Efficacy of BAY 60-2770, a Soluble Guanylate Cyclase Activator, for Coronary Spasm in Animal Models.

Ischemia with non-obstructive coronary arteries (INOCA), caused by coronary artery spasm, has gained increasing attention owing to the poor quality of life of impacted patients. Therapeutic options to address INOCA remain limited, and developing new therapeutic agents is desirable. Here, we examined whether soluble guanylate cyclase (sGC) activators could be beneficial in preventing coronary spasms. In organ chamber experiments with isolated canine coronary arteries, prostaglandin F2 α -induced, endothelin-1-induced, 5-hydroxytryptamine-induced, and potassium chloride-induced contractions were suppressed by the sGC activator BAY 60-2770 (0.1, 1, and 10 nM). In isolated pig coronary arteries, BAY 60-2770 (0.1, 1, and 10 nM) could prolong the cycle length of phasic contractions induced by 3,4-diaminopyridine, as well as lower the peak and bottom tension of the contraction in a concentration-dependent manner. Additionally, BAY 60-2770 (1 pM-0.1 µM) evoked a concentration-related relaxation to a greater extent in small (first diagonal branch) coronary arteries than in large (left anterior descending) coronary arteries. In vasopressin-induced angina model rats, pretreatment with BAY 60-2770 (3 µg/kg) suppressed electrocardiogram S-wave depression induced by arginine vasopressin without affecting changes in mean blood pressure and heart rate. These findings suggest that BAY 60-2770 could be valuable in preventing both large and small coronary spasms. Therefore, sGC activators could represent a novel and efficacious therapeutic option for INOCA. SIGNIFICANCE STATEMENT: The soluble guanylate cyclase (sGC) activator BAY 60-2770 exerted antispastic effects on the coronary arteries in animal vasospasm models as proof-of-concept studies. These data can help to support potential clinical development with sGC activators, suitable for human use in patients with vasospastic angina.

Effects of beetroot juice supplementation on vascular functional and structural changes in aged mice.

This study investigated whether beetroot juice (BRJ) ingestion ameliorates aging-induced functional and structural changes in vasculature. Aged mice (98-100 weeks old) were supplemented with BRJ (nitrate: 3.5 mmol/L) or drinking water for 4 weeks and compared with young mice (12-15 weeks old). The vasorelaxant response of isolated aortas to acetylcholine was markedly weaker in aged mice than in young mice, but the attenuated relaxation was significantly improved in BRJ-supplemented aged mice. The acetylcholine-induced relaxation was completely abolished by Nω -nitro-l-arginine methyl ester in all groups. Additionally, the response to sodium nitroprusside was comparable among the three groups. The aortic medial thickness was significantly greater in aged mice than in young mice, and BRJ supplementation did not suppress this thickening. Plasma nitrate levels were significantly higher in BRJ-supplemented aged mice than in non-supplemented aged mice. Conversely, non-supplemented aged mice had high plasma levels of thiobarbituric acid-reactive substances, but the levels were suppressed in BRJ-supplemented aged mice. These findings suggest that BRJ ingestion improves vascular endothelial dysfunction associated with aging, at least in part, by enhancing nitric oxide bioavailability and reducing oxidative stress. Therefore, beetroot ingestion may be a highly useful self-medication option to prevent vascular aging.

Factors influencing the soluble guanylate cyclase heme redox state in blood vessels.

Soluble guanylate cyclase (sGC) plays an important role in maintaining vascular homeostasis, as an acceptor for the biological messenger nitric oxide (NO). However, only reduced sGC (with a ferrous heme) can be activated by NO; oxidized (ferric heme) and apo (absent heme) sGC cannot. In addition, the proportions of reduced, oxidized, and apo sGC change under pathological conditions. Although diseased blood vessels often show decreased NO bioavailability in the vascular wall, a shift of sGC heme redox balance in favor of the oxidized/apo forms can also occur. Therefore, sGC is of growing interest as a drug target for various cardiovascular diseases. Notably, the balance between NO-sensitive reduced sGC and NO-insensitive oxidized/apo sGC in the body is regulated in a reversible manner by various biological molecules and proteins. Many studies have attempted to identify endogenous factors and determinants that influence this redox state. For example, various reactive nitrogen and oxygen species are capable of inducing the oxidation of sGC heme. Conversely, a heme reductase and some antioxidants reduce the ferric heme in sGC to the ferrous state. This review summarizes the factors and mechanisms identified by these studies that operate to regulate the sGC heme redox state.

