Bradykinin-induced Vasodilation Research Articles

Association of cerebral microvascular dysfunction and white matter injury in Alzheimer's disease.

Patients with Alzheimer's disease (AD) often have cerebral white matter (WM) hyperintensities on MRI and microinfarcts of presumed microvascular origin pathologically. Here, we determined if vasodilator dysfunction of WM-penetrating arterioles is associated with pathologically defined WM injury and disturbances in quantitative MRI-defined WM integrity in patients with mixed microvascular and AD pathology. We analyzed tissues from 28 serially collected human brains from research donors diagnosed with varying degrees of AD neuropathologic change (ADNC) with or without cerebral microinfarcts (mVBI). WM-penetrating and pial surface arteriolar responses to the endothelium-dependent agonist bradykinin were quantified ex vivo with videomicroscopy. Vascular endothelial nitric oxide synthase (eNOS) and NAD(P)H-oxidase (Nox1,2 and 4 isoforms) expression were measured with quantitative PCR. Glial fibrillary acidic protein (GFAP)-labeled astrocytes were quantified by unbiased stereological approaches in regions adjacent to the sites of WM-penetrating vessel collection. Post-mortem diffusion tensor imaging (DTI) was used to measure mean apparent diffusion coefficient (ADC) and fractional anisotropy (FA), quantitative indices of WM integrity. In contrast to pial surface arterioles, white matter-penetrating arterioles from donors diagnosed with high ADNC and mVBI exhibited a significantly reduced dilation in response to bradykinin when compared to the other groups. Expression of eNOS was reduced, whereas Nox1 expression was increased in WM arterioles in AD and mVBI cases. WM astrocyte density was increased in AD and mVBI, which correlated with a reduced vasodilation in WM arterioles. Moreover, in cases with low ADNC, bradykinin-induced WM arteriole dilation correlated with lower ADC and higher FA values. Comorbid ADNC and mVBI appear to synergistically interact to selectively impair bradykinin-induced vasodilation in WM-penetrating arterioles, which may be related to reduced nitric oxide- and excess reactive oxygen species-mediated vascular endothelial dysfunction. WM arteriole vasodilator dysfunction is associated with WM injury, as supported by reactive astrogliosis and MRI-defined disrupted WM microstructural integrity.

Hepatic Encephalopathy-Associated Cerebral Vasculopathy in Acute-on-Chronic Liver Failure: Alterations on Endothelial Factor Release and Influence on Cerebrovascular Function.

The acute-on-chronic liver failure (ACLF) is a syndrome characterized by liver decompensation, hepatic encephalopathy (HE) and high mortality. We aimed to determine the mechanisms implicated in the development of HE-associated cerebral vasculopathy in a microsurgical liver cholestasis (MHC) model of ACLF. Microsurgical liver cholestasis was induced by ligating and extracting the common bile duct and four bile ducts. Sham-operated and MHC rats were maintained for eight postoperative weeks Bradykinin-induced vasodilation was greater in middle cerebral arteries from MHC rats. Both Nω-Nitro-L-arginine methyl ester and indomethacin diminished bradykinin-induced vasodilation largely in arteries from MHC rats. Nitrite and prostaglandin (PG) F1α releases were increased, whereas thromboxane (TX) B2 was not modified in arteries from MHC. Expressions of endothelial nitric oxide synthase (eNOS), inducible NOS, and cyclooxygenase (COX) 2 were augmented, and neuronal NOS (nNOS), COX-1, PGI2 synthase, and TXA2S were unmodified. Phosphorylation was augmented for eNOS and unmodified for nNOS. Altogether, these endothelial alterations might collaborate to increase brain blood flow in HE.

Does eNOS derived nitric oxide protect the young from severe COVID-19 complications?

