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Phytomedicine 13 (2006) 442–445

www.elsevier.de/phymed

Vascular effects of 7-epiclusianone, a prenylated benzophenone from

Rheedia gardneriana, on the rat aorta

A.J. Cruza, V.S. Lemosb, M.H. dos Santosc, T.J. Nagemd, S.F. Cortesa,�

aDepartamento de Farmacologia, Universidade Federal de Minas Gerais, Belo Horizonte-MG, BrazilbDepartamento de Fisiologia e Biofısica, Universidade Federal de Minas Gerais, Belo Horizonte-MG, BrazilcDepartamento de Farmacia – Escola de Farmacia e Odontologia de Alfenas/Ceufe, Alfenas-MG, BrazildDepartamento de Quımica, Universidade Federal de Ouro Preto, Ouro Preto-MG, Brazil

Abstract

The vascular effects of 7-epiclusianone on the rat aorta were investigated. In the rat aortic rings with functionalendothelia, 7-epiclusianone up to 10 mM induced a concentration-dependent vasodilatation of the sustainedcontractions induced by phenylephrine (0.3 mM). At concentrations higher than 10 mM, 7-epiclusianone induced aconcentration-dependent contraction in the aortic rings. The vasodilator effect of 7-epiclusianone was drasticallydecreased with L-NAME (100 mM) as well as in endothelium-denuded aortic rings. Moreover, indomethacin (10 mM)induced a significant shift to the left in the vasodilator but did not modify the vasoconstrictor effect of 7-epiclusianone.In arteries without pre-contraction, 7-epiclusianone (3–100 mM) induced concentration-dependent contraction only inendothelium-intact and in the presence of L-NAME (100 mM). This effect was inhibited by indomethacin (10 mM) andZM230487 (1 mM), selective inhibitors of cyclooxygenase and of 5-lipoxygenase, respectively. We can conclude that atlow concentrations 7-epiclusianone induces an endothelium-dependent vasodilator effect in rat aortic rings. At higherconcentrations and in conditions where NO synthase was inhibited, 7-epiclusianone induces a vasocontractile effect.Nitric oxide seems to participate in the vasodilatation, while endothelial cyclooxygenase- and 5-lipoxygenase-derivedproducts play a role in the vasoconstrictor effect.r 2005 Elsevier GmbH. All rights reserved.

Keywords: Rheedia gardneriana; Benzophenone; Endothelium; Nitric oxide; Lipoxygenase; Cyclooxygenase

Introduction

Polyprenylated benzophenones have been describedas antioxidant, free radical scavengers; as antiulceragents; as inhibitors of iNOS and COX-2 expression incolon carcinoma; and as apoptosis inducers (Pan et al.,2001; Tanaka et al., 2000; Yamaguchi et al., 2000a, b),demonstrating a broad spectrum of biological activities

e front matter r 2005 Elsevier GmbH. All rights reserved.

ymed.2005.01.014

ing author. Tel.: +55 31 3499 2726;

9 2695.

ess: sfcortes@icb.ufmg.br (S.F. Cortes).

for this class of compounds. The association ofantioxidant and anti-inflammatory effect with vasodila-tation would be relevant for the treatment of cardio-vascular illnesses such as atherosclerosis and ischemia.

7-epiclusianone (Fig. 1) is a prenylated benzophenoneisolated as the main constituent from Rheedia gardneri-

ana Miers ex Planch & Triana (Santos et al., 1998). Thebiological activities of this substance remain poorlyinvestigated. There is only one previous report demon-strating in vitro activity of 7-epiclusianone againsttrypomastigotes of Trypanosoma cruzi (Alves et al.,1999). In the present work, we began the investigation

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OO

OHO

CH3

CH3

H3C

CH3

H3C

CH3

CH3

CH3

Fig. 1. Chemical structure of 7-epiclusianone.

A.J. Cruz et al. / Phytomedicine 13 (2006) 442–445 443

of the effects of 7-epiclusianone on the vascularsystem.

