Effect of two phenanthrene alkaloids on angiotensin II‐induced leukocyte–endothelial cell interactions in vivo

Abstract

1 The present study has evaluated the effect of two phenanthrene alkaloids, uvariopsine and stephenanthrine, on angiotensin II (Ang-II)-induced leukocyte -endothelial cell interactions in vivo and the mechanisms involved in their activity. Intravital microscopy within the rat mesenteric microcirculation was used. 2 A 60 min superfusion with 1 nM Ang-II induced a significant increase in the leukocyte -endothelial cell interactions that were completely inhibited by 1 mM uvariopsine cosuperfusion. A lower dose of 0.1 mM significantly reduced Ang-II-induced leukocyte adhesion by 75%. 3 When Ang-II was cosuperfused with 1 and 0.1 mM stephenanthrine, Ang-II-induced leukocyte responses were significantly diminished. A lower dose of 0.01 mM only affected Ang-II-induced leukocyte adhesion. 4 Both alkaloids inhibited Ang-II-induced endothelial P-selectin upregulation and the generation of reactive oxygen species (ROS) in endothelial cells stimulated with Ang-II, in fMLP-stimulated human neutrophils (PMNs) and in the hypoxanthine -xanthine oxidase system. However, cyclic AMP levels in PMNs stimulated with fMLP were not affected. 5 Uvariopsine and stephenanthrine inhibited PAF-induced elevations in intracellular calcium levels in PMNs (IC 50 values: 15.1 and 6.1 mM respectively) and blocked the binding of [ 3 H]PAF to these leukocytes. They also reduced PAF-induced increases in intracellular levels of superoxide anion and hydrogen peroxide. 6 In conclusion, stephenanthrine and uvariopsine are potent inhibitors of Ang-II-induced leukocyte accumulation in vivo. This effect appears to be mediated through ROS scavenging activity and blockade of PAF receptor. Thus, they have potential therapeutic interest for the control of leukocyte recruitment that occurs in cardiovascular disease states in which Ang-II is involved.

Effect of boldine, secoboldine, and boldine methine on angiotensin II-induced neurtrophil recruitment in vivo

“…In this context, BM was capable of inhibiting the release of this chemoattractant in vivo when animals were i.p.-injected with Ang-II. Additionally, some phenanthrene alkaloids bind to the PAF receptor and behave as antagonists of this inflammatory mediator, as it has been demonstrated by using binding assays in rabbit platelets and human neutrophils [10,26]. In the present study, we have encountered that only BM was capable of blocking the PAF receptor in a concentration-dependent manner (IC 50 28.2 M), and B and SB only inhibited [ 3 H]PAF binding to PMNs when they were assayed at 100 M. The effect displayed by BM is of a lesser magnitude than that observed with uvariposine and stephenanthrine, which were twice and six times more potent than the alkaloid tested [10].…”

“…Nevertheless, all of these alkaloids are less potent than the well-known PAF receptor antagonist WEB2086 (IC 50 0.15 M). As opposed to uvariopsine and stephenanthrine, which present a methylenedioxy group in their structure [10], BM lacks this functional group, which would result in a lower potency as a PAF receptor antagonist.…”

“…with 20 nmol/kg B, SB, or BM, 15 min prior to Ang-II injection. The dose selected for testing these alkaloids was that which caused the total inhibition of Ang-II-induced, leukocyte-endothelial cell interactions in vivo by two related alkaloids, uvariopsine and stephenanthrine, previously studied by us [10]. A concentration response on this assay was not carried out as a result of the amount of animals to be used.…”

“…The experimental preparation used in this study was similar to that described previously [10]. Male Sprague-Dawley rats (200 -250 g) were fasted for 24 h and anesthetized with sodium pentobarbital (50 mg/kg, i.p.).…”

“…Conversely, our group isolated two phenanthrene alkaloids, uvariopsine and stephenanthrine, from the fresh root of Dennettia tripetala [9] and proved that they could inhibit Ang-IIinduced leukocyte-endothelial cell interactions in vivo [10]. This effect was partly mediated through inhibition of the generation of reactive oxygen species (ROS), down-regulation of P-selectin expression on the endothelial cell, and blockade of platelet-activating factor (PAF)-induced responses by interacting with its receptor [10]. Based on their structural features, we semisynthesized four phenanthrene and one aporphine alkaloid from natural boldine (B) and evaluated their inhibitory effect on ROS generation [11].…”