The COVID-19 pandemic poses an imminent threat to humanity, especially to the elderly. The molecular mechanisms underpinning the age-dependent disparity for disease progression is not clear. COVID-19 is both a respiratory and a vascular disease in severe patients. The damage endothelial system provides a good explanation for the various complications seen in COVID-19 patients. These observations lead us to suspect that endothelial cells are a barrier that must be breached before progression to severe disease. Endothelial intracellular defences are largely dependent of the activation of the interferon (IFN) system. Nevertheless, low type I and III IFNs are generally observed in COVID-19 patients suggesting that other intracellular viral defence systems are also activated to protect the young. Intriguingly, Nitric oxide (NO), which is the main intracellular antiviral defence, has been shown to inhibit a wide array of viruses, including SARS-CoV-1. Additionally, the increased risk of death with diseases that have underlying endothelial dysfunction suggest that endothelial NOS-derived nitric oxide could be the main defence mechanism. NO decreases dramatically in the elderly, the hyperglycaemic and the patients with low levels of vitamin D. However, eNOS derived NO occurs at low levels, unless it is during inflammation and co-stimulated by bradykinin. Regrettably, the bradykinin-induced vasodilation also progressively declines with age, thereby decreasing anti-viral NO production as well. Intriguingly, the inverse correlation between the percentage of WT eNOS haplotype and death per 100K population could potentially explain the disparity of COVID-19 mortality between Asian and non-Asian countries. These changes with age, low bradykinin and NO, may be the fundamental reasons that intracellular innate immunity declines with age leading to more severe COVID-19 complications.

Acute vascular effects of vascular endothelial growth factor inhibition in the forearm arterial circulation.

Objective:Although vascular endothelial growth factor inhibition (VEGFi) represents a major therapeutic advance in oncology, it is associated with hypertension and adverse vascular thrombotic events. Our objective was to determine whether VEGFi caused direct vascular dysfunction through increased endothelin-1 (ET-1) activity or impaired endothelial vasomotor or fibrinolytic function.Methods:Using forearm venous occlusion plethysmography, we measured forearm blood flow during intra-arterial infusions of bevacizumab (36–144 μg/dl forearm volume per minute) administered for 15–60 min in healthy volunteers (n = 6–8). On two separate occasions in 10 healthy volunteers, we further measured forearm blood flow and tissue plasminogen activator (t-PA) release during intra-arterial bradykinin infusion (100 and 1000 pmol/min) in the presence and absence of bevacizumab (144 μg/dl forearm volume per minute), and the presence and absence of endothelin A receptor antagonism with BQ-123 (10 nmol/min). Plasma t-PA and plasminogen activator inhibitor-1 (PAI-1) concentrations were measured at baseline and with each dose of bradykinin.Results:Baseline blood flow and plasma ET-1, t-PA and PAI-1 concentrations were unaffected by bevacizumab. Bradykinin caused dose-dependent vasodilatation (P < 0.0001) and t-PA release (P < 0.01) but had no effect on plasma PAI-1 concentrations. Neither bevacizumab nor BQ-123 affected bradykinin-induced vasodilatation and t-PA release.Conclusion:Acute exposure to bevacizumab does not directly cause endothelial vasomotor or fibrinolytic dysfunction in healthy young volunteers.

Pharmacological Targeting of KCa Channels to Improve Endothelial Function in the Spontaneously Hypertensive Rat

Systemic hypertension is a major risk factor for the development of cardiovascular disease and is often associated with endothelial dysfunction. KCa2.3 and KCa3.1 channels are expressed in the vascular endothelium and contribute to stimulus-evoked vasodilation. We hypothesized that acute treatment with SKA-31, a selective activator of KCa2.x and KCa3.1 channels, would improve endothelium-dependent vasodilation and transiently lower mean arterial pressure (MAP) in male, spontaneously hypertensive rats (SHRs). Isolated vascular preparations exhibited impaired vasodilation in response to bradykinin (i.e., endothelial dysfunction) compared with Wistar controls, which was associated with decreased bradykinin receptor expression in mesenteric arteries. In contrast, similar levels of endothelial KCa channel expression were observed, and SKA-31 evoked vasodilation was comparable in vascular preparations from both strains. Addition of a low concentration of SKA-31 (i.e., 0.2–0.3 μM) failed to augment bradykinin-induced vasodilation in arteries from SHRs. However, responses to acetylcholine were enhanced. Surprisingly, acute bolus administration of SKA-31 in vivo (30 mg/kg, i.p. injection) modestly elevated MAP compared with vehicle injection. In summary, pharmacological targeting of endothelial KCa channels in SHRs did not readily reverse endothelial dysfunction in situ, or lower MAP in vivo. SHRs thus appear to be less responsive to endothelial KCa channel activators, which may be related to their vascular pathology.

