Vasorelaxation by Amentoflavone Isolated from Selaginella tamariscina

Abstract

In the courses of in vitro screening for the vasorelaxant effect of the various extracts from medicinal plants, an ethyl acetate-soluble extract of Selaginella tamariscina was found to exhibit distinctive vasorelaxant activity. Further purifications of the extract as guided by in vitro vasorelaxant assay afforded an active biflavonoid, amentoflavone. Amentoflavone induced concentration-dependent relaxation of the phenylephrine-precontracted aorta, which disappeared by removal of functional endothelium. Pretreatment of the aortic tissues with N ^G-nitro-L-arginine methyl ester (L-NAME), methylene blue, or 1H-[1] [2] [4]-oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) inhibited the relaxation induced by amentoflavone. Amentoflavone-induced relaxations were also markedly attenuated by addition of tetraethylammonium (TEA) or verapamil. However, the relaxant effect of amentoflavone was not blocked by pretreatment with indomethacin, glibenclamide, atropine, or propranolol. Incubation of endothelium-intact aortic rings with amentoflavone increased the production of cGMP, but this effect was blocked by endothelium-denudation or pretreatment with L-NAME or ODQ. These results suggest that amentoflavone relaxes vascular smooth muscle via endothelium-dependent nitric oxide-cGMP signaling, with possible involvement of non-specific K⁺ and Ca²⁺ channels.

Abbreviations

EDRF:endothelium-derived relaxing factor

EDHF:endothelium-derived hyperpolarizing factor

NO:nitric oxide

cGMP:guanosine 3′,5′-cyclic monophosphate

DMSO:dimethyl sulfoxide

L-NAME:N ^G-nitro-L-arginine methyl ester

ODQ:1H-[1,2,4]-oxadiazole-[4,3-a]-quinoxalin-1-one

IBMX:3-isobutyl-1-methylxanthine

K_Ca:Ca²⁺-dependent K⁺ channel

K_ATP:adenosine triphosphate (ATP)-sensitive K⁺ channel

TEA:tetraethylammonium

Selaginella tamariscina - amentoflavone - vasorelaxation - nitric oxide - cGMP

References

• 1 Furchgott R F, Zawadzki J V. The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine.  Nature. 1980;  288 373-6
• 2 Jaffe E A. Physiologic functions of normal endothelial cells.  Ann N Y Acad Sci. 1985;  454 279-91
• 3 Beny J L, Brunet P C. Neither nitric oxide nor nitroglycerin accounts for all the characteristics of endothelially mediated vasodilatation of pig coronary arteries.  Blood Vessels.. 1988;  25 308-11
• 4 Palmer R M, Ashton D S, Moncada S. Vascular endothelial cells synthesize nitric oxide from L-arginine.  Nature. 1988;  333 664-6
• 5 Hirano S, Agata N, Hara Y, Iguchi H, Shirai M, Tone H, Urakawa N. Pirarubicin-induced endothelium-dependent relaxation in rat isolated aorta.  J Pharm Pharmacol. 1991;  43 848-54
• 6 Gray D W, Marshall I. Human alpha-calcitonin gene-related peptide stimulates adenylate cyclase and guanylate cyclase and relaxes rat thoracic aorta by releasing nitric oxide.  Br J Pharmacol. 1992;  107 691-6
• 7 Sato K, Ozaki H, Karaki H. Changes in cytosolic calcium level in vascular smooth muscle strip measured simultaneously with contraction using fluorescent calcium indicator fura 2.  J Pharmacol Exp Ther. 1988;  246 294-300
• 8 Lee H S, Oh W K, Kim B Y, Ahn S C, Kang D O, Shin D I, Kim J, Mheen T I, Ahn J S. Inhibition of phospholipase C gamma 1 activity by amentoflavone isolated from Selaginella tamariscina .  Planta Medica. 1996;  62 293-6
• 9 Lin Y M, Chen F C, Lee K H. Hinokiflavone, a cytotoxic principle from Rhus succedanea and the cytotoxicity of the related biflavonoids.  Planta Medica. 1989;  55 166-8
• 10 Lobstein-Guth A, Briancon-Scheid F, Victoire C, Haag-Berrurier M, Anton R. Isolation of amentoflavone from Ginkgo biloba .  Planta Medica. 1988;  54 555-6
• 11 Mora A, Paya M, Rios J L, Alcaraz M J. Structure-activity relationships of polymethoxyflavones and other flavonoids as inhibitors of non-enzymic lipid peroxidation.  Biochem Pharmacol. 1990;  40 793-7
• 12 Lin Y M, Anderson H, Flavin M T, Pai Y H, Mata-Greenwood E, Pengsuparp T, Pezzuto J M, Schinazi R F, Hughes S H, Chen F C. In vitro anti-HIV activity of biflavonoids isolated from Rhus succedanea and Garcinia multiflora .  J Nat Prod. 1997 ;  60 884-8
• 13 Suzuki A, Matsunaga K, Mimaki Y, Sashida Y, Ohizumi Y. Properties of amentoflavone, a potent caffeine-like Ca²⁺ releaser in skeletal muscle sarcoplasmic reticulum.  Eur J Pharmacol. 1999;  372 97-102
• 14 Butterweck V, Nahrstedt A, Evans J, Hufeisen S, Rauser L, Savage J, Popadak B, Ernsberger P, Roth B L. In vitro receptor screening of pure constituents of St. John’s wort reveals novel interactions with a number of GPCRs.  Psychopharmacology. 2002;  162 193-202
• 15 Chakravarthy B K, Rao Y V, Gambhir S S, Gode K D. Isolation of amentoflavone from Selaginella rupestris and its pharmacological activity on central nervous system, smooth muscles and isolated frog heart preparations.  Planta Medica. 1981;  43 64-70
• 16 Kim S Z, Kim S H, Park J K, Koh G Y, Cho K W. Presence and biological activity of C-type natriuretic peptide-dependent guanylate cyclase-coupled receptor in the penile corpus cavernosum.  J Urol. 1998;  159 1741-6
• 17 Murad F. Cyclic guanosine monophosphate as a mediator of vasodilation.  J Clin Invest. 1986;  78 1-5
• 18 Gordon J L, Martin W. Stimulation of endothelial prostacyclin production plays no role in endothelium-dependent relaxation of the pig aorta.  Br J Pharmacol. 1983;  80 179-86
• 19 Satake N, Kiyoto S, Shibata S, Gandhi V, Jones D J, Morikawa M. Possible mechanisms of inhibition with atropine against noradrenaline-induced contraction in the rabbit aorta.  Br J Pharmacol. 1992;  107 553-8
• 20 Loeb A L, Izzo NJ J r, Johnson R M, Garrison J C, Peach M J. Endothelium-derived relaxing factor release associated with increased endothelial cell inositol trisphosphate and intracellular calcium.  Am J Cardiol. 1988;  62 36G-40G
• 21 Standen N B, Quayle J M, Davies N W, Brayden J E, Huang Y, Nelson M T. Hyperpolarizing vasodilators activate ATP-sensitive K⁺ channels in arterial smooth muscle.  Science. 1989;  245 177-80
• 22 Mehrke G, Daut J. The electrical response of cultured guinea-pig coronary endothelial cells to endothelium-dependent vasodilators.  J Physiol. 1990;  430 251-72