Acute Kynurenine Exposure of Rat Thoracic Aorta Induces Vascular Dysfunction via Superoxide Anion Production.

The accumulation of uremic toxins is known to be one of the causes of cardiovascular disorder related to renal disease. Among the many uremic toxins, we focused on kynurenine (kyn), whose levels have been shown to be positively correlated with vascular endothelial dysfunction markers, and directly evaluated the influence of kyn on the rat thoracic aorta. Exposure of the endothelium-intact aorta to kyn markedly attenuated the acetylcholine (ACh)-induced relaxation and significantly increased superoxide anion (O2·-) production. These effects were ameliorated by pretreatment with ascorbic acid, an antioxidant, and CH223191, an aryl hydrocarbon receptor (AhR) inhibitor, but not by apocynin, a reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor. In the endothelium-denuded aorta, kyn significantly attenuated the nitric oxide (NO) donor sodium nitroprusside (SNP)-induced vasorelaxation and increased the O2·- production. Ascorbic acid treatment significantly ameliorated these effects, whereas CH223191 and apocynin treatments did not. Kyn had no influence on the vasorelaxant response to BAY 41-2272, a soluble guanylate cyclase stimulator. This suggested that kyn attenuates the NO-mediated vasorelaxation response by promoting O2·- production in thoracic aorta to inactivate NO. O2·- production is likely stimulated in both vascular endothelium and smooth muscle, the former of which may be mediated by AhR activation.

Vascular Endothelial Dysfunction in the Thoracic Aorta of Rats with Ischemic Acute Kidney Injury: Contribution of Indoxyl Sulfate.

Chronic kidney disease (CKD) and cardiovascular disease are known to be linked, and the involvement of indoxyl sulfate (IS), a type of uremic toxin, has been suggested as one of the causes. It is known that IS induces vascular dysfunction through overproduction of reactive oxygen species (ROS). On the other hand, the involvement of IS in the vascular dysfunction associated with acute kidney injury (AKI) is not fully understood. Therefore, we investigated this issue using the thoracic aorta of rats with ischemic AKI. Ischemic AKI was induced by occlusion of the left renal artery and vein for 45 min, followed by reperfusion 2 weeks after contralateral nephrectomy. One day after reperfusion, there was marked deterioration in renal function evidenced by an increase in plasma creatinine. Furthermore, blood IS levels increased markedly due to worsening renal function. Seven days and 28 days after reperfusion, blood IS levels decreased with the improvement in renal function. Of note, acetylcholine-induced vasorelaxation deteriorated over time after reperfusion, contradicting the recovery of renal function. In addition, 28 days after reperfusion, we observed a significant increase in ROS production in the vascular tissue. Next, we administered AST-120, a spherical adsorbent charcoal, after reperfusion to assess whether the vascular endothelial dysfunction associated with the ischemic AKI was due to a temporary increase in blood IS levels. AST-120 reduced the temporary increase in blood IS levels after reperfusion without influencing renal function, but did not restore the impaired vascular reactivity. Thus, in ischemic AKI, we confirmed that the vascular endothelial function of the thoracic aorta is impaired even after the recovery of kidney injury, probably with excessive ROS production. IS, which increases from ischemia to early after reperfusion, may not be a major contributor to the vascular dysfunction associated with ischemic AKI.

Soluble Guanylate Cyclase-Mediated Relaxation in Aortas from Rats with Renovascular Hypertension.