Ascorbate blocks acetylcholine- and bradykinin-induced vasodilatation in retinal vascular bed of the bovine isolated perfused eye

Ascorbate blocks acetylcholine- and bradykinin-induced vasodilatation in retinal vascular bed of the bovine isolated perfused eye

Combination of β Adrenergic Receptor Block and Renin-Angiotensin System Inhibition Diminished the Angiotensin II-Induced Vasoconstriction and Increased Bradykinin-Induced Vasodilation in Hypertension.

In hypertension, the combination therapy is frequently used to obtain a better therapeutic effect and reduce adverse effects. One effective combination is with inhibitors and β-blockers of renin–angiotensin system. Although the mechanisms of action of each drug are already known, the antihypertensive mechanism is more complex and therefore the combined treatment mechanism is unclear. Specifically, the effect of the treatments of angiotensin-converting enzyme inhibitor or AT1 receptor antagonist with β-blocker on the angiotensin II and bradykinin reactivity has not been studied. For this reason, we evaluated the interaction between propranolol and captopril or losartan on vascular reactivity to bradykinin and angiotensin II in spontaneously hypertensive rat. We constructed concentration–response curves to angiotensin II and bradykinin after treatment of SHR with propranolol–captopril or propranolol–losartan by using rat aortic rings. While losartan or captopril with propranolol potentiated bradykinin-induced vasodilation effect, the propranolol–losartan interaction decreased the angiotensin II-induced vasoconstriction. In addition, the combinations did not reduce the heart rate significantly. These results suggest that the combined therapy decreased blood pressure to normotensive values and showed less effect for angiotensin II and greater effect for bradykinin than monotherapy which could contribute in the antihypertensive effect.

Cardiometabolic effects of a novel SIRT1 activator, SRT2104, in people with type 2 diabetes mellitus

BackgroundThe cardiometabolic effects of SRT2104, a novel SIRT1 activator, were investigated in people with type 2 diabetes mellitus (T2DM).MethodsFifteen adults with T2DM underwent a randomised, double-blind, placebo-controlled cross-over trial and...

Sex differences in pulmonary vascular control: focus on the nitric oxide pathway.

Although the incidence of pulmonary hypertension is higher in females, the severity and prognosis of pulmonary vascular disease in both neonates and adults have been shown to be worse in male subjects. Studies of sex differences in pulmonary hypertension have mainly focused on the role of sex hormones. However, the contribution of sex differences in terms of vascular signaling pathways regulating pulmonary vascular function remains incompletely understood. Consequently, we investigated pulmonary vascular function of male and female swine in vivo, both at rest and during exercise, and in isolated small pulmonary arteries in vitro, with a particular focus on the NO‐cGMP‐PDE5 pathway. Pulmonary hemodynamics at rest and during exercise were virtually identical in male and female swine. Moreover, NO synthase inhibition resulted in a similar degree of pulmonary vasoconstriction in male and female swine. However, NO synthase inhibition blunted bradykinin‐induced vasodilation in pulmonary small arteries to a greater extent in male than in female swine. PDE5 inhibition resulted in a similar degree of vasodilation in male and female swine at rest, while during exercise there was a trend towards a larger effect in male swine. In small pulmonary arteries, PDE5 inhibition failed to augment bradykinin‐induced vasodilation in either sex. Finally, in the presence of NO synthase inhibition, the pulmonary vasodilator effect of PDE5 inhibition was significantly larger in female swine both in vivo and in vitro. In conclusion, the present study demonstrated significant sex differences in the regulation of pulmonary vascular tone, which may contribute to understanding sex differences in incidence, treatment response, and prognosis of pulmonary vascular disease.

