Anti-inflammatory, anti-proliferative and anti-atherosclerotic effects of quercetin in human in vitro and in vivo models - ScienceDirect

Atherosclerosis

Volume 218, Issue 1, September 2011, Pages 44-52

Atherosclerosis

https://doi.org/10.1016/j.atherosclerosis.2011.04.023 Get rights and content

Abstract

Objective

Polyphenols such as quercetin may exert several beneficial effects, including those resulting from anti-inflammatory activities, but their impact on cardiovascular health is debated. We investigated the effect of quercetin on cardiovascular risk markers including human C-reactive protein (CRP) and on atherosclerosis using transgenic humanized models of cardiovascular disease.

Methods

After evaluating its anti-oxidative and anti-inflammatory effects in cultured human cells, quercetin (0.1%, w/w in diet) was given to human CRP transgenic mice, a humanized inflammation model, and ApoE*3Leiden transgenic mice, a humanized atherosclerosis model. Sodium salicylate was used as an anti-inflammatory reference.

Results

In cultured human endothelial cells, quercetin protected against H₂O₂-induced lipid peroxidation and reduced the cytokine-induced cell-surface expression of VCAM-1 and E-selectin. Quercetin also reduced the transcriptional activity of NFκB in human hepatocytes. In human CRP transgenic mice (quercetin plasma concentration: 12.9 ± 1.3 μM), quercetin quenched IL1β-induced CRP expression, as did sodium salicylate. In ApoE*3Leiden mice, quercetin (plasma concentration: 19.3 ± 8.3 μM) significantly attenuated atherosclerosis by 40% (sodium salicylate by 86%). Quercetin did not affect atherogenic plasma lipids or lipoproteins but it significantly lowered the circulating inflammatory risk factors SAA and fibrinogen. Combined histological and microarray analysis of aortas revealed that quercetin affected vascular cell proliferation thereby reducing atherosclerotic lesion growth. Quercetin also reduced the gene expression of specific factors implicated in local vascular inflammation including IL-1R, Ccl8, IKK, and STAT3.

Conclusion

Quercetin reduces the expression of human CRP and cardiovascular risk factors (SAA, fibrinogen) in mice in vivo. These systemic effects together with local anti-proliferative and anti-inflammatory effects in the aorta may contribute to the attenuation of atherosclerosis.

Introduction

Cardiovascular disease (CVD) is an important health problem and one of the main causes of morbidity and mortality in industrialized countries. Specific dietary patterns are positively associated with cardiovascular risk factors [1] and the development of atherosclerosis [2]. For example, diets with a high saturated fat content coincide with elevated levels of circulating C-reactive protein (CRP), a sensitive inflammatory marker predictive of future CVD [3], and with an increased incidence of cardiovascular events [1], [2]. On the other hand, diets rich in fruit and vegetables are inversely related to inflammatory CVD markers and atherosclerosis [1], [2]. A clear understanding of the mechanisms underlying the protective effects of food constituents has not been reached thus far, and the role of individual dietary factors remains enigmatic.

Among the dietary constituents that may influence development of CVD are polyphenols, and epidemiological studies of diet-disease relationships support a possible protective role [4], [5]. However, controlled human studies testing the effect of specific polyphenol-rich foods on circulating risk factors for CVD (e.g. CRP, fibrinogen) are not consistent, with some studies showing reductions in risk factor concentrations [6], [7], while others report a lack of effect [8].

Quercetin is a potent dietary polyphenol that can exert anti-inflammatory, anti-proliferative and anti-oxidative effects [9], [10], [11]. The combination of these putatively beneficial properties makes quercetin a most promising ‘nutriceutical’ for CVD prevention. However, a substantial part of the experimental evidence that suggests health effects comes from in vitro studies and short-term in vivo studies, that often require relatively high doses of compound and lack clear-cut biological endpoints.

In this study, we evaluated quercetin in cultured human cells and two humanized models of CVD, with particular emphasis on the inflammatory aspects of the disease. In cell culture, we have analyzed the anti-oxidative and anti-inflammatory properties of quercetin. In these assays, vitamin E (α-tocopherol) and genistein served as an anti-oxidative and anti-inflammatory reference control, respectively. Direct anti-inflammatory effects of quercetin in vivo were investigated in mice expressing the human CRP gene, both before and after cytokine administration. Of note, the murine form of CRP is not an acute phase reactant or inflammatory marker. Long-term effects of quercetin on the development of atherosclerosis were investigated in human ApoE*3Leiden transgenic mice (ApoE*3L), a model with humanized lipoprotein metabolism [12]. Sodium salicylate served as an anti-inflammatory reference control in the in vivo experiments.

We confirm the putative direct anti-inflammatory effects of quercetin in short-term in vitro and in vivo experiments and show that long-term quercetin treatment attenuates atherosclerotic lesion formation. Bioinformatical pathway analysis of aortic microarray gene expression data revealed that quercetin affects vascular cell proliferation and inflammatory state.

Section snippets

Endothelial cell culture assay for quantifying oxidative stress

Oxidative stress was determined essentially as described [13]. Briefly, endothelial cells from human umbilical veins (HUVEC) were isolated and confluent HUVEC monolayers of passage 1 or 2 were used. The hydrogen peroxide (H₂O₂)-induced oxidative stress was assessed with a fluorescence assay using fluorophore C11-BODIPY^581/591. HUVEC were incubated for 30 min at 37 °C with C11-BODIPY^581/591 (1 μmol/L). Subsequently, cells were washed with PBS and oxidative stress was induced with PBS containing 5

Quercetin exerts anti-oxidative and anti-inflammatory effects in cultured human cells

The anti-oxidative and anti-inflammatory capacity of quercetin was first evaluated in human cell culture models and was analyzed together with two reference control compounds with established anti-oxidative (vitamin E) or anti-inflammatory (genistein) effects.