Soluble guanylate cyclase (sGC) plays an important role in nitric oxide (NO)-mediated regulation of vascular tone; however, NO bioavailability is often reduced in diseased blood vessels. Accumulating evidence suggests that a shift of sGC from the NO-sensitive form to the NO-insensitive form could be an underlying cause contributing to this reduction. Herein, we investigated the impact of renovascular hypertension on NO-sensitive and NO-insensitive sGC-mediated relaxation in rat aortas. Renovascular hypertension was induced by partially clipping the left renal artery (2-kidneys, 1-clip; 2K1C) for 10 weeks. Systolic, diastolic, and mean arterial pressures were significantly increased in the 2K1C group when compared with the sham group. In addition, plasma thiobarbituric acid reactive substances and aortic superoxide generation were significantly enhanced in the 2K1C group when compared with those in the sham group. The vasorelaxant response of isolated aortas to the sGC stimulator BAY 41-2272 (NO-sensitive sGC agonist) was comparable between the sham and 2K1C groups. Likewise, the sGC activator BAY 60-2770 (NO-insensitive sGC agonist)-induced relaxation did not differ between the sham and 2K1C groups. In addition, the cGMP mimetic 8-Br-cGMP (protein kinase G agonist) induced similar relaxation in both groups. Furthermore, there were no differences in BAY 41-2272-stimulated and BAY 60-2770-stimulated cGMP generation between the groups. These findings suggest that the balance between NO-sensitive and NO-insensitive forms of sGC is maintained during renovascular hypertension. Therefore, sGC might not be responsible for the reduced NO bioavailability observed during renovascular hypertension.

Alteration of the soluble guanylate cyclase system in coronary arteries of high cholesterol diet-fed rabbits.

This study aimed to investigate how atherosclerosis affects the soluble guanylate cyclase (sGC) system in coronary arteries. Rabbits were fed a normal diet for 12 weeks (N group) or a diet containing high cholesterol (1%) for 4 weeks (S-HC group) and 12 weeks (L-HC group). Cholesterol deposition in the intima of coronary arteries was observed in the S-HC group, but the formation of an atherosclerotic plaque was not observed. In contrast, a major plaque developed in the L-HC group. The relaxant response of isolated coronary arteries to sodium nitroprusside (SNP, nitric oxide donor) was not different between the N and S-HC groups, whereas the response in the L-HC group was markedly attenuated. The relaxation induced by BAY 60-2770 (sGC activator) tended to be augmented in the S-HC group, but it was significantly impaired in the L-HC group compared to that in the N group. sGC ß1 immunostaining was equally detected in the medial layer of the arteries among the N, S-HC, and L-HC groups. In addition, a strong staining was observed in the plaque region of the L-HC group. cGMP levels in the arteries stimulated with SNP were identical in the N and S-HC groups and slightly lower in the L-HC group than the other groups. BAY 60-2770-stimulated cGMP formation tended to be increased in the S-HC and L-HC groups. These findings suggest that the sGC system was not normal in atherosclerotic coronary arteries. The redox state of sGC and the distribution pattern are likely to change with the progression of atherosclerosis.

Preventive effects of nitrate-rich beetroot juice supplementation on monocrotaline-induced pulmonary hypertension in rats.

Beetroot (Beta vulgaris L.) has a high level of nitrate; therefore, its dietary intake could increase nitric oxide (NO) level in the body, possibly preventing the development of pulmonary hypertension (PH). In this study, we examined the effects of beetroot juice (BJ) supplementation on PH and the contribution of nitrate to such effects using a rat model of monocrotaline (MCT, 60 mg/kg s.c.)-induced PH. Rats were injected subcutaneously with saline or 60 mg/kg MCT and were sacrificed 28 days after the injection. In some rats injected with MCT, BJ was supplemented from the day of MCT injection to the day of sacrifice. First, MCT-induced right ventricular systolic pressure elevation, pulmonary arterial medial thickening and muscularization, and right ventricular hypertrophy were suppressed by supplementation with low-dose BJ (nitrate: 1.3 mmol/L) but not high-dose BJ (nitrate: 4.3 mmol/L). Of the plasma nitrite, nitrate, and their sum (NOx) levels, only the nitrate levels were found to be increased by the high-dose BJ supplementation. Second, in order to clarify the possible involvement of nitrate in the preventive effects of BJ on PH symptoms, the effects of nitrate-rich BJ (nitrate: 0.9 mmol/L) supplementation were compared with those of the nitrate-depleted BJ. While the former exerted preventive effects on PH symptoms, such effects were not observed in rats supplemented with nitrate-depleted BJ. Neither supplementation with nitrate-rich nor nitrate-depleted BJ affected plasma nitrite, nitrate, and NOx levels. These findings suggest that a suitable amount of BJ ingestion, which does not affect systemic NO levels, can prevent the development of PH in a nitrate-dependent manner. Therefore, BJ could be highly useful as a therapy in patients with PH.

