Sildenafil reduces aortic endothelial dysfunction and structural damage in spontaneously hypertensive rats: Role of NO, NADPH and COX-1 pathways

doi:10.1016/j.vph.2019.106601

Vascular Pharmacology

Volume 124, January 2020, 106601

https://doi.org/10.1016/j.vph.2019.106601 Get rights and content

Abstract

Arterial hypertension is a condition associated with endothelial dysfunction, accompanied by an imbalance in the production of reactive oxygen species (ROS) and NO. The aim of this study was to investigate and elucidate the possible mechanisms of sildenafil, a selective phosphodiesterase-5 inhibitor, actions on endothelial function in aortas from spontaneously hypertensive rats (SHR).

SHR treated with sildenafil (40 mg/kg/day, p.o., 3 weeks) were compared to untreated SHR and Wistar-Kyoto (WKY) rats. Systolic blood pressure (SBP) was measured by tail-cuff plethysmography and vascular reactivity was determined in isolated rat aortic rings. Circulating endothelial progenitor cells and systemic ROS were measured by flow cytometry. Plasmatic total antioxidant capacity, NO production and aorta lipid peroxidation were determined by spectrophotometry. Scanning electron microscopy was used for structural analysis of the endothelial surface.

Sildenafil reduced high SBP and partially restored the vasodilator response to acetylcholine and sodium nitroprusside in SHR aortic rings. Using selective inhibitors, our experiments revealed an augmented participation of NO, with a simultaneous decrease of oxidative stress and of cyclooxygenase-1 (COX-1)-derived prostanoids contribution in the endothelium-dependent vasodilation in sildenafil-treated SHR compared to non-treated SHR. Also, the relaxant responses to sildenafil and 8-Br-cGMP were normalized in sildenafil-treated SHR and sildenafil restored the pro-oxidant/antioxidant balance and the endothelial architecture.

In conclusion, sildenafil reverses endothelial dysfunction in SHR by improving vascular relaxation to acetylcholine with increased NO bioavailability, reducing the oxidative stress and COX-1 prostanoids, and improving cGMP/PKG signaling. Also, sildenafil reduces structural endothelial damage. Thus, sildenafil is a promising novel pharmacologic strategy to treat endothelial dysfunction in hypertensive states reinforcing its potential role as adjuvant in the pharmacotherapy of cardiovascular diseases.

Graphic abstract

Introduction

Arterial hypertension is the most prevalent cardiovascular risk factor, contributing to an increase in global morbidity and mortality and is estimated that 1.56 billion people will suffer from high blood pressure by 2025 ([1,59]). It is defined as a complex multifactorial condition in which about 90% of cases are classified as essential hypertension and its precise cause is unknown [4]. The development of hypertension has been associated with endothelial dysfunction of conductance and resistance vessels, characterized by oxidative stress and decreased bioavailability of relaxing factors derived from the endothelium, such as nitric oxide (NO), in large vessels and endothelium-derived hyperpolarizing factor in small vessels ([9,14]).

Endothelial cells play an important role in the preservation of the vascular function and its dysfunction is considered an important hallmark that participates in metabolic disease states and arterial hypertension [10,27]. A considerable number of studies on vascular dysfunction have been performed with spontaneously hypertensive rats (SHR), which exhibit a decreased NO bioavailability and an excessive generation of NADPH oxidase-driven reactive oxygen species (ROS) ([15,39]). Thus, SHR have been used as an important tool for medical research in experimental hypertension and vascular dysfunction as well as for the screening of antihypertensive agents [18].

Current studies have evidenced that selective phosphodiesterase type 5 (PDE5) inhibitors, among them sildenafil, display beneficial effects on altered systemic hemodynamics, endothelial dysfunction, systemic arterial hypertension and atherosclerosis, in addition to the known effects on sexual erectile dysfunction and pulmonary hypertension ([32,37,49]). Moreover, our group have published many data demonstrating that chronic treatment with sildenafil improves vascular dysfunction in renovascular hypertension and atherosclerotic apolipoprotein E-deficient murine model (apoE^−/−) [[2], [3],11,31].

However, the vascular effects and the mechanisms involved in sildenafil action in conductance vessels of SHR animals have been little evaluated and explored. Therefore, the present study was designed to determine the effects of sildenafil on the vascular dysfunction of conductance vessels of SHR, focusing the role of NADPH oxidase, cyclooxygenase-1 (COX-1), NO/cyclic GMP (cGMP), cyclic GMP-dependent protein kinase (PKG) and cyclic AMP (cAMP)-dependent protein kinase (PKA) in aortas of SHR.

Section snippets

Animals

The present study was performed in male 4-month old SHR and in age-matched Wistar-Kyoto rats (WKY), both of which were maintained in the animal care facility of the Federal University of Espirito Santo, Brazil. The rats were acclimatized and housed in individual plastic cages with a controlled temperature (22–23 °C) and light-dark cycle (12:12 h) and were fed with a standard rat chow and provided with water ad libitum. All experimental procedures were performed in accordance with the guidelines

Effects of sildenafil on SBP and vasodilator responses to acetylcholine in hypertensive rats

As expected, SHR exhibit high SBP compared with age-gender-match WKY rats. The 3-week oral treatment with sildenafil caused a significant fall in SBP of SHR of approximately 15% (Table 1).

