Corine Girard-Thernier

Vasorelaxant effects and mechanisms of action of Heracleum sphondylium L. (Apiaceae) in rat thoracic aorta

ETHNOPHARMACOLOGICAL RELEVANCE Aerial parts of Heracleum sphondylium L. (HS) are used in traditional medicine to treat hypertension. To provide pharmacological basis for this use, we investigated the vasorelaxant effects of a dichloromethane extract of HS (HSDE) and the mechanisms involved. MATERIALS AND METHODS Activity of HSDE was evaluated on rat isolated thoracic aortic rings. RESULTS HSDE induced vasorelaxation in phenylephrine (PE, 10(-6)mol/L) and high KCl-(6×10(-2)mol/L) pre-contracted aortic rings that was independent on the presence of endothelium. HSDE markedly decreased extracellular Ca(2+)-induced contraction in high-KCl and PE pre-challenged rings. It also inhibited the intracellular Ca(2+) release sensitive to PE (10(-6)M). The relaxant effect of HSDE were blunted by 4-amino-pyridine (4-AP, 10(-3)mol/L), an inhibitor of voltage-dependent K(+) channels. CONCLUSION Our results provide the first evidence that a dichloromethane extract of Heracleum sphondylium L. exhibits vasorelaxant properties through endothelium-independent mechanisms involving the inhibition of Ca(2+) mobilization and changes in Kv channel conductances. These data argue for its use as antihypertensive therapy in traditional medicine.

Arginase Inhibitors: A Rational Approach Over One Century.

Arginase (EC 3.5.3.1) is the bimanganese enzyme that converts L‐arginine into ornithine and urea. This enzyme was discovered more than a century ago and early α‐amino acids were identified as weak inhibitors. It was only during the 90s, after nitric oxide (NO) was reported as one of the most important biological mediators and when tight interrelation of arginase and NO synthase was found, that the development of arginase inhibitors was accelerated. The regulation of arginase activity by the N‐hydroxy‐L‐arginine (3, NOHA) intermediate of the NO synthesis was the starting point of the N‐hydroxy‐nor‐arginine (21, nor‐NOHA) that proved to be the first micromolar inhibitor. The previously known manganese and arginase binding by borate inspired the 2(S)‐amino‐6‐boronohexanoic acid (39, ABH) and S‐(2‐boronoethyl)‐L‐cysteine (40, BEC) now both considered as reference compounds in arginase inhibition. The high‐resolution crystal structure of arginase and molecular modeling has rendered possible the recent design of (53) the strongest α,α‐disubstituted derivatives of ABH. Simultaneously, traditional medicinal plants have contributed as a source of molecular diversity to the discovery of arginase inhibitors. This rational, step‐by‐step approach serves as guide in the present review where emphasis is placed on structure activity relationships.

New insights into the mechanisms of the vasorelaxant effects of apocynin in rat thoracic aorta.

Apocynin is a naturally occurring acetophenone widely used as an inhibitor of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Recent data suggested that apocynin might exert NADPH oxidase‐independent pharmacological properties. Among them, vasorelaxant properties have been described, but the mechanisms still give rise to debates. The present study investigated the mechanisms involved in the vasorelaxant effect of apocynin on the in vitro model of rat isolated thoracic aortic rings. Apocynin (30 μm to 10 mm) induced a dose‐dependent relaxation in both endothelium‐intact and endothelium‐denuded aortic rings with respective EC50 values of 0.78 ± 0.08 and 1.91 ± 0.21 mm. Endothelium removal or inhibition of nitric oxide (NO) synthase with Nω‐nitro‐l‐arginine‐methyl ester (l‐NAME) significantly decreased but did not abolish the effect of apocynin. By contrast, apocynin‐induced relaxation was unchanged after incubation with indomethacin or charybdotoxin plus apamin. In endothelium‐denuded aortas, the vasorelaxant effect of apocynin was significantly reduced by glibenclamide but not by 4‐aminopyridine nor by iberiotoxin. Apocynin significantly decreased Ca2+‐induced contraction and inhibited intracellular Ca2+mobilization after contraction with phenylephrine. Finally, the acute intravenous injection of apocynin led to an immediate and transient hypotensive effect in spontaneously hypertensive rats (SHR). In conclusion, our data demonstrated that apocynin induces both endothelium‐independent relaxant effects involving inhibition of Ca2+mobilization and activation of KATP channels in vascular smooth muscle cells and endothelium‐dependent effects mediated by NO. These results should provide a basis for caution when interpreting results on the vascular effects of apocynin.