Materials and methods

Experimental procedure

Animal experiments were performed according to therecommendations of the Brazilian Council for AnimalCare and to the directives of the Ethics Committee ofthe Universidade Federal de Minas Gerais. Male Wistarrats (200–250 g each) were killed by cervical dislocationand exsanguinated. The aortic rings were mounted andmaintained in Krebs-Henseleit solution of the followingcomposition (mM): NaCl 110.8, KCl 5.9, NaHCO3 25.0,MgSO4 1.07, CaCl2 2.49, NaH2PO4 2.33 and glucose11.51, as described by Guedes et al. (2002). 7-epiclusia-none was added cumulatively during the tonic contrac-tions induced by phenylephrine (0.3 mM) or in aorticrings without pre-contraction. The results obtained withthe vasodilator effect induced by 7-epiclusianone wereexpressed as a percentage of the decrease in maximalcontraction induced by phenylephrine, taking as 100%relaxation the point when the basal line was reached.For the contractile effects of 7-epiclusianone the resultswere expressed as mN/mm. Fifty-percent inhibitoryconcentrations (IC50 values) were calculated graphicallyfrom concentration–response curves.

Drugs

Acetylcholine chloride, L-NAME, indomethacin andPhenylephrine chloride were purchased from Sigma(USA). ZM230487 was a gift from Zeneca Pharmaceu-ticals. Pure 7-epiclusianone (99.82% by analyticalcalculation) was obtained as described previously(Santos et al., 1998). It was solubilized in a mixture of

distilled water/cremophor as a 10�2M solution anddiluted to the desired concentration with distilled waterjust before use. The final concentration of cremophornever exceeded 0.1%, which was without effect whenexposed to control preparations. Indomethacin wasdissolved in 0.5%w/v sodium bicarbonate. The othercompounds were freely dissolved in water.

Statistics

Results are expressed as the mean7s.e.m of fiveexperiments. Student’s t-test (for IC50 values) and One-Way ANOVA followed by Bonferroni’s MultipleComparison Test (for concentration-response curves)were used to analyze the results and statistical sig-nificance was determined at po0:05.

Results and discussion

In endothelium-intact aortic rings, 7-epiclusianoneinduced concentration-dependent relaxation with amaximal effect (Emax) of 61.773.3% and an IC50 valueof 8.171.5 mM (Fig. 2) in rat aortic rings pre-contractedwith phenylephrine (0.3 mM). This vasodilator effect wasdramatically reduced (Emax ¼ 25:7� 7:9%) in endothe-lium-denuded arteries (Fig. 2A). Acetylcholine, a well-known endothelium-dependent vasodilator, also in-duced a concentration-dependent vasodilator effectwith an Emax of 70.676.2% and an IC50 value of0.1570.05 mM (n ¼ 5), approximately 50 times morepotent than 7-epiclusianone (not shown). In addition,pre-treatment of the aortic rings with L-NAME(100 mM), a selective inhibitor of NO synthase at aconcentration that completely inhibited the vasodilatoreffect of acetylcholine (Rees et al., 1990), induced asignificant reduction in the vasodilator effect induced by7-epiclusianone (Fig. 2B). However, a significant shift tothe left (IC50 value ¼ 2.570.9 mM; po0:05) withoutmodification of Emax was observed in aortic rings pre-treated with indomethacin (10 mM; Fig. 2B), a selectiveinhibitor of cyclooxygenase at a concentration thatcompletely inhibited the production of cylcooxygenase-derived factors (Cole et al., 1986). The above resultsdemonstrate that 7-epiclusianone at low concentrationsinduces a vasodilator effect via a mechanism dependenton the presence of a functional endothelium and on theproduction of NO. They also suggest the participationof cyclooxygenase-derived contractile factor(s).

