell (L). (M–O) ANG-2 blockage inhibited tumor growth of subcutaneously growing Colo205 xenografts after anti–ANG-2 treatment (*P < 0.05, mean ± SEM, n = 20) (M). Colo205 xenograft tumor sections were stained for CD34 and analyzed by high-resolution analysis to detect intratumoral endothelial tip cells (scale bars: 50 μm) (N). The number of intratumoral tip cells was significantly reduced in the ANG-2 antibody treatment group (*P < 0.05, mean ± SEM, n = 5) (O).