Bioassay-guided isolation of vasorelaxant compounds from Ziziphora clinopodioides Lam. (Lamiaceae)

Ziziphora clinopodioides Lam. (Lamiaceae) is traditionally used in Uighurs medicine for the treatment of hypertension. Our study determined and evaluated the bioactive compounds by performing an activity-guided fractionation of a hydroalcoholic extract of the whole plant, using an in vitro model of rat isolated thoracic aortic rings. Seven compounds were identified as active principles: acacetin, apigenin, chrysin, thymonin, acetovanillone, 4-hydroxyacetophenone and ethyl 4-coumarate. Apigenin, chrysin and ethyl 4-coumarate were found to be the most effective. Our results provide the first evidence that the vasodilation induced by Z. clinopodioides Lam. is mediated, at least in part, by phenolic components.

Investigation of Mammal Arginase Inhibitory Properties of Natural Ubiquitous Polyphenols by Using an Optimized Colorimetric Microplate Assay

Polyphenols are plant secondary metabolites which possess many positive effects on human health. Although these beneficial effects could be mediated through an increase in nitric oxide synthase activity, little is known regarding the inhibitory effect of polyphenols on mammal arginase, an enzyme which competes with nitric oxide synthase for their common substrate, L-arginine. The aim of the present study was to determine the potential of a series of polyphenols as mammalian arginase inhibitors and to identify some structure-activity relationships. For this purpose, we first developed a simple and cost-effective in vitro colorimetric microplate method using commercially-available mammal bovine liver arginase (b-ARG 1). Among the ten tested polyphenolic compounds [chlorogenic acid, piceatannol, resveratrol, (-)-epicatechin, taxifolin, quercetin, fisetin, caffeic acid, quinic acid, and kaempferol], cholorogenic acid and piceatannol exhibited the highest inhibitory activities (IC50 = 10.6 and 12.1 µM, respectively) but were however less active as (S)-(2-Boronoethyl)-L-cysteine (IC50 = 3.3 µM), used as reference compound. Enzyme kinetic studies showed that both chlorogenic acid and piceatannol are competitive arginase inhibitors. Structural data identified the importance of the caffeoyl (3,4-dihydroxycinnamoyl)-part and of the catechol function in the inhibitory activity of the tested compounds. These results identified chlorogenic acid and piceatannol as two potential core structures for the design of new arginase inhibitors.

Cinnamide Derivatives as Mammalian Arginase Inhibitors: Synthesis, Biological Evaluation and Molecular Docking

Arginases are enzymes that are involved in many human diseases and have been targeted for new treatments. Here a series of cinnamides was designed, synthesized and evaluated in vitro and in silico for their inhibitory activity against mammalian arginase. Using a microassay on purified liver bovine arginase (b-ARG I), (E)-N-(2-phenylethyl)-3,4-dihydroxycinnamide, also named caffeic acid phenylamide (CAPA), was shown to be slightly more active than our natural reference inhibitor, chlorogenic acid (IC50 = 6.9 ± 1.3 and 10.6 ± 1.6 µM, respectively) but it remained less active that the synthetic reference inhibitor Nω-hydroxy-nor-l-arginine nor-NOHA (IC50 = 1.7 ± 0.2 µM). Enzyme kinetic studies showed that CAPA was a competitive inhibitor of arginase with Ki = 5.5 ± 1 µM. Whereas the activity of nor-NOHA was retained (IC50 = 5.7 ± 0.6 µM) using a human recombinant arginase I (h-ARG I), CAPA showed poorer activity (IC50 = 60.3 ± 7.8 µM). However, our study revealed that the cinnamoyl moiety and catechol function were important for inhibitory activity. Docking results on h-ARG I demonstrated that the caffeoyl moiety could penetrate into the active-site pocket of the enzyme, and the catechol function might interact with the cofactor Mn2+ and several crucial amino acid residues involved in the hydrolysis mechanism of arginase. The results of this study suggest that 3,4-dihydroxycinnamides are worth being considered as potential mammalian arginase inhibitors, and could be useful for further research on the development of new arginase inhibitors.

