Endothelium-Dependent Contractions of Isolated Arteries to Thymoquinone Require Biased Activity of Soluble Guanylyl Cyclase with Subsequent Cyclic IMP Production

Abstract

Preliminary experiments on isolated rat arteries demonstrated that thymoquinone, a compound widely used for its antioxidant properties and believed to facilitate endothelium-dependent relaxations, as a matter of fact caused endothelium-dependent contractions. The present experiments were designed to determine the mechanisms underlying this unexpected response. Isometric tension was measured in rings (with and without endothelium) of rat mesenteric arteries and aortae and of porcine coronary arteries. Precontracted preparations were exposed to increasing concentrations of thymoquinone, which caused concentration-dependent, sustained further increases in tension (augmentations) that were prevented by endothelium removal, Nv-nitro-L-arginine methyl ester [L-NAME; nitric oxide (NO) synthase inhibitor], and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; soluble guanylyl cyclase [sGC] inhibitor). In L-NAME-treated rings, the NO-donor diethylenetriamine NONOate restored the thymoquinone-induced augmentations; 5-[1-(phenylmethyl)-1H-indazol-3-yl]-2-furanmethanol (sGC activator) and cyclic IMP (cIMP) caused similar restorations. By contrast, in ODQ-treated preparations, the cell-permeable cGMP analog did not restore the augmentation by thymoquinone. The compound augmented the content (measured with ultra-high performance liquid chromatography-tandem mass spectrometry) of cIMP, but not that of cGMP; these increases in cIMP content were prevented by endothelium removal, L-NAME, and ODQ. The augmentation of contractions caused by thymoquinone was prevented in porcine arteries, but not in rat arteries, by 1-(5-isoquinolinylsulfonyl)homopiperazine dihydrochloride and trans-4-[(1R)-1-aminoethyl]-N-4-pyridinylcyclohexanecarboxamide dihydrochloride (Rho-kinase inhibitors); in the latter, but not in the former, it was reduced by 3,5-dichloro-N-[[(1a,5a,6-exo,6a)-3-(3,3-dimethylbutyl)-3-azabicyclo[3.1.0]hex-6-yl]methyl]-benzamide hydrochloride (T-type calcium channel inhibitor), demonstrating species/vascular bed differences in the impact of cIMP on calcium handling. Thymoquinone is the first pharmacological agent that causes endothelium-dependent augmentation of contractions of isolated arteries, which requires endothelium-derived NO and biased sGC activation, resulting in the augmented production of cIMP favoring the contractile process.

Recent progress in the field of cIMP research

“…The occurrence of cIMP in only certain pathophysiological settings could be an explanation for the failure to detect cIMP in several biological systems under normal conditions, i.e., without hypoxia (Beste et al, 2013;Hartwig et al, 2014;Bähre et al, 2015). Detremmerie et al (2016) provide evidence in favor of the notion that the pharmacological actions of thymoquinone depend on sGC-catalyzed cIMP formation. Hence, thymoquinone constitutes a novel pharmacological tool that can be used to increase intracellular cIMP concentrations.…”

“…However, convincing and wellcontrolled clinical studies are still missing. In their study, Detremmerie et al (2016) show that thymoquinone at moderately high concentrations (10-100 μM) can exhibit paradoxical vasoconstrictory effects via Ca 2+ influx by T-type-or L-type Ca 2+ channels. cIMP-induced vasoconstriction may play a role in coronary ischemia/reperfusion injury and/or cardiovascular complications in sleep apnea (Detremmerie et al, 2016).…”

“…In their study, Detremmerie et al (2016) show that thymoquinone at moderately high concentrations (10-100 μM) can exhibit paradoxical vasoconstrictory effects via Ca 2+ influx by T-type-or L-type Ca 2+ channels. cIMP-induced vasoconstriction may play a role in coronary ischemia/reperfusion injury and/or cardiovascular complications in sleep apnea (Detremmerie et al, 2016). The occurrence of cIMP in only certain pathophysiological settings could be an explanation for the failure to detect cIMP in several biological systems under normal conditions, i.e., without hypoxia (Beste et al, 2013;Hartwig et al, 2014;Bähre et al, 2015).…”

“…In an unrelated recent study, the group of Vanhoutte followed up their previous study on cIMP-induced vasoconstriction (Chen et al, 2014) and examined the effects of thymoquinone on blood vessels (Detremmerie et al, 2016). Thymoquinone is the pharmacologically active constituent of Nigella sativa and exhibits pleiotropic pharmacological effects including antioxidant properties (Amin and Hosseinzadeh, 2016).…”

“…The precise molecular mechanism of action of thymoquinone in the sGC-cGMP/cIMP system is not yet known, and it is unlikely that the pleiotropic pharmacological effects of the natural product (Amin and Hosseinzadeh, 2016) can all be attributed to a single molecular mechanism. To this end, it is clear that for the cIMP increase, the quinone moiety of thymoquinone is required (Detremmerie et al, 2016), a first step towards the elucidation of structure/activity relationships.…”