Indoxyl sulfate induces ROS production via the aryl hydrocarbon receptor-NADPH oxidase pathway and inactivates NO in vascular tissues.

AIMS: The uremic toxin indoxyl sulfate (IS) was reported to be the cause of cardiovascular disease associated with chronic kidney disease. Therefore, we evaluated the direct influences of IS on vascular function, focusing on the superoxide anion (O2-) and nitric oxide (NO)/soluble guanylate cyclase (sGC) pathways. MAIN METHODS: Isolated rat thoracic aortas with and without vascular endothelium were incubated with IS for 4 h in a physiological solution. In some experiments, several inhibitors were treated 30 min before the addition of IS. O2- production was measured by the chemiluminescence method, and the vascular reactivity to different vasorelaxants was examined using organ chamber technique. KEY FINDINGS: 1) Experiments using endothelium-intact vascular rings: IS significantly increased O2- production. The increase was suppressed by addition of the NADPH oxidase inhibitor apocynin, the antioxidant ascorbic acid and the aryl hydrocarbon receptor (AhR) inhibitor CH223191. Furthermore, IS attenuated the acetylcholine (ACh)-induced vasorelaxantion, which was suppressed by addition of the above drugs. 2) Experiments using endothelium-denuded vascular rings: IS significantly increased O2- production and also attenuated sodium nitroprusside (SNP)-induced vasorelaxation. These influences of IS were normalized only by ascorbic acid addition. On the other hand, IS did not affect the vasorelaxation by the sGC stimulator BAY 41-2272. SIGNIFICANCE: This study suggested that IS causes O2- production in vascular tissues, thereby attenuating ACh- and SNP-induced vasorelaxation, probably through NO inactivation. Furthermore, it is reasonable to consider that IS-promoted O2- production in the presence of vascular endothelium is through binding to AhR and the activation of NADPH oxidase.

Sensitization of glutamate receptor-mediated pain behaviour via nerve growth factor-dependent phosphorylation of transient receptor potential V1 under inflammatory conditions.

BACKGROUND AND PURPOSE: Glutamate and metabotropic glutamate (mGlu) receptors on primary sensory neurons are crucial in modulating pain sensitivity. However, it is unclear how inflammation affects mGlu receptor-mediated nociceptive responses. We therefore investigated the effects of mGlu1/5 receptor agonists on pain-related behaviour during persistent inflammation and their underlying mechanisms. EXPERIMENTAL APPROACH: Effects of a mGlu1/5 receptor agonist on pain-related behaviour during inflammation was assessed in mice. Intracellular calcium responses, membrane current responses, and protein expression in primary sensory neurons were examined using cultured dorsal root ganglion (DRG) neurons, dissociated from wild-type and gene knockout mice. KEY RESULTS: Persistent inflammation induced by complete Freund's adjuvant increased the duration of mGlu1/5 receptor-mediated pain behaviour, which was antagonized by inhibition of nerve growth factor (NGF)-tropomyosin receptor kinase A (TrkA) signalling. Calcium imaging revealed that NGF treatment increased the number of cultured DRG neurons responding to mGlu1/5 receptor activation. Stimulation of mGlu1/5 receptors in NGF-treated DRG neurons induced inward currents through TRPV1 channels in association with PLC but not with IP3 receptors. NGF treatment also increased the number of neurons responding to a DAG analogue via TRPV1 channel activation. Furthermore, NGF up-regulated expression of TRPV1 and A-kinase anchoring protein 5 (AKAP5), resulting in increased AKAP5-dependent TRPV1 phosphorylation. AKAP5 knockout mice did not exhibit mGlu1/5 receptor-mediated excitation in NGF-treated DRG neurons or pain response facilitation under inflammatory conditions. CONCLUSIONS AND IMPLICATIONS: NGF augments glutamate- and mGlu1/5 receptor-mediated excitation of nociceptive neurons by AKAP5-dependent phosphorylation of TRPV1 channels, potentiating hypersensitivity to glutamate in inflamed tissues.