Research of novel anticancer agents targeting arginase inhibition.

Arginase plays an important part in l-arginine metabolism. This metalloenzyme also regulates polyamine biosynthesis, nitric oxide production and the T-cell-mediated immune response, which are all involved in the growth and control of cancer. Research over the past decades has reported arginase as an attractive target for cancer treatment, and inhibition of arginase could be a promising strategy for cancer therapy. Herein, we present the available data on the role of arginase in cancer development. The principal synthetic and natural arginase inhibitors are outlined and followed by their mechanism of action. Among them, some molecules have shown their anticancer effects. The perspectives of arginase inhibitors as new anticancer agents will be discussed.

Treatment with an extract of Terminalia superba Engler & Diels decreases blood pressure and improves endothelial function in spontaneously hypertensive rats.

ETHNOPHARMACOLOGICAL RELEVANCE The stem bark of Terminalia superba (TS) is widely used as a decoction by Cameroonian folk medicine for the treatment of hypertension. The aim of the present study was to evaluate the effect of a chronic treatment with a TS extract on spontaneously hypertensive rats (SHR) with respect to efficacy, biochemical mechanisms and safety. MATERIALS AND METHODS Eleven-week-old SHR and normotensive Wistar Kyoto rats (WKY) were daily treated by gavage with a methylene chloride extract of stem bark of Terminalia superba (TMSE, 150mg/kg) or with the vehicle for 5 weeks. Systolic blood pressure (SBP) was measured weekly using the tail-cuff method. At the end of the treatment period, vascular function was assessed on isolated thoracic rings, urinary 8-iso-PGF2α levels were measured and cytochrome P-450 3A (CYP 3A) activity was evaluated in liver microsomes. RESULTS TMSE reduced SBP (P<0.001) in SHR but not in WKY rats. In SHR, the vasorelaxant response to acetylcholine was significantly improved by TMSE as a result of increased nitric oxide synthase (NO) activity and decreased superoxide anion production. In addition, TMSE reduced the vasoconstrictive effect of phenylephrine and improved the sensitivity of smooth muscle cells to NO. TMSE dramatically decreased 8-iso-PGF2α levels in SHR. By contrast, TMSE did not affect all these parameters in WKY rats. Neither diuresis nor the hepatic CYP 3A activity was modified in both animal groups. CONCLUSIONS This study demonstrated that Terminalia superba has a potent antihypertensive activity in SHR which is partly due to endothelium-dependent and endothelium-independent effects as well as decreased oxidative stress. The data also provide evidence for the lack of herb-drug interaction through hepatic CYP 3A.

Cyperaceae Species Are Potential Sources of Natural Mammalian Arginase Inhibitors with Positive Effects on Vascular Function

The inhibition of arginase is of substantial interest for the treatment of various diseases of public health interest including cardiovascular diseases. Using an ex vivo experiment on rat aortic rings and an in vitro assay with liver bovine purified arginase, it was demonstrated that several polyphenolic extracts from Cyperus and Carex species possess vasorelaxant properties and mammalian arginase inhibitory capacities. Phytochemical studies performed on these species led to the identification of eight compounds, including monomers, dimers, trimers, and tetramers of resveratrol. The potential of these stilbenes as inhibitors of mammalian arginase was assessed. Five compounds, scirpusin B (5), ε-viniferin (4), cyperusphenol B (6), carexinol A (7), and the new compound virgatanol (1), showed significant inhibition of arginase, with percentage inhibition ranging from 70% to 95% at 100 μg/mL and IC50 values between 12.2 and 182.1 μM, confirming that these stilbenes may be useful for the development of new pharmaceutical products.