7-epiclusianone at concentrations higher than 10 mMinduced concentration-dependent contractions in aorticrings pre-contracted with phenylephrine (0.3 mM). In anattempt to clarify this effect, the contractile effect of 7-epiclusianone was investigated in aortic rings withoutpre-contraction. We observed that 7-epiclusianone was

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Fig. 2. Relaxant effect induced by 7-epiclusianone in rat aortic rings pre-contracted with phenylephrine (0.3 mM). (A) Effect of 7-

epiclusianone in aortic rings in the presence (control; J) and in the absence of a functional endothelium (K). (B) Effect of 7-

epiclusianone in rat aortic rings with a functional endothelium in the absence (control;J) and in the presence of L-NAME (100mM;

K) and indomethacin (10 mM; E). Results are expressed as mean7s.e.m. of five experiments. *po0:05, **po0:01 and ***po0:001compared to control curves by One-Way ANOVA.

Con

trac

tion

(mN

/mm

) 4

2

0

4

2

0

4

2

0-8 -7 -4-6 -5 -8 -7 -4-6 -5 -8 -7 -4-6 -5

log [7-epiclusianone] (M)log [7-epiclusianone] (M) log [7-epiclusianone] (M)

A B C

***

**

***

*** * ***

Fig. 3. Contractile effect induced by 7-epiclusianone in rat aortic rings. (A) Effect of 7-epiclusianone in rings with (K) and without

(E) a functional endothelium, and in rings with endothelium treated with L-NAME (100 mM; control; J). (B) Effect of 7-

epiclusianone in aortic rings with endothelium and L-NAME (100 mM; control; J) in the presence (K) of indomethacin (10 mM).

(C) Effect of 7-epiclusianone in aortic rings with endothelium and L-NAME (100 mM; control;J) in the presence (K) of ZM230487

(1 mM). Results are expressed as mean7s.e.m. of five experiments. *po0:05 and **po0:01 compared to control curves by One-Way

ANOVA.

A.J. Cruz et al. / Phytomedicine 13 (2006) 442–445444

able to induce concentration-dependent contractions atconcentrations higher than 3 mM, in rings with afunctional endothelium and in the presence of L-NAME(Fig. 3A). No significant contraction was observed inendothelium-denuded rings (Fig. 3A), suggesting thatthe contractile-effect induced by 7-epiclusianone wasdependent on the production of endothelium-derivedcontractile factors, and that this effect was masked bythe production of NO. Products derived from arachi-donic acid metabolism are well-known to induceendothelium-dependent contraction in the vasculature(Cole et al., 1986; Walch et al., 2000), for this reason weinvestigated the participation of cyclooxygenase- andlipoxygenase-derived contractile factors in the vasocon-striction induced by 7-epiclusianone. We observed thatindomethacin (10 mM) induced a significant reduction ofthe contractile curve of 7-epiclusianone (Fig. 3B). In thepresence of a selective inhibitor of 5-lipoxygenase(Negro et al., 1997), ZM230487 (1 mM), the contractileeffect of 7-epiclusianone was also significantly reduced

(Fig. 3C). The above results are indicative of theparticipation of cyclooxygenase- and 5-lipoxygenase-derived contractile factors in the vasoconstrictioninduced by 7-epiclusianone.

We can conclude that low concentrations of 7-epiclusianone induce an endothelium- and NO-depen-dent vasodilator effect in rat aortic rings. At higherconcentrations, 7-epiclusianone induces a contractileeffect where cyclooxygenase and 5-lipoxygenase-derivedproducts are involved. The vasodilator effect of 7-epiclusianone could be of interest for the treatment ofcardiovascular illnesses. However, care should be takendue to the vasoconstrictor effect observed at higherconcentrations.

Acknowledgements

To the National Research Council (CNPq; Brazil) forresearch fellowships (A.J.C., V.S.L., M.H.S., T.J.N.,

ARTICLE IN PRESSA.J. Cruz et al. / Phytomedicine 13 (2006) 442–445 445

S.F.C.) and Research foundation from Minas Gerais(FAPEMIG; Brazil) for financial support.

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