Effect of Betanin, a Beetroot Component, on Vascular Tone in Isolated Porcine Arteries.

BACKGROUND: Beetroot has attracted much attention because of its blood pressure-lowering properties. Although beetroot contains various nutritional compounds, including inorganic nitrate, some of their physiological properties are not fully understood. In this study, we examined whether betanin, a beetroot component, has a regulatory effect on vascular tone. METHODS: Mechanical responses of isolated porcine coronary, mesenteric, and pulmonary arteries were assessed by organ chamber technique. In some cases, the vascular reactivity was observed in the presence of a physiological concentration of betanin (10 µM). RESULTS: Betanin did not induce vasorelaxation at physiological concentrations both in endothelium-intact and -denuded coronary, mesenteric, and pulmonary arteries. The endothelium-dependent agonists, bradykinin and A23187 induced vasorelaxation of endothelium-intact coronary arteries, both of which were not affected by exposure to betanin. Likewise, endothelium-independent vasorelaxation induced by sodium nitrite and sodium nitroprusside was also not affected by the presence of betanin. In addition, exposure of endothelium-intact coronary arteries to betanin did not attenuate prostaglandin F2α- and endothelin-1-induced vasocontraction. CONCLUSIONS: These findings suggest that betanin does not have a vasorelaxant activity. It is unlikely that betanin is a component directly responsible for the beetroot-induced acute blood pressure-lowering effect in a nitrate-independent manner.

Impact of cigarette smoking on nitric oxide-sensitive and nitric oxide-insensitive soluble guanylate cyclase-mediated vascular tone regulation.

Cigarette smoking induces vascular endothelial dysfunction characterized by impaired nitric oxide (NO) bioavailability. There are two types of soluble guanylate cyclase (sGC), which is a cellular target of NO: NO-sensitive reduced form (the heme moiety with a ferrous iron) and NO-insensitive oxidized (the heme moiety with a ferric iron)/heme-free form. This study investigated the influence of cigarette smoking on NO-sensitive and NO-insensitive sGC-mediated vascular tone regulation in organ chamber experiments with isolated rat and human arteries. The rats were subcutaneously administered phosphate-buffered saline (PBS), nicotine-free cigarette smoke extract (N(-)-CSE) or nicotine-containing cigarette smoke extract (N(+)-CSE) for 4 weeks. Plasma thiobarbituric acid reactive substance (TBARS) levels were higher in the N(+)-CSE group than those in the N(-)-CSE group, and TBARS levels for these groups were higher than those for the PBS group. In the aorta and the pulmonary artery in rats administered N(-)-CSE or N(+)-CSE, acetylcholine-induced relaxation was significantly impaired compared with that in rats administered PBS; there was no significant difference in the relaxation between the N(-)-CSE and N(+)-CSE groups. However, sodium nitroprusside (NO-sensitive sGC stimulant)- and BAY 60-2770 (NO-insensitive sGC stimulant)-induced relaxations were not different among the three groups, regardless of the vessel type. In addition, in the human gastroepiploic artery, the relaxant responses to these sGC-targeting drugs were identical between nonsmokers and smokers. These findings suggest that NO-sensitive and NO-insensitive sGC-mediated vascular tone regulation functions normally even in blood vessels damaged by cigarette smoking.

Chronic Tear Deficiency Sensitizes Transient Receptor Potential Vanilloid 1-Mediated Responses in Corneal Sensory Nerves.

