Shear stress–induced endothelial adrenomedullin signaling regulates vascular tone and blood pressure
Andras Iring, … , Lee S. Weinstein, Stefan Offermanns
Andras Iring, … , Lee S. Weinstein, Stefan Offermanns
Published July 1, 2019; First published June 17, 2019
Citation Information: J Clin Invest. 2019;129(7):2775-2791. https://doi.org/10.1172/JCI123825.
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Categories: Research Article Vascular biology

Shear stress–induced endothelial adrenomedullin signaling regulates vascular tone and blood pressure

Abstract

Hypertension is a primary risk factor for cardiovascular diseases including myocardial infarction and stroke. Major determinants of blood pressure are vasodilatory factors such as nitric oxide (NO) released from the endothelium under the influence of fluid shear stress exerted by the flowing blood. Several endothelial signaling processes mediating fluid shear stress–induced formation and release of vasodilatory factors have been described. It is, however, still poorly understood how fluid shear stress induces these endothelial responses. Here we show that the endothelial mechanosensitive cation channel PIEZO1 mediated fluid shear stress–induced release of adrenomedullin, which in turn activated its Gs-coupled receptor. The subsequent increase in cAMP levels promoted the phosphorylation of endothelial NO synthase (eNOS) at serine 633 through protein kinase A (PKA), leading to the activation of the enzyme. This Gs/PKA-mediated pathway synergized with the AKT-mediated pathways leading to eNOS phosphorylation at serine 1177. Mice with endothelium-specific deficiency of adrenomedullin, the adrenomedullin receptor, or Gαs showed reduced flow-induced eNOS activation and vasodilation and developed hypertension. Our data identify fluid shear stress–induced PIEZO1 activation as a central regulator of endothelial adrenomedullin release and establish the adrenomedullin receptor and subsequent Gs-mediated formation of cAMP as a critical endothelial mechanosignaling pathway regulating basal endothelial NO formation, vascular tone, and blood pressure.

Authors

Andras Iring, Young-June Jin, Julián Albarrán-Juárez, Mauro Siragusa, ShengPeng Wang, Péter T. Dancs, Akiko Nakayama, Sarah Tonack, Min Chen, Carsten Künne, Anna M. Sokol, Stefan Günther, Alfredo Martínez, Ingrid Fleming, Nina Wettschureck, Johannes Graumann, Lee S. Weinstein, Stefan Offermanns

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Figure 1

Gs mediates flow-induced cAMP formation and specific eNOS phosphorylation at serine 635 in BAECs.

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Gs mediates flow-induced cAMP formation and specific eNOS phosphorylatio...
(A) BAECs were pretreated with the PKA inhibitor PKI (1 μM) or solvent (control) for 20 minutes and were then kept under static conditions or were exposed to flow (15 dyn/cm2) for 30 minutes. Total and phosphorylated eNOS was determined by immunoblotting. Graphs show the densitometric evaluation (n = 3). (B–G) BAECs were transfected with scrambled (control) siRNA or siRNA directed against Gαs as indicated and were exposed to flow for 30 minutes or the indicated time periods (15 dyn/cm2 in B–E) or were incubated with isoproterenol (Isopr., 10 μM, 10 minutes) or VEGF (50 ng/ml, 10 minutes) (F and G). (B) Intracellular cAMP levels were determined (n = 4, control; n = 6, Gαs). (C) Phosphorylation of various eNOS sites was determined by LC-MS/MS under static conditions and after application of shear stress (15 dyn/cm2) for 30 minutes. Intensity values of the phoshpo-sites shown were normalized to eNOS intensity and thus protein abundance in the proteome. Bar diagrams show the fold change of the corrected ratios of flow versus static conditions (n = 2–4). Logarithmization of the y axis emphasizes the directional deviation from the static condition. (D–G) Total and phosphorylated eNOS and AKT were analyzed by immunoblotting. Graphs and bar diagrams show the densitometric evaluation (n = 3). Analysis of Gαs and GAPDH expression served as control (D). Data represent the mean ± SEM; *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, 1-way ANOVA with Tukey’s post hoc test (A, F, and G), 2-way ANOVA with Bonferroni’s post hoc test (B and E).