Mechanisms of Vascular Dysfunction in COPD and Effects of a Novel Soluble Epoxide Hydrolase Inhibitor in Smokers

BackgroundSmoking and COPD are risk factors for cardiovascular disease, and the pathogenesis may involve endothelial dysfunction. We tested the hypothesis that endothelium-derived epoxyeicosatrienoic acid (EET)-mediated endothelial function is impaired in patients with COPD and that a novel soluble epoxide hydrolase inhibitor, GSK2256294, attenuates EET-mediated endothelial dysfunction in human resistance vessels both in vitro and in vivo.MethodsEndogenous and stimulated endothelial release of EETs was assessed in 12 patients with COPD, 11 overweight smokers, and two matched control groups, using forearm plethysmography with intraarterial infusions of fluconazole, bradykinin, and the combination. The effects of GSK2256294 on EET-mediated vasodilation in human resistance arteries were assessed in vitro and in vivo in a phase I clinical trial in healthy overweight smokers.ResultsCompared with control groups, there was reduced vasodilation with bradykinin (P = .005), a blunted effect of fluconazole on bradykinin-induced vasodilation (P = .03), and a trend toward reduced basal EET/dihydroxyepoxyeicosatrienoic acid ratio in patients with COPD (P = .08). A similar pattern was observed in overweight smokers. In vitro, 10 μM GSK2256294 increased 11,12-EET-mediated vasodilation compared with vehicle (90% ± 4.2% vs 72.6% ± 6.2% maximal dilatation) and shifted the bradykinin half-maximal effective concentration (EC50) (–8.33 ± 0.172 logM vs –8.10 ± 0.118 logM; P = .001 for EC50). In vivo, 18 mg GSK2256294 improved the maximum bradykinin response from 338% ± 46% before a dose to 566% ± 110% after a single dose (P = .02) and to 503% ± 123% after a chronic dose (P = .003).ConclusionsGSK2256294 attenuates smoking-related EET-mediated endothelial dysfunction, suggesting potential therapeutic benefits in patients with COPD.Trial RegistryClinicalTrials.gov; No.: NCT01762774; URL: www.clinicaltrials.gov

Bradykinin –induced vasodilatation: Role of age, ACE1-inhibitory peptide, mas- and bradykinin receptors

Bradykinin exerts its vascular actions via two types of receptors, the non-constitutively expressed bradykinin receptor type 1 (BR1) and the constitutive type 2 receptor (BR2). Bradykinin-induced vasorelaxation is age-dependent, a phenomenon related to the varying amounts of BR1 and BR2 in the vasculature.Isoleucine-proline-proline (Ile-Pro-Pro), a bioactive tripeptide, lowers elevated blood pressure and improves impaired endothelium-dependent vasorelaxation in hypertensive rats. It inhibits angiotensin converting enzyme 1 (ACE1). Other mechanisms of action have also been postulated.The aims of the study were to clarify the underlying mechanisms of the age-dependency of bradykinin-induced vasodilatation such as the roles of the two bradykinin receptors, the mas-receptor and synergism with Ile-Pro-Pro.The vascular response studies were conducted using mesenteric artery and aorta rings from normotensive 6 wk. (young) and 22 wk. (old) Wistar rats. Cumulative dosing of acetylcholine, bradykinin and angiotensin(1–7) (Ang(1–7))were tested in phenylephrine-induced vasoconstriction with or without 10min pre-incubation with antagonists against BR1-, BR2- or mas-receptors, Ang(1–7) or ACE1-inhibitors captopril and Ile-Pro-Pro.The bradykinin-induced vasorelaxation in vitro was age-dependent and it was improved by pre-incubation with Ile-Pro-Pro, especially in old rats with endothelial dysfunction. The mas-receptor antagonist, D-Pro7-Ang(1–7) abolished bradykinin-induced relaxation totally. Interestingly, BR1 and BR2 antagonists only slightly reduced bradykinin-induced vasorelaxation, as an evidence for the involvement of other mechanisms in addition to receptor activation.In conclusion, bradykinin-induced vasorelaxation was age-dependent and Ile-Pro-Pro improved it. Mas receptor antagonist abolished relaxation while bradykinin receptor antagonist only slightly reduced it, suggesting that bradykinin-induced vasorelaxation is regulated also by other mechanisms than the classical BR1/BR2 pathway.