To evaluate its anti-oxidative capacity, we measured the effect of quercetin on H₂O₂-induced lipid peroxidation using fluorophore C-11 BODIPY^581/591. Pre-incubation of HUVEC with quercetin for 18 h dose-dependently (1-100 μmol/L) protected

Discussion

Polyphenols such as quercetin have become the subject of considerable attention, mainly because of their broad spectrum of putatively beneficial effects for vascular health (‘nutriceutical’). Here, we focused on the anti-inflammatory and anti-atherogenic effects of the polyphenol quercetin. Using human CRP transgenic mice, we show that short-term (14 days) treatment with dietary quercetin fully quenches the cytokine-induced expression of human CRP, a sensitive inflammation marker and risk

Funding

This work was supported by the Nederlandse Organisatie voor Toegepast Wetenschappelijk Onderzoek (TNO) research project ‘New Initiative Systems Biology/Personalized Health’.

Author contribution

LV and SZ carried out the in vivo experiments. SZ, PW, and LV were responsible for data collection and analysis. MvE analyzed the micro-array gene expression data. RK, MM and TK designed the study, analyzed data and wrote the manuscript.

Conflict of interest

The authors declare to have no conflict of interest.

Acknowledgements

We thank Karin Toet, Erik Offerman and Annie Jie for excellent technical assistance.

References (48)

J.A. Nettleton et al.
Dietary patterns are associated with biochemical markers of inflammation and endothelial activation in the Multi-Ethnic Study of Atherosclerosis (MESA)
Am J Clin Nutr
(2006)
J.A. Nettleton et al.
Dietary patterns and incident cardiovascular disease in the Multi-Ethnic Study of Atherosclerosis
Am J Clin Nutr
(2009)
T.L. Zern et al.
Cardioprotective effects of dietary polyphenols
J Nutr
(2005)
P.C. Hollman et al.
Dietary flavonol intake may lower stroke risk in men and women
J Nutr
(2010)
A.W. Boots et al.
Health effects of quercetin: from antioxidant to nutraceutical
Eur J Pharmacol
(2008)
R. Kleemann et al.
Fibrates down-regulate IL-1-stimulated C-reactive protein gene expression in hepatocytes by reducing nuclear p50-NFkappa B-C/EBP-beta complex formation
Blood
(2003)
B. Haslinger et al.
Simvastatin suppresses tissue factor expression and increases fibrinolytic activity in tumor necrosis factor-alpha-activated human peritoneal mesothelial cells
Kidney Int
(2003)
D. Rein et al.
Transgenic flavonoid tomato intake reduces C-reactive protein in human C-reactive protein transgenic mice more than wild-type tomato
J Nutr
(2006)
V.C.J. de Boer et al.
Tissue distribution of quercetin in rats and pigs
J Nutr
(2005)
L. Verschuren et al.
LXR agonist suppresses atherosclerotic lesion growth and promotes lesion regression in apoE*3Leiden mice: time course and mechanisms
J Lipid Res
(2009)

R. Kleemann et al.

Evidence for anti-inflammatory activity of statins and PPARalpha activators in human C-reactive protein transgenic mice in vivo and in cultured human hepatocytes in vitro

Blood

(2004)

G.J. Blake et al.

C-reactive protein and other inflammatory risk markers in acute coronary syndromes

J Am Coll Cardiol

(2003)

O.K. Chun et al.

Serum C-reactive protein concentrations are inversely associated with dietary flavonoid intake in U.S. adults

J Nutr

(2008)

E. Sacanella et al.

Down-regulation of adhesion molecules and other inflammatory biomarkers after moderate wine consumption in healthy women: a randomized trial

Am J Clin Nutr

(2007)

B.H. Kim et al.

Quercetin 3-O-beta-(2”-galloyl)-glucopyranoside inhibits endotoxin LPS-induced IL-6 expression and NF-kappa B activation in macrophages

Cytokine

(2007)

J.Y. Yang et al.

Enhanced inhibition of adipogenesis and induction of apoptosis in 3T3-L1 adipocytes with combinations of resveratrol and quercetin

Life Sci

(2008)

V. Noe et al.

Epicatechin and a cocoa polyphenolic extract modulate gene expression in human Caco-2 cells

J Nutr

(2004)

T.L. Zern et al.

Grape polyphenols exert a cardioprotective effect in pre- and postmenopausal women by lowering plasma lipids and reducing oxidative stress

J Nutr

(2005)

J.S. Choi et al.

Flavones mitigate tumor necrosis factor-alpha-induced adhesion molecule upregulation in cultured human endothelial cells: role of nuclear factor-kappa B

J Nutr

(2004)

S.B. Lotito et al.

Dietary flavonoids attenuate tumor necrosis factor alpha-induced adhesion molecule expression in human aortic endothelial cells. Structure-function relationships and activity after first pass metabolism

J Biol Chem

(2006)

C. Boesch-Saadatmandi et al.

Effect of quercetin and its metabolites isorhamnetin and quercetin-3-glucuronide on inflammatory gene expression: role of miR-155

J Nutr Biochem

(2011)

M. Karakas et al.

CRP in cardiovascular disease

Herz

(2009)

V. Konstantinidou et al.

In vivo nutrigenomic effects of virgin olive oil polyphenols within the frame of the Mediterranean diet: a randomized controlled trial

FASEB J

(2010)

A. Karlsen et al.

Bilberry juice modulates plasma concentration of NF-kappaB related inflammatory markers in subjects at increased risk of CVD

Eur J Nutr

(2010)

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