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 2

Tip cells have weak TIE2, but strong ANG-2 expression.

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Tip cells have weak TIE2, but strong ANG-2 expression. (A and B) TIE2 ex...
(A and B) TIE2 expression in lectin- and TIE2-double-stained retinas from adult mice (A) and postnatal mouse pups (B) (n = 3 each; arrows: TIE2-positive ECs; arrowheads: TIE2-negative ECs). (C) Specimen of human melanoma (n = 6) and healthy skin specimens (n = 3) were stained for CD34 and TIE2. The resting vasculature in the control specimen uniformly coexpressed TIE2 and CD34 (arrows, top row). In contrast, TIE2-positive (arrows) and TIE2-negative ECs (arrowheads) were detected in the vasculature of melanomas (bottom row). (D) Co-localization of CD34 and TIE2 in the spheroidal EC xenografting assay (30, 31). Lumenized vessels stained positive for TIE2 (arrow indicating yellow co-localization; left). ECs in non-lumenized vascular structures stained for CD34 but not for TIE2 (arrowhead, right). (E) Comparative TIE2 expression of migrating and confluent HUVECs (scratch wound assay; image shown in pseudocolors; n = 3). (F) Quantitative assessment of mean TIE2 expression in migrating and confluent ECs (*P < 0.05). (G) FACS analysis of confluent and subconfluent ECs for uPAR and TIE2 expression. (H) Quantitative assessment of the mean EC subpopulation of TIE2-negative and uPAR-positive ECs under non-permeating conditions. sub, subonfluent; con, confluent. (I) Abundant Ang2 mRNA expression in tip cells of the developing retina. Whole mount retinas were analyzed by in situ hybridization against Ang2, followed by immunoreactivity against collagen IV. Merged signals were pseudocolored using Adobe Photoshop CS software. Scale bars: A, 75 μm; B, 300 μm (insets, 50 μm); C, 75 μm; D, 75 μm; E, 100 μm (insets, 20 μm); I, 20 μm.