Asymmetric dimethylarginine (ADMA) elevation and arginase up-regulation contribute to endothelial dysfunction related to insulin resistance in rats and morbidly obese humans.

The presence of insulin resistance (IR) is determinant for endothelial dysfunction associated with obesity. Although recent studies have implicated the involvement of mitochondrial superoxide and inflammation in the defective nitric oxide (NO)-mediated responses and subsequent endothelial dysfunction in IR, other mechanisms could compromise this pathway. In the present study, we assessed the role of asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of endothelium-dependent vasodilatation in human morbid obesity and in a non-obese rat model of IR. We show that both increased ADMA and up-regulated arginase are determinant factors in the alteration of the l-arginine/NO pathway associated with IR in both models and also that acute treatment of arteries with arginase inhibitor or with l-arginine significantly alleviate endothelial dysfunction. These results help to expand our knowledge regarding the mechanisms of endothelial dysfunction that are related to obesity and IR and establish potential therapeutic targets for intervention. Insulin resistance (IR) is determinant for endothelial dysfunction in human obesity. Although we have previously reported the involvement of mitochondrial superoxide and inflammation, other mechanisms could compromise NO-mediated responses in IR. We evaluated the role of the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA) and arginase with respect to IR-induced impairment of l-arginine/NO-mediated vasodilatation in human morbid obesity and in a non-obese rat model of IR. Bradykinin-induced vasodilatation was evaluated in microarteries derived from insulin-resistant morbidly obese (IR-MO) and non-insulin-resistant MO (NIR-MO) subjects. Defective endothelial vasodilatation in IR-MO was improved by l-arginine supplementation. Increased levels of ADMA were detected in serum and adipose tissue from IR-MO. Serum ADMA positively correlated with IR score and negatively with pD2 for bradykinin. Gene expression determination by RT-PCR revealed not only the decreased expression of ADMA degrading enzyme dimethylarginine dimethylaminohydrolase (DDAH)1/2 in IR-MO microarteries, but also increased expression of arginase-2. Arginase inhibition improved endothelial vasodilatation in IR-MO. Analysis of endothelial vasodilatation in a non-obese IR model (fructose-fed rat) confirmed an elevation of circulating and aortic ADMA concentrations, as well as reduced DDAH aortic content and increased aortic arginase activity in IR. Improvement of endothelial vasodilatation in IR rats by l-arginine supplementation and arginase inhibition provided functional corroboration. These results demonstrate that increased ADMA and up-regulated arginase contribute to endothelial dysfunction as determined by the presence of IR in human obesity, most probably by compromising arginine availability. The results provide novel insights regarding the mechanisms of endothelial dysfunction related to obesity and IR and establish potential therapeutic targets for intervention.

Metformin as a prevention and treatment for preeclampsia: effects on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion and endothelial dysfunction