Chronic tear deficiency enhances the excitability of corneal cold-sensitive nerves that detect ocular dryness, which can lead to discomfort in patients with dry eye disease (DED). However, changes in corneal nerve excitations through the polymodal nociceptor "transient receptor potential vanilloid 1" (TRPV1) and the potential link between this receptor and symptoms of DED remain unclear. In this study, we examined the firing properties of corneal cold-sensitive nerves expressing TRPV1 and possible contributions of chronic tear deficiency to corneal nerve excitability by TRPV1 activation. The bilateral excision of lacrimal glands in guinea pigs decreased the tear volume and increased the frequency of spontaneous eyeblinks 1-4 weeks after surgery. An analysis of the firing properties of the cold-sensitive nerves was performed by single-unit recordings of corneal preparations 4 weeks after surgery in both the sham-operated and gland-excised groups. Perfusion of the TRPV1 agonist, capsaicin (1 µM), transiently increased the firing frequency in approximately 46-48% of the cold-sensitive nerves characterized by low-background activity and high threshold (LB-HT) cold thermoreceptors in both groups. Gland excision significantly decreased the latency of capsaicin-induced firing in cold-sensitive nerves; however, its magnitude was unchanged. Calcium imaging of cultured trigeminal ganglion neurons from both groups showed that intracellular calcium elevation of corneal neurons induced by a low concentration of capsaicin (0.03 µM) was significantly larger in the gland excision group, regardless of responsiveness to cold. An immunohistochemical study of the trigeminal ganglion revealed that gland excision significantly increased the proportion of corneal neurons enclosed by glial fibrillary acidic protein (GFAP)-immunopositive satellite glial cells. Topical application of the TRPV1 antagonist, A784168 (30 µM), on the ocular surface attenuated eye-blink frequency after gland excision. Furthermore, gland excision enhanced blink behavior induced by a low concentration of capsaicin (0.1 µM). These results suggest that chronic tear deficiency sensitizes the TRPV1-mediated response in the corneal LB-HT cold thermoreceptors and cold-insensitive polymodal nociceptors, which may be linked to dry eye discomfort and hyperalgesia resulting from nociceptive stimuli in aqueous-deficient dry eyes.

Involvement of γ-Glutamyl Transpeptidase in Ischemia/Reperfusion-Induced Cardiac Dysfunction in Isolated Rat Hearts.

GGsTop is a highly potent and specific, and irreversible γ-glutamyl transpeptidase (GGT) inhibitor without any influence on glutamine amidotransferases. The aim of the present study was to investigate the involvement of GGT in ischemia/reperfusion-induced cardiac dysfunction by assessing the effects of a treatment with GGsTop. Using a Langendorff apparatus, excised rat hearts underwent 40 min of global ischemia without irrigation and then 30 min of reperfusion. GGT activity was markedly increased in cardiac tissues exposed to ischemia, and was inhibited by the treatment with GGsTop. Exacerbation of cardiac functional parameters caused by ischemia and reperfusion, namely the reduction of left ventricular (LV) developed pressure and the maximum and negative minimum values of the first derivative of LV pressure, and the increment in LV end-diastolic pressure was significantly attenuated by GGsTop treatment. The treatment with GGsTop suppressed excessive norepinephrine release in the coronary perfusate, a marker for myocardial dysfunction, after ischemia/reperfusion. In addition, oxidative stress indicators in myocardium, including superoxide and malondialdehyde, after ischemia/reperfusion were significantly low in the presence of GGsTop. These observations demonstrate that enhanced GGT activity contributes to cardiac damage after myocardial ischemia/reperfusion, possibly via increased oxidative stress and subsequent norepinephrine overflow. GGT inhibitors have potential as a therapeutic strategy to prevent myocardial ischemia/reperfusion injury in vivo.

Chronological Change of Vascular Reactivity to cGMP Generators in the Balloon-Injured Rat Carotid Artery.

