Nitroxyl, the novel redox sibling of NO, suppresses cerebrovascular NADPH oxidase

Background: Nitroxyl (HNO), the reduced and protonated congener of nitric oxide (NO), is emerging as a novel entity with distinct pharmacology and therapeutic advantages over NO• [1]. Importantly, HNO has vasoprotective actions with the potential to serve as an antioxidant. Here we explored the ability of HNO to modulate cerebrovascular NADPH oxidase activity, a major source of superoxide (.O2-) in the vasculature. Materials and methods: Intracranial (pooled middle cerebral and basilar) and extracranial (carotid) cerebral arteries from male C57BL/6J mice were treated with angiotensin II (10 nM) acutely (30 min) and chronically (24 h), respectively, in the absence and presence of the HNO donor, Angeli's salt (AS). NADPH (100 μM)-stimulated .O2- production was then measured using lucigenin (5 μM)-enhanced chemiluminescence. Results: AS (1 μM) did not scavenge .O2- generated in a cell free xanthine (100 μM)/xanthine oxidase (0.05 U/ml) activity assay (control: 447.9 ± 90.8; AS 507.1 ± 113.3 counts, n = 4). In contrast, acute and chronic treatment with AS (0.01–1 μM) caused a concentration-dependent decrease in NADPH oxidase-derived .O2- production by intracranial and extracranial cerebral arteries, respectively (carotid 0.59 ± 0.05; AS 0.1 μM 0.33 ± 0.08; AS 1 μM 0.16 ± 0.03 103 counts/s/mg, P < 0.05, n = 8). The effects of AS were reversed by the HNO scavenger, L-cysteine (3 mM) but unchanged in the presence of the NO• scavenger carboxy-PTIO (200 μM) and sGC inhibitor, ODQ (10 μM). Conclusion: HNO suppresses vascular NADPH-oxidase activity both acutely and chronically, possibly via a cGMP-independent mechanism. Such antioxidant actions of HNO may confer therapeutic advantages in the treatment of cerebrovascular disorders

Targeting the NO/sGC/cGMP signaling pathway in health and disease

The therapeutic utility of the nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine 3’5’-monophosphate (cGMP) pathway is well-recognised, with NO donors used in the treatment of diseases such as heart failure, acute hypertensive crisis and pulmonary hypertension. However, traditional nitrovasodilator therapy is somewhat limited due to the development of tolerance following prolonged administration. Furthermore, in vascular disease states such as hypertension, the NO/sGC/cGMP pathway appears to be dysfunctional. Such dysfunction is thought to arise as a consequence of oxidative stress and the associated increase in vascular superoxide anion radical (•O2-) levels, leading to enhanced scavenging of NO to form the oxidant, peroxynitrite (ONOO-). Indeed, ONOO- is able to oxidise the reduced (Fe2+) heme group of sGC, converting it into its NO-insensitive ferric (Fe3+) or heme-free forms. Under such conditions, the efficacy of NO donors/nitrovasodilators are compromised, due to their inability to target these altered states of sGC, underlining the need for novel non-NO based compounds. As such, this thesis examines the vaso-protective actions and therapeutic potential of the redox sibling of NO, nitroxyl (HNO), as well as NO-independent sGC stimulators (BAY 41-2272) and activators (BAY 58-2667) in a number of cardiovascular disease states. From a clinical perspective, NO-independent compounds such as HNO donors and/or stimulators and activators of sGC may offer considerable therapeutic advantages over traditional nitrovasodilator therapies due to their well-documented resistance to •O2- scavenging and vascular tolerance development, as well as the ability to target oxidised/heme-free sGC (i.e. BAY 58-2667) and the potential to mediate their effects via cGMP-independent mechanisms of action. Such compounds may also offer the basis from which future therapies, for the treatment of cardiovascular diseases, are designed