Preeclampsia is associated with placental ischemia/hypoxia and secretion of soluble fms-like tyrosine kinase 1 and soluble endoglin into the maternal circulation. This causes widespread endothelial dysfunction that manifests clinically as hypertension and multisystem organ injury. Recently, small molecule inhibitors of hypoxic inducible factor 1α have been found to reduce soluble fms-like tyrosine kinase 1 and soluble endoglin secretion. However, their safety profile in pregnancy is unknown. Metformin is safe in pregnancy and is also reported to inhibit hypoxic inducible factor 1α by reducing mitochondrial electron transport chain activity. The purposes of this study were to determine (1) the effects of metformin on placental soluble fms-like tyrosine kinase 1 and soluble endoglin secretion, (2) to investigate whether the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion are regulated through the mitochondrial electron transport chain, and (3) to examine its effects on endothelial dysfunction, maternal blood vessel vasodilation, and angiogenesis. We performed functional (in vitro and ex vivo) experiments using primary human tissues to examine the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion from placenta, endothelial cells, and placental villous explants. We used succinate, mitochondrial complex II substrate, to examine whether the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin secretion were mediated through the mitochondria. We also isolated mitochondria from preterm preeclamptic placentas and gestationally matched control subjects and measured mitochondrial electron transport chain activity using kinetic spectrophotometric assays. Endothelial cells or whole maternal vessels were incubated with metformin to determine whether it rescued endothelial dysfunction induced by either tumor necrosis factor-α (to endothelial cells) or placenta villous explant-conditioned media (to whole vessels). Finally, we examined the effects of metformin on angiogenesis on maternal omental vessel explants. Metformin reduced soluble fms-like tyrosine kinase 1 and soluble endoglin secretion from primary endothelial cells, villous cytotrophoblast cells, and preterm preeclamptic placental villous explants. The reduction in soluble fms-like tyrosine kinase 1 and soluble endoglin secretion was rescued by coadministration of succinate, which suggests that the effects of metformin on soluble fms-like tyrosine kinase 1 and soluble endoglin were likely to be regulated at the level of the mitochondria. In addition, the mitochondrial electron transport chain inhibitors rotenone and antimycin reduced soluble fms-like tyrosine kinase 1 secretion, which further suggests that soluble fms-like tyrosine kinase 1 secretion is regulated through the mitochondria. Mitochondrial electron transport chain activity in preterm preeclamptic placentas was increased compared with gestation-matched control subjects. Metformin improved features of endothelial dysfunction relevant to preeclampsia. It reduced endothelial cell messenger RNA expression of vascular cell adhesion molecule 1 that was induced by tumor necrosis factor-α (vascular cell adhesion molecule 1 is an inflammatory adhesion molecule up-regulated with endothelial dysfunction and is increased in preeclampsia). Placental conditioned media impaired bradykinin-induced vasodilation; this effect was reversed by metformin. Metformin also improved whole blood vessel angiogenesis impaired by fms-like tyrosine kinase 1. Metformin reduced soluble fms-like tyrosine kinase 1 and soluble endoglin secretion from primary human tissues, possibly by inhibiting the mitochondrial electron transport chain. The activity of the mitochondrial electron transport chain was increased in preterm preeclamptic placenta. Metformin reduced endothelial dysfunction, enhanced vasodilation in omental arteries, and induced angiogenesis. Metformin has potential to prevent or treat preeclampsia.

Microcirculation and left-ventricular hypertrophy

Microcirculation and left-ventricular hypertrophy

CytochromeP-450 2C9 exerts a vasoconstrictor influence on coronary resistance vessels in swine at rest and during exercise

A significant endothelium-dependent vasodilation persists after inhibition of nitric oxide synthase (NOS) and cyclooxygenase (COX) in the coronary vasculature, which has been linked to the activation of cytochrome P-450 (CYP) epoxygenases expressed in endothelial cells and subsequent generation of vasodilator epoxyeicosatrienoic acids. Here, we investigated the contribution of CYP 2C9 metabolites to regulation of porcine coronary vasomotor tone in vivo and in vitro. Twenty-six swine were chronically instrumented. Inhibition of CYP 2C9 with sulfaphenazole (5 mg/kg iv) alone had no effect on bradykinin-induced endothelium-dependent coronary vasodilation in vivo but slightly attenuated bradykinin-induced vasodilation in the presence of combined NOS/COX blockade with N(ω)-nitro-L-arginine (20 mg/kg iv) and indomethacin (10 mg/kg iv). Sulfaphenazole had minimal effects on coronary resistance vessel tone at rest or during exercise. Surprisingly, in the presence of combined NOS/COX blockade, a significant coronary vasodilator response to sulfaphenzole became apparent, both at rest and during exercise. Subsequently, we investigated in isolated porcine coronary small arteries (∼250 μm) the possible involvement of reactive oxygen species (ROS) in the paradoxical vasoconstrictor influence of CYP 2C9 activity. The vasodilation by bradykinin in vitro in the presence of NOS/COX blockade was markedly potentiated by sulfaphenazole under control conditions but not in the presence of the ROS scavenger N-(2-mercaptoproprionyl)-glycine. In conclusion, CYP 2C9 can produce both vasoconstrictor and vasodilator metabolites. Production of these metabolites is enhanced by combined NOS/COX blockade and is critically dependent on the experimental conditions. Thus production of vasoconstrictors slightly outweighed the production of vasodilators at rest and during exercise. Pharmacological stimulation with bradykinin resulted in vasodilator CYP 2C9 metabolite production when administered in vivo, whereas vasoconstrictor CYP 2C9 metabolites, most likely ROS, were dominant when administered in vitro.

