Activation of Notch signalling by soluble Dll4 decreases vascular permeability via a cAMP/PKA-dependent pathway

Boardman, Rachel; Pang, Vincent; Malhi, Naseeb; Lynch, Amy P; Leach, Lopa; Benest, Andrew V; Bates, David O.; Machado, Maria JC

Abstract

© 2019 the American Physiological Society. The Notch ligand delta-like ligand 4 (Dll4), upregulated by VEGF, is a key regulator of vessel morphogenesis and function, controlling tip and stalk cell selection during sprouting angiogenesis. Inhibition of Dll4 results in hypersprouting, nonfunctional, poorly perfused vessels, suggesting a role for Dll4 in the formation of mature, reactive, functional vessels, with low permeability and able to restrict fluid and solute exchange. We tested the hypothesis that Dll4 controls transvascular fluid exchange. A recombinant protein expressing only the extracellular portion of Dll4 [soluble Dll4 (sDll4)] induced Notch signaling in endothelial cells (ECs), resulting in increased expression of vascular-endothelial cadherin, but not the tight junctional protein zonula occludens 1, at intercellular junctions. sDll4 decreased the permeability of FITC-labeled albumin across EC monolayers, and this effect was abrogated by coculture with the γ-secretase inhibitor N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester. One of the known molecular effectors responsible for strengthening EC-EC contacts is PKA, so we tested the effect of modulation of PKA on the sDll4-mediated reduction of permeability. Inhibition of PKA reversed the sDll4-mediated reduction in permeability and reduced expression of the Notch target gene Hey1. Knockdown of PKA reduced sDLL4-mediated vascular-endothelial cadherin junctional expression. sDll4 also caused a significant decrease in the hydraulic conductivity of rat mesenteric microvessels in vivo. This reduction was abolished upon coperfusion with the PKA inhibitor H89 dihydrochloride. These results indicate that Dll4 signaling through Notch activation acts through a cAMP/PKA pathway upon intercellular adherens junctions, but not tight junctions, to regulate endothelial barrier function. NEW & NOTEWORTHY Notch signaling reduces vascular permeability through stimulation of cAMP-dependent protein kinase A.

Citation

Boardman, R., Pang, V., Malhi, N., Lynch, A. P., Leach, L., Benest, A. V., Bates, D. O., & Machado, M. J. (2019). Activation of Notch signalling by soluble Dll4 decreases vascular permeability via a cAMP/PKA-dependent pathway. AJP - Heart and Circulatory Physiology, 316(5), H1065-H1075. https://doi.org/10.1152/ajpheart.00610.2018

• Journal Article Type: Article
• Acceptance Date: Jan 22, 2019
• Online Publication Date: Apr 25, 2019
• Publication Date: May 1, 2019
• Deposit Date: Apr 1, 2019
• Publicly Available Date: Apr 26, 2020
• Journal: American Journal of Physiology-Heart and Circulatory Physiology
• Print ISSN: 0363-6135
• Electronic ISSN: 1522-1539
• Publisher: American Physiological Society
• Peer Reviewed: Peer Reviewed
• Volume: 316
• Issue: 5
• Pages: H1065-H1075
• DOI: https://doi.org/10.1152/ajpheart.00610.2018
• Keywords: delta-like ligand 4; endothelial cells; Notch; vascular permeabililty
• Public URL: https://nottingham-repository.worktribe.com/output/1507643
• Publisher URL: https://www.physiology.org/doi/full/10.1152/ajpheart.00610.2018
• Contract Date: May 20, 2019