Thymoquinone as a pharmacological tool to study the endothelium-dependent, soluble guanylyl cyclase-dependent augmentation caused by hypoxia in isolated arteries

Abstract

BACKGROUND: Experiments were designed to study the effects of thymoquinone on isolated arteries and to determine the mechanism underlying the endothelium-dependent, soluble guanylyl cyclase-dependent augmentation the compound is causing. METHODS: Rings, with or without endothelium, of porcine coronary arteries and rat arteries were suspended in conventional organ chambers for isometric tension recording. Certain rings were incubated with inhibitors of nitric oxide (NO) synthase (L-NG-nitroarginine methyl ester, L-NAME) or soluble guanylyl cyclase (1H-[1,2,4]oxadiazolo [4,3-a]quinoxalin-1-one, ODQ). They were contracted with phenylephrine (rat arteries) or prostaglandin F2a (porcine coronaries) and exposed to increasing concentrations of thymoquinone. Cyclic nucleotides were measured by HPLC-MS/MS. RESULTS: Thymoquinone caused a sustained further increase of tension in rings with endothelium. This augmentation was prevented by endothelium-removal, L-NAME and ODQ. Thymoquinone did not change the tissue levels of cyclic GMP, measured by immunoassay. Incubation with the NO-donor DETA NONOate in L-NAME-treated rings restored and even increased the contractile response to thymoquinone. Treatment with 8-bromo cyclic GMP or pyrophospate did not restore the augmentation by thymoquinone. By contrast, administration of cyclic IMP restored contractions to thymoquinone in rings without endothelium. HPLC-MS/MS measurements revealed that the compound increased the production of cyclic IMP. CONCLUSIONS: The endothelium-dependent augmentation caused by thymoquinone requires endothelium-derived NO and activation of soluble guanylyl cyclase, as described under hypoxic conditions. In addition, both thymoquinone- and hypoxia-induced augmentation require production of cyclic IMP but not the presence of either pyrophosphate or cyclic GMP. Thus, thymoquinone can serve as a pharmacological tool to mimic the endothelium-dependent vasoconstrictor effects of intermittent hypoxia, as seen in the clinical setting of sleep apnea patients.