Role of Calcium-Activated Potassium Channels with Small Conductance in Bradykinin-Induced Vasodilation of Porcine Retinal Arterioles

Endothelial dysfunction and impaired vasodilation may be involved in the pathogenesis of retinal vascular diseases. In the present study, the mechanisms underlying bradykinin vasodilation were examined and whether calcium-activated potassium channels of small (SK(Ca)) and intermediate (IK(Ca)) conductance are involved in regulation of endothelium-dependent vasodilation in retinal arterioles was investigated. Porcine retinal arterioles (diameter approximately 112 microm, N = 119) were mounted in microvascular myographs for isometric tension recordings. The arterioles were contracted with the thromboxane analogue, U46619, and concentration-response curves were constructed for bradykinin and a novel opener of SK(Ca) and IK(Ca) channels, NS309. In U46619-contracted arterioles, bradykinin and NS309 induced concentration-dependent relaxations. In vessels without endothelium, bradykinin relaxation was abolished and NS309 relaxation was attenuated. Inhibition of NO synthase with asymmetric dimethylarginine and/or cyclooxygenase with indomethacin markedly reduced bradykinin and NS309 relaxation. NO synthase and cyclooxygenase inhibition together with oxyhemoglobin abolished bradykinin relaxation and attenuated NS309 relaxation. Blocking of SK(Ca) and IK(Ca) channels with apamin plus charybdotoxin or blocking of SK(Ca) channels alone in the absence and the presence of indomethacin markedly reduced bradykinin and NS309 relaxation, whereas blocking of IK(Ca) channels had no significant effect. In vessels without endothelium, blocking of SK(Ca) channels alone had no effect on sodium nitroprusside-induced relaxation. In porcine retinal arterioles, NO and prostaglandins mediate endothelium-dependent relaxation to bradykinin and NS309. Moreover, these findings suggest that SK(Ca) channels contribute to NO-mediated relaxation induced by bradykinin and NS309 and, hence, may play an important role in retinal arterial endothelial function.

Vascular B1 Kinin Receptors in Patients With Congestive Heart Failure

Animal models suggest a vasomotor role for the B1 kinin receptor in cardiovascular disease states. In patients with heart failure treated with angiotensin-converting enzyme inhibition (ACEi), or combined B1/B2 receptor antagonism, but not B2 receptor antagonism alone, causes vasoconstriction. However, B1 agonism has no effect on vasomotor or fibrinolytic function. Findings from transgenic animals lacking the B2 receptor suggest that these conflicting data may be explained by cross-talk between B1 and B2 receptors. We hypothesized that B1 stimulation causes vasodilatation and tissue plasminogen activator release in the human forearm when B2 receptor signaling is inhibited. Forearm blood flow was measured in 16 patients with heart failure receiving ACEi. In double-blinded crossover studies, intrabrachial Lys-[Leu8]-des-Arg9-bradykinin (B1 antagonist), lys-des-Arg9-bradykinin (B1 agonist), bradykinin (B2 agonist), and sodium nitroprusside (endothelium-independent vasodilator) were infused alone or with HOE-140 (B2 antagonist). HOE-140 did not affect basal vascular tone or t-PA release, but it abolished bradykinin-induced vasodilatation and t-PA release (P < 0.0001). Blood flow and t-PA release were unaffected by B1 agonism or antagonism in the presence and absence HOE-140. Our findings do not support a role for crosstalk between the B1 and B2 kinin receptors in the human peripheral circulation.

