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.
View: Text | PDF
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

×

Figure 7

ANG-2 binds to αvβ3, αvβ5, and α5β1 integrins with lower affinity than TIE2 receptor.

Options: View larger image (or click on image) Download as PowerPoint
ANG-2 binds to αvβ3, αvβ5, and α5β1 integrins with lower affinity than T...
(A) ELISA quantification of ANG-2 adhesion to TIE2 or integrins at physiological pH or in an acidic environment. ANG-2 bound in an acidic environment with significantly higher affinity to TIE2 compared with the integrin heterodimers (*P < 0.05; n = 3). (B–D) HUVEC monolayers were stimulated with Mn2+ or ANG-2. The integrin conformation was studied with β1 integrin active conformation antibodies (HUTS-21, 9EGF) (40, 41). The mean intensity was determined and the relative conformation expression quantified (n = 3). In contrast to Mn2+ stimulation, ANG-2 did not induce conformational changes in β1 integrins. (E) Effect of Mn2+ on HUVEC adhesion to immobilized ANG-2. HUVECs adhered to ANG-2–coated plates with or without Mn2+. Adherent cells were visualized with crystal violet. (F) Competition binding of ANG-2 and RGD-containing proteins. After preincubation of ECs with ANG-2, cells adhered to plates coated with fibronectin (FN) or vitronectin (VN). Adherent cells were visualized with crystal violet. Preincubation with ANG-2 did not reduce binding to RGD-containing FN or VN but enhanced HUVEC adhesion to FN (*P < 0.05; n = 3). (G) Antibody blocking of control transduced and TIE2-silenced HUVEC adhesion to ANG-2. HUVECs were preincubated with the indicated integrin heterodimer antibodies (cocktail: combination of αvβ3, αvβ5, and α5β1 integrin antibodies), followed by adhesion to ANG-2. Adherent cells were visualized with crystal violet. Antibodies against αvβ3, α5β1, and the integrin cocktail inhibited HUVEC adhesion to the ANG-2 matrix to baseline levels in TIE2lo ECs (*P < 0.05; n = 3).