Thrombin Inhibition Prevents Endothelial Dysfunction and Reverses 20-HETE Overproduction without Affecting Blood Pressure in Angiotensin II-Induced Hypertension in Mice.

Agnieszka Kij,Lukasz Mateuszuk,Kamil Przyborowski,Boye Jensen,Pernille B Lærkegaard Hansen,Anna Bar,Brygida Marczyk,Karolina Matyjaszczyk-Gwarda,Anna Kieronska-Rudek,Stefan Chlopicki,Camilla Enggaard,Anna Tworzydlo,Agnieszka Jasztal,Bartosz Proniewski,Maria Walczak

doi:10.3390/ijms22168664

Abstract

Angiotensin II (Ang II) induces hypertension and endothelial dysfunction, but the involvement of thrombin in these responses is not clear. Here, we assessed the effects of the inhibition of thrombin activity by dabigatran on Ang II-induced hypertension and endothelial dysfunction in mice with a particular focus on NO- and 20-HETE-dependent pathways. As expected, dabigatran administration significantly delayed thrombin generation (CAT assay) in Ang II-treated hypertensive mice, and interestingly, it prevented endothelial dysfunction development, but it did not affect elevated blood pressure nor excessive aortic wall thickening. Dabigatran’s effects on endothelial function in Ang II-treated mice were evidenced by improved NO-dependent relaxation in the aorta in response to acetylcholine in vivo (MRI measurements) and increased systemic NO bioavailability (NO2− quantification) with a concomitant increased ex vivo production of endothelium-derived NO (EPR analysis). Dabigatran treatment also contributed to the reduction in the endothelial expression of pro-inflammatory vWF and ICAM-1. Interestingly, the fall in systemic NO bioavailability in Ang II-treated mice was associated with increased 20-HETE concentration in plasma (UPLC-MS/MS analysis), which was normalised by dabigatran treatment. Taking together, the inhibition of thrombin activity in Ang II-induced hypertension in mice improves the NO-dependent function of vascular endothelium and normalises the 20-HETE-depedent pathway without affecting the blood pressure and vascular remodelling.

Highlights

The endothelium constitutes a monolayer of endothelial cells (ECs) lining the inner surface of all blood vessels and is responsible for regulating the vascular tone and permeability, smooth muscle cell proliferation, blood cells adhesion, thrombotic processes, and vascular inflammation [1,2]
We demonstrated that the direct inhibition of thrombin activity by dabigatran effectively prevented the development of angiotensin II (Ang II)-induced endothelial dysfunction and endothelial inflammation, without affecting elevated blood pressure and vascular remodelling
nitric oxide (NO) bioavailability and overproduction of 20-hydroxyeicosatetraenoic acid (20-hydroxyeicosatetraenoic acids (HETEs)), which were both reversed by dabigatran treatment

Summary

Introduction

The endothelium constitutes a monolayer of endothelial cells (ECs) lining the inner surface of all blood vessels and is responsible for regulating the vascular tone and permeability, smooth muscle cell proliferation, blood cells adhesion, thrombotic processes, and vascular inflammation [1,2]. A disturbance of vascular homeostasis leads to the development of endothelial dysfunction defined as a reduction in nitric oxide (NO)-dependent vessel function [3]. The impairment of endothelial function can be a cause or a consequence of many cardiovascular diseases, including hypertension [4,5], stroke, and myocardial infarction [6]. The overactivation of RAS in hypertension is associated with the excessive generation of arachidonic acid-derived 20-hydroxyeicosatetraenoic acid (20-HETE), a strong vasoconstrictor, which potentiates systemic vascular bed responses to angiotensin II (Ang II), and impairs endothelial function [8,9]. Impairment of endothelial function is often associated with a reduction in the biosynthesis of vasodilatory epoxyeicosatrienoic acids (e.g., 14,15-EET) identified as an endothelium-derived hyperpolarising factor [10]

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Conclusion