Potential role of vasomotor effects of fibrinogen in bradykinin-induced angioedema

Although bradykinin is known to play a major role in the pathophysiology of hereditary and angiotensin-converting enzyme inhibitor (ACEi)-induced angioedema, other factors acting as triggers or enhancers are likely important as well. We hypothesized that fibrinogen might contribute to ACEi-induced angioedema (eg, through direct actions on vascular tone). Plasma levels of fibrinogen were determined in 59 patients with acute angioedema. Vascular activity of human and bovine fibrinogen and its effects on bradykinin-induced vasodilation and phosphorylation of vasodilator-stimulated phosphoprotein were investigated in small (0.8-1.4 mm in diameter) porcine coronary artery and human internal thoracic artery (ITA) segments. In patients with ACEi-induced angioedema, fibrinogen levels (481 +/- 22 mg/dL, n = 39) were significantly higher than in patients with idiopathic angioedema (302 +/- 15 mg/dL, P < .001). Fibrinogen (1-15 mumol/L) induced a concentration-dependent vasodilation in preconstricted small porcine coronary arteries (n = 13), reaching a maximum vasodilator effect of 70% +/- 4.7%. Likewise, fibrinogen induced a 52.1% +/- 9.1% (n = 7) vasodilation in ITA rings. Fibrinogen vasorelaxations were completely inhibited by abciximab and diminished by endothelial denudation and treatment with the nitric oxide synthase inhibitor L-nitroargininemethylester and glibenclamide (P < .01). Importantly, fibrinogen increased the vasodilator potency of bradykinin by 10-fold (P < .0001) and increased bradykinin-induced vasodilator-stimulated phosphoprotein phosphorylation (P < .01). The increase of plasma fibrinogen levels, its vasodilator activity in human ITAs, and the potentiation of bradykinin-induced vasodilation suggest that fibrinogen might contribute to the pathophysiology of ACEi-induced angioedema. Thus acute-phase proteins, such as fibrinogen, might be viewed as risk factors for bradykinin-induced angioedema.

TNF-α Alters Nitric Oxide- and Endothelium-Derived Hyperpolarizing Factor-Mediated Vasodilatation in Human Omental Arteries

Objective: To investigate the effect of TNF-α on the endothelial function of human systemic arteries. Methods: Omental arteries were obtained from healthy pregnant women undergoing Cesarean section and examined using isometric wire myography. Results: Incubation with TNF-α (1nM) alone did not alter bradykinin-mediated endothelium-dependent relaxation of arteries. However, TNF-α did attenuate nitric oxide- (NO) and prostacyclin-independent endothelial-mediated relaxation. Similarly, in vessels constricted with a high potassium solution (60 mM), which inhibits vasodilatation via endothelial-derived hyperpolarising factor (EDHF), TNF-α incubation also attenuated bradykinin-induced vasodilatation. Conclusions: The vasorelaxant capacity of human systemic arteries is compromised by TNF-α incubation in the presence of NO/prostacyclin or EDHF-blockade.

Melatonin inhibits nitric oxide production by microvascular endothelial cells in vivo and in vitro

We have previously shown that melatonin inhibits bradykinin-induced NO production by endothelial cells in vitro. The purpose of this investigation was to extend this observation to an in vivo condition and to explore the mechanism of action of melatonin. RT-PCR assays were performed with rat cultured endothelial cells. The putative effect of melatonin upon arteriolar tone was investigated by intravital microscopy while NO production by endothelial cells in vitro was assayed by fluorimetry, and intracellular Ca(2+) measurements were assayed by confocal microscopy. No expression of the mRNA for the melatonin synthesizing enzymes, arylalkylamine N-acetyltransferase and hydroxyindole-O-methyltransferase, or for the melatonin MT(2) receptor was detected in microvascular endothelial cells. Melatonin fully inhibited L-NAME-sensitive bradykinin-induced vasodilation and also inhibited NO production induced by histamine, carbachol and 2-methylthio ATP, but did not inhibit NO production induced by ATP or alpha, beta-methylene ATP. None of its inhibitory effects was prevented by the melatonin receptor antagonist, luzindole. In nominally Ca(2+)-free solution, melatonin reduced intracellular Ca(2+) mobilization induced by bradykinin (40%) and 2-methylthio ATP (62%) but not Ca(2+) mobilization induced by ATP. We have confirmed that melatonin inhibited NO production both in vivo and in vitro. In addition, the melatonin effect was selective for some G protein-coupled receptors and most probably reflects an inhibition of Ca(2+) mobilization from intracellular stores.