Targeting the NO/sGC/cGMP signaling pathway in health and disease

The therapeutic utility of the nitric oxide (NO)/soluble guanylyl cyclase (sGC)/cyclic guanosine 3’5’-monophosphate (cGMP) pathway is well-recognised, with NO donors used in the treatment of diseases such as heart failure, acute hypertensive crisis and pulmonary hypertension. However, traditional nitrovasodilator therapy is somewhat limited due to the development of tolerance following prolonged administration. Furthermore, in vascular disease states such as hypertension, the NO/sGC/cGMP pathway appears to be dysfunctional. Such dysfunction is thought to arise as a consequence of oxidative stress and the associated increase in vascular superoxide anion radical (•O2-) levels, leading to enhanced scavenging of NO to form the oxidant, peroxynitrite (ONOO-). Indeed, ONOO- is able to oxidise the reduced (Fe2+) heme group of sGC, converting it into its NO-insensitive ferric (Fe3+) or heme-free forms. Under such conditions, the efficacy of NO donors/nitrovasodilators are compromised, due to their inability to target these altered states of sGC, underlining the need for novel non-NO based compounds. As such, this thesis examines the vaso-protective actions and therapeutic potential of the redox sibling of NO, nitroxyl (HNO), as well as NO-independent sGC stimulators (BAY 41-2272) and activators (BAY 58-2667) in a number of cardiovascular disease states. From a clinical perspective, NO-independent compounds such as HNO donors and/or stimulators and activators of sGC may offer considerable therapeutic advantages over traditional nitrovasodilator therapies due to their well-documented resistance to •O2- scavenging and vascular tolerance development, as well as the ability to target oxidised/heme-free sGC (i.e. BAY 58-2667) and the potential to mediate their effects via cGMP-independent mechanisms of action. Such compounds may also offer the basis from which future therapies, for the treatment of cardiovascular diseases, are designed

Nitroxyl donors retain their depressor effects in hypertension

2013; doi:10.1152/ajpheart.00630.2012.—Nitroxyl (HNO), the redox congener of nitric oxide, has numerous vasoprotective actions includ-ing an ability to induce vasodilation and inhibit platelet aggregation. Given HNO is resistant to scavenging by superoxide and does not develop tolerance, we hypothesised that HNO would retain its in vivo vasodilatory action in the setting of hypertension. The in vitro and in vivo vasodilator properties of the HNO donors Angeli’s salt (AS) and isopropylamine/NONOate (IPA/NO) were compared with the NO˙ donor diethylamine/NONOate (DEA/NO) in spontaneously hyperten-sive rats (SHR) and normotensive [Wistar-Kyoto (WKY) rats]. AS (10, 50, and 200 g/kg), IPA/NO (10, 50, and 200 g/kg), and DEA/NO (1, 5, and 20 g/kg) caused dose-dependent depressor responses in conscious WKY rats of similar magnitude. Depressor responses to AS and IPA/NO were significantly attenuated (P 0.01) after infusion of the HNO scavenger N-acetyl-L-cysteine (NAC)

Vampire Venom: Vasodilatory Mechanisms of Vampire Bat (Desmodus rotundus) Blood Feeding

Animals that specialise in blood feeding have particular challenges in obtaining their meal, whereby they impair blood hemostasis by promoting anticoagulation and vasodilation in order to facilitate feeding. These convergent selection pressures have been studied in a number of lineages, ranging from fleas to leeches. However, the vampire bat (Desmondus rotundus) is unstudied in regards to potential vasodilatory mechanisms of their feeding secretions (which are a type of venom). This is despite the intense investigations of their anticoagulant properties which have demonstrated that D. rotundus venom contains strong anticoagulant and proteolytic activities which delay the formation of blood clots and interfere with the blood coagulation cascade. In this study, we identified and tested a compound from D. rotundus venom that is similar in size and amino acid sequence to human calcitonin gene-related peptide (CGRP) which has potent vasodilatory properties. We found that the vampire bat-derived form of CGRP (i.e., vCGRP) selectively caused endothelium-independent relaxation of pre-contracted rat small mesenteric arteries. The vasorelaxant efficacy and potency of vCGRP were similar to that of CGRP, in activating CGRP receptors and Kv channels to relax arteriole smooth muscle, which would facilitate blood meal feeding by promoting continual blood flow. Our results provide, for the first time, a detailed investigation into the identification and function of a vasodilatory peptide found in D. rotundus venom, which provides a basis in understanding the convergent pathways and selectivity of hematophagous venoms. These unique peptides also show excellent drug design and development potential, thus highlighting the social and economic value of venomous animals