BACKGROUND/AIMS: Soluble guanylate cyclase (sGC) exists as reduced, oxidized, and heme-free forms. Currently, it is unclear whether endovascular mechanical stenosis has an impact on vascular tone control by drugs targeting sGC, namely cGMP generators. METHODS: Pharmacological responses to acidified sodium nitrite (reduced sGC stimulant) and BAY 60-2770 (oxidized/heme-free sGC stimulant) were studied in balloon-injured rat carotid arteries at several time points. In addition, sGC expression was detected by immunohistochemistry. RESULTS: At 1 day after injury, acidified sodium nitrite-induced relaxation was attenuated in the injured artery, whereas BAY 60-2770-induced relaxation was augmented. Similar attenuation of response to acidified sodium nitrite was seen at 7 and 14 days after injury. On the other hand, the augmentation of response to BAY 60-2770 disappeared at 7 and 14 days after injury. At 1 day after injury, the immunohistochemical expression pattern of sGC in the smooth muscle layer of the injured artery was not different from that of the uninjured artery. However, in the injured artery, the intensity of sGC staining was weak at 7 and 14 days after injury. CONCLUSION: Balloon injury alters vascular responsiveness to cGMP generators, which seems to be associated with the form and/or expression of sGC.

Responsiveness of rat aorta and pulmonary artery to cGMP generators in the presence of thiol or heme oxidant.

This study investigated the effects of thiol and heme oxidants on responsiveness to cGMP generators in isolated rat aorta and pulmonary artery using an organ chamber. The nitric oxide (NO) donor sodium nitroprusside (SNP)-induced relaxation was impaired by exposure to the thiol oxidant diamide in both the aorta and the pulmonary artery, whereas the soluble guanylate cyclase (sGC) stimulator BAY 41-2272- or the sGC activator BAY 60-2770-induced relaxation was not affected. The impairment by diamide of SNP-induced aortic and pulmonary arterial relaxation was completely restored by post-treatment with the thiol reductant dithiothreitol. However, regardless of the vessel type, the relaxant response to SNP or BAY 41-2272 was impaired by exposure to the heme oxidant ODQ, whereas the response to BAY 60-2770 was enhanced. The ODQ-induced effects were reversed partially by post-treatment with the heme reductant dithionite. These findings indicate that thiol oxidation attenuates only the vascular responsiveness to NO donors and that heme oxidation attenuates the responsiveness to NO donors and sGC stimulators but augments that to sGC activators. Therefore, under oxidative stress, the order of usability of the vasodilators is suggested to be: NO donors < sGC stimulators < sGC activators.

Effects of Beet Juice Supplementation on Monocrotaline-Induced Pulmonary Hypertension in Rats.

BACKGROUND: Recently, attention has been focused on the cardiovascular protective effects of beet juice (BJ) with high amounts of nitrate. In this study, we examined the effect of BJ supplementation in a rat model of monocrotaline (MCT)-induced pulmonary hypertension (PH). METHODS: MCT (60 mg/kg) was subcutaneously administered to rats, and BJ (prepared by dissolving BJ powder at a concentration of 1 g/l or 10 g/l in drinking water) supplementation was started from the day of, 1 week before, and 2 weeks after MCT injection. Saline-injected rats given drinking water were used as controls. RESULTS: Low-dose BJ supplementation starting from the day of MCT injection exerted protective effects on the MCT-induced elevation of right ventricular systolic pressure, right ventricular hypertrophy, and pulmonary arterial remodeling, without causing a significant increase in plasma nitrite plus nitrate (NOx) levels. On the other hand, such beneficial effects were not observed with high-dose BJ supplementation, although the NOx levels were slightly higher than those in the low-dose group. In addition, low-dose BJ supplementation starting from 1 week before MCT injection did not improve PH symptoms, as described above. Furthermore, low-dose BJ supplementation starting from 2 weeks after MCT injection was ineffective against functional and morphological alterations in pulmonary circulation associated with MCT-induced PH. CONCLUSIONS: Habitual ingestion of a suitable amount of BJ could be a potential option for preventing PH. However, beneficial effects cannot be expected when PH has developed to some degree.

Stimulation of nitric oxide-sensitive soluble guanylate cyclase in monocrotaline-induced pulmonary hypertensive rats.

