Tie1 controls angiopoietin function in vascular remodeling and inflammation
Emilia A. Korhonen, … , Kari Alitalo, Pipsa Saharinen
Emilia A. Korhonen, … , Kari Alitalo, Pipsa Saharinen
Published August 22, 2016
Citation Information: J Clin Invest. 2016;126(9):3495-3510. https://doi.org/10.1172/JCI84923.
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Research Article Angiogenesis Vascular biology

Tie1 controls angiopoietin function in vascular remodeling and inflammation

Abstract

The angiopoietin/Tie (ANG/Tie) receptor system controls developmental and tumor angiogenesis, inflammatory vascular remodeling, and vessel leakage. ANG1 is a Tie2 agonist that promotes vascular stabilization in inflammation and sepsis, whereas ANG2 is a context-dependent Tie2 agonist or antagonist. A limited understanding of ANG signaling mechanisms and the orphan receptor Tie1 has hindered development of ANG/Tie-targeted therapeutics. Here, we determined that both ANG1 and ANG2 binding to Tie2 increases Tie1-Tie2 interactions in a β1 integrin–dependent manner and that Tie1 regulates ANG-induced Tie2 trafficking in endothelial cells. Endothelial Tie1 was essential for the agonist activity of ANG1 and autocrine ANG2. Deletion of endothelial Tie1 in mice reduced Tie2 phosphorylation and downstream Akt activation, increased FOXO1 nuclear localization and transcriptional activation, and prevented ANG1- and ANG2-induced capillary-to-venous remodeling. However, in acute endotoxemia, the Tie1 ectodomain that is responsible for interaction with Tie2 was rapidly cleaved, ANG1 agonist activity was decreased, and autocrine ANG2 agonist activity was lost, which led to suppression of Tie2 signaling. Tie1 cleavage also occurred in patients with hantavirus infection. These results support a model in which Tie1 directly interacts with Tie2 to promote ANG-induced vascular responses under noninflammatory conditions, whereas in inflammation, Tie1 cleavage contributes to loss of ANG2 agonist activity and vascular stability.

Authors

Emilia A. Korhonen, Anita Lampinen, Hemant Giri, Andrey Anisimov, Minah Kim, Breanna Allen, Shentong Fang, Gabriela D’Amico, Tuomas J. Sipilä, Marja Lohela, Tomas Strandin, Antti Vaheri, Seppo Ylä-Herttuala, Gou Young Koh, Donald M. McDonald, Kari Alitalo, Pipsa Saharinen

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Figure 2

Effect of α5β1 integrin on angiopoietin-induced Tie receptor activation and downstream signaling in endothelial cells.

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Effect of α5β1 integrin on angiopoietin-induced Tie receptor activation ...
(A) Representative confocal (bottom) and FLIM (top) images of CAng1-stimulated scramble (shScr) or β1 integrin (shβ1) silenced HUVECs expressing Tie2-GFP alone (left) and with Tie1-V5 (middle and right). Scale bars: 20 μm. (B) Quantification of GFP lifetimes (ns) n(ROI) = 6 for unstimulated shScr, n(ROI) = 4 for unstimulated shβ1, n(ROI) = 12 for CAng1-stimulated shScr, n(ROI) = 16 for CAng1-stimulated shβ1, n(ROI) = 14 for ANG2-stimulated shScr, n(ROI) = 13 for ANG2-stimulated shβ1. A representative experiment is shown (n = 2). **P < 0.01; ***P < 0.001, Welch’s unequal variances t test, followed by Bonferroni’s post hoc test. Error bars indicate SD. (C–F) HUVECs were silenced using shRNA lentivirus for Scr (n = 10), α5 (n = 2), β1 (n = 6), or β3 integrin (n = 2), stimulated with CAng1 (n = 10) or VEGF (n = 1), immunoprecipitated (IP) for Tie1, Tie2, or VEGFR2, and analyzed for phosphotyrosine. The membranes were reprobed using Tie1, Tie2, and VEGFR2 antibodies. Total lysates (TL) were analyzed for Hsc70, α5, β1, or β3 integrin, Tie1, Tie2, VEGFR2, phospho-Akt (n = 2), or Akt, as indicated. (G) Quantification of FOXO1-positive nuclei relative to DAPI-positive nuclei (%) in shScr and shβ1 HUVECs stimulated with ANG1. n(cells) = 506 for unstimulated shScr, n(cells) = 283 for unstimulated shβ1 integrin, n(cells) = 443 for ANG1-stimulated shScr, n(cells) = 293 for Ang1-stimulated β1 integrin silenced cells. A representative experiment is shown (n = 2). ***P < 0.001, 1-way ANOVA followed by Tukey’s post hoc test.