Fig. 1 summarizes the results of the acetylcholine-induced relaxations in aortic rings from SHR and WKY groups. The maximal tension induced by phenylephrine (10 μmol/L) in these experiments was of 0.91 ± 0.10 g, 1.14 ± 0.14 g and 0.98 ± 0.16 g for WKY, SHR and SHR treated with sildenafil, respectively. There

Discussion

The present study shows, for the first time, that sildenafil, a selective PDE5 inhibitor, significantly reverses endothelial dysfunction in SHR, which is characterized by an impaired relaxation response to acetylcholine, an asymmetry in the NO/cGMP and ROS pathways and damage to endothelial surface layer. Our data provide a clear demonstration that the central mechanism for the beneficial effect of sildenafil involves repair in the vascular endothelial architecture, reduction of the oxidative

Author contribution

Contributed to the study design: MASL, TMCP and ECV.

Contributed to techniques of morphological analysis of blood vessels: MASL, VBM.

Contributed to biochemical analysis: MASL, TP

Contributed to the blind analysis of morphological changes: RA and SSM.

Contributed to the data collection using flow cytometry: RA, MASL and BPC.

Contributed to the vascular responsiveness data collection: MASL and TP.

Contributed to the data analysis, and data interpretation: MASL, BPC, TMCP, SSM, ECV and MCT.

Contributed

Declaration of Competing Interest

The authors declare no conflicts of interest.

Acknowedgements

The authors gratefully acknowledge research funding from the State Agency for the Development of Science and Technology (FAPES/Universal/Proc. 67597483) and the National Council for the Development of Science and Technology (CNPq; Ref. Grant #303001/2015-1) to ECV. State Research Foundation (FAPES Grants to ECV: CNPq/FAPES #24/2018 – PRONEx #569/2018). We greatly appreciated the open access to use the electronic microscopy facilities, headed by Dr. Breno Valentim Nogueira from the Federal

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Sildenafil reduces aortic endothelial dysfunction and structural damage in spontaneously hypertensive rats: Role of NO, NADPH and COX-1 pathways

Abstract

Graphic abstract

Introduction

Section snippets

Animals

Effects of sildenafil on SBP and vasodilator responses to acetylcholine in hypertensive rats

Discussion

Author contribution

Declaration of Competing Interest

Acknowedgements

Eur. J. Intern. Med.

LWT Food Sci. Technol.

Life Sci.

Vasc. Pharmacol.

Biochem. Pharmacol.

Am. Heart J.

Redox Biol.

J. Nutr. Biochem.

Vasc. Pharmacol.

J. Biol. Chem.

Life Sci.

Nitric Oxide

Sildenafil restores endothelial function in the apolipoprotein E knockout mouse

J. Transl. Med.

Protective effect of sildenafil on the genotoxicity and cytotoxicity in apolipoprotein E-deficient mice bone marrow cells

Lipids Health Dis.

Essential hypertension: an approach to its etiology and neurogenic pathophysiology

Int. J. Hypertens.

cGMP-dependent protein kinase expression restores contractile function in cultured vascular smooth muscle cells

J. Vasc. Res.

Effects of sildenafil on pulmonary hypertension and levels of ET-1, eNOS, and cGMP in aorta-banded rats

Exp. Biol. Med. (Maywood)

New therapeutic implications of endothelial nitric oxide synthase (eNOS) function/dysfunction in cardiovascular disease

Int. J. Mol. Sci.

Endothelial function and dysfunction: testing and clinical relevance

Circulation

Beneficial morphofunctional changes promoted by sildenafil in resistance vessels in the angiotensin II-induced hypertension model

Curr. Pharm. Biotechnol.

Sildenafil ameliorates oxidative stress and DNA damage in the stenotic kidneys in mice with renovascular hypertension

J. Transl. Med.

Inhibition of phosphodiesterase 5 restores endothelial function in renovascular hypertension

J. Transl. Med.

Roles of inflammation, oxidative stress, and vascular dysfunction in hypertension

Biomed. Res. Int.

Animal models for the study of arterial hypertension

J. Biosci.

Interactions between thromboxane A2, thromboxane/prostaglandin (TP) receptors, and endothelium-derived hyperpolarization

Cardiovasc. Res.

Sildenafil improves vascular endothelial structure and function in renovascular hypertension

Curr. Pharm. Biotechnol.

Endothelial dysfunction: a multifaceted disorder (The Wiggers Award Lecture)

Am. J. Physiol. Heart Circ. Physiol.

Chronic administration of the probiotic kefir improves the endothelial function in spontaneously hypertensive rats

J. Transl. Med.

Regulation of the expression of soluble guanylyl cyclase by reactive oxygen species

Br. J. Pharmacol.

Impaired endothelial repair capacity of early endothelial progenitor cells in prehypertension: relation to endothelial dysfunction

Hypertension.

Aliskiren improves endothelium-dependent relaxation of thoracic aorta by activating PI3K/Akt/eNOS signal pathway in SHR

Clin. Exp. Pharmacol. Physiol.