AIMS: In this study, we examined whether a disruption in the balance between nitric oxide (NO)-sensitive and -insensitive soluble guanylate cyclase (sGC) is observed in pulmonary hypertension (PH) and whether treatment with NO-enhancing drugs can halt disease progression. MAIN METHODS: Rats were injected subcutaneously with saline or 60 mg/kg monocrotaline (MCT). At 14 days after injection, the vascular reactivity of isolated extralobar pulmonary arteries was assessed by organ chamber technique. In a separate experiment, isosorbide mononitrate (0.3 or 1 g/L) or sodium nitrite (30 or 300 mg/L) was administered in drinking water for the last 14 days (from day 15 to day 28), and their therapeutic potential was evaluated. KEY FINDINGS: The NO-sensitive sGC stimulant BAY 41-2272 and the NO-insensitive sGC stimulant BAY 60-2770 both relaxed the pulmonary arteries, which was comparable between saline- and MCT-injected rats. Treatment with isosorbide mononitrate suppressed the MCT-induced right ventricular systolic pressure (RVSP) elevation and pulmonary arterial medial thickening but not right ventricular hypertrophy. However, the beneficial effects on RVSP and pulmonary vascular remodeling were not observed when a high dose was administered. The same results were obtained following the sodium nitrite treatment. Interestingly, NO-enhancing drugs did not increase plasma nitrite plus nitrate levels at a dose that provided the greatest therapeutic advantage. SIGNIFICANCE: These findings suggest that the balance between NO-sensitive and -insensitive sGC is not disrupted in the early stage of MCT-induced PH. Furthermore, supplementation with an adequate amount of NO may be a useful therapy to prevent the progression of PH.

Responsiveness of internal thoracic arteries to nitroglycerin in patients with renal failure.

Nitroglycerin is commonly used as an antispasmodic for treating spasm of coronary artery bypass grafts. This study investigated whether the presence of renal failure affects reactivity to nitroglycerin in internal thoracic arteries obtained from patients undergoing coronary bypass surgery. The patients were divided into three groups according to estimated glomerular filtration rate (eGFR, mL/min/1.73 m2): without renal failure (60 ≤ eGFR, n = 13), with moderate renal failure (30 ≤ eGFR < 60, n = 10), and with severe renal failure (eGFR < 30, n = 10). Organ chamber technique was used to evaluate concentration-related responses of isolated internal thoracic arteries to vasodilators. Nitroglycerin induced a concentration-dependent relaxation, which was significantly augmented in patients with severe but not moderate renal failure than in those without renal failure. In addition, there was a negative correlation between eGFR and the relaxant efficacy of nitroglycerin (P = 0.016). On the other hand, relaxant responses to BAY 60-2770 (which enhances cGMP generation as with nitroglycerin) were similar among three grades of renal function. An inverse relationship of eGFR to the relaxant efficacy of BAY 60-2770 was not observed, either (P = 0.314). These findings suggest that severe renal failure specifically potentiates nitroglycerin-induced relaxation in internal thoracic artery grafts.

Segmental Difference in Vasoreactivity of the Human Right Gastroepiploic Artery.

BACKGROUND: The gastroepiploic artery (GEA) plays an important role in the era of multiple arterial revascularization, but spasm is a major matter of concern. The internal thoracic artery has been shown to have a strong tendency to spasm in its distal bifurcating part, whereas the segmental difference in vasoreactivity of the GEA has never been performed.Methods and Results:The full length of the GEA obtained from 21 patients undergoing a total gastrectomy was divided into 3 sections: proximal (5 cm from the origin), middle, and distal (5 cm from the end). Concentration-response curves for vasoconstrictors (phenylephrine, prostaglandin F2α, and endothelin-1) and vasodilators (carperitide, nitroglycerin, and nifedipine) were then established using organ baths. All the vasoconstrictors and vasodilators produced concentration-dependent responses in each section. As the concentration of the vasoconstrictors increased, segments at the distal section showed a significantly greater contraction than those at the middle and proximal sections regardless of the type of vasoconstrictor. The effective concentration of drugs that caused 50% of the maximal response for endothelin-1 was significantly greater in the distal section than that in the proximal sections. No significant difference was found in vasodilators-induced relaxation. CONCLUSIONS: The contractility increases toward to the end of the GEA. Clinically, the distal portion of the GEA should be trimmed off and not be used as an anastomotic site wherever possible.