Objective: To investigate the effect of osthole on isolated thoracic aortic rings, and to determine the potential mechanism of action.
Methods: Thoracic aortas were isolated from Wistar rats, and were suspended in tissue organ chambers for vascular tension measurement. The effect of cumulative osthole (10－?, 10－?, 10－?, 10－?, and 10－? mol/L) on endothelium-intact and endothelium-denuded thoracic aortic rings pre-contracted with phenylephrine (PE, 10－? mol/L) or KCl (6 × 10－? mol/L) was recorded. Histomorphological changes of thoracic aorta were analyzed by hematoxylin-eosin. The effects of different osthole concentrations on endothelium-intact aortic rings, which were pre-inhibited with the non-selective nitric oxide synthase inhibitor L-Arg(NO2)-OMe·HCl (3 × 10－4 mol/L), endothelium-derived nitric oxide synthase inhibitor Nω-nitro-L-arginine (3 × 10－4 mol/L), guanylate cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3-α] quinoxaline-1-one (10－5 mol/L), cyclooxygenase inhibitor indometacin (10－5 mol/L), and the Ca2+-activated potassium channel inhibitor tetraethylammonium nitrate (10－5 mol/L), and then contracted with PE, were examined. Aortic rings incubated with osthole (10－5 mol/L), phentolamine (10－5 mol/L), or verapamil (10－5 mol/L) in Ca2+-free Krebs-Henseleit solution (KHS) were stimulated with PE or KCl.
Results: There was a dose-dependent increase in vasorelaxation of isolated thoracic aortic rings (both with and without endothelium) with increasing osthole concentration. Hematoxylin-eosin staining showed that osthole significantly improved thoracic aorta ring morphology. Compared with the control group, there were also significant differences after incubation with L-Arg(NO2)-OMe·HCl, Nω-nitro-L-arginine, and 1H-[1,2,4] oxadiazolo [4,3-α] quinoxaline-1-one (P < 0.05 for all). The relaxation rate of the rings in the osthole group incubated with indometacin and tetraethylammonium nitrate were similar to controls. In Ca2+-free KHS, the PE-induced contraction was similar between the osthole (4.37% ± 0.41%) and control (4.21% ± 1.33%) groups. However, after cumulative CaCl2 (0.5, 1, 1.5, 2, 2.5, and 3 mmol/L), the Ca2+-induced contraction was significantly inhibited in the osthole and phentolamine groups compared with controls (P < 0.05). After cumulative CaCl2 was added to Ca2+-free KHS (high K+ concentration), the contraction rate was significantly higher than both of the control and the osthole groups (P < 0.05). The contraction rate in the osthole group was higher than the verapamil group (P < 0.05).
Conclusion: Osthole has a vasorelaxant effect on isolated rat thoracic aortic rings, via inhibition of both receptor-operated and voltage-dependent Ca2+ channels in arterial smooth muscle, leading to decreased Ca2+ influx, and via inhibition of nitric oxide release on arterial endothelial cells.
Keywords: Aorta, thoracic; Calcium; Nitric oxide; Osthole; Phentolamine; Vasorelaxation; Verapamil.