Combined glucocorticoid resistance and hyperlactatemia contributes to lethal shock in sepsis.

Sepsis is a potentially lethal syndrome resulting from a maladaptive response to infection. Upon infection, glucocorticoids are produced as a part of the compensatory response to tolerate sepsis. This tolerance is, however, mitigated in sepsis due to a quickly induced glucocorticoid resistance at the level of the glucocorticoid receptor. Here, we show that defects in the glucocorticoid receptor signaling pathway aggravate sepsis pathophysiology by lowering lactate clearance and sensitizing mice to lactate-induced toxicity. The latter is exerted via an uncontrolled production of vascular endothelial growth factor, resulting in vascular leakage and collapse with severe hypotension, organ damage, and death, all being typical features of a lethal form of sepsis. In conclusion, sepsis leads to glucocorticoid receptor failure and hyperlactatemia, which collectively leads to a lethal vascular collapse.

Characterization of the retina-induced relaxation in mice.

PURPOSE: The retinal relaxing factor (RRF) is a continuously released factor from the retina that causes vasorelaxation, the identity and potential role in physiology of which remain largely unknown. Experiments were performed to find out whether the RRF-induced relaxation is influenced by serotonin, glutamate, L-cysteine, the cytochrome P450 pathway, the cyclooxygenase pathway, or oxidative stress. In addition, the sensitivity of retinal and non-retinal arteries towards the RRF was compared. METHODS: In vitro tension measurements were performed on isolated mouse femoral or bovine retinal arteries to study the vasorelaxing effect of the RRF, induced by mouse or bovine retinas. RESULTS: The presence of serotonin, glutamate, or L-cysteine did not alter the RRF-induced relaxation. Increasing oxidative stress by hydroquinone and diethyldithiocarbamic acid sodium salt enhanced the RRF response. Inhibition of the cytochrome P450 or the cyclooxygenase pathway did not cause any alteration. Surprisingly, the RRF-induced relaxation was enhanced by the presence of flufenamic acid or carbenoxolone. Furthermore, bringing retinal tissue in close contact with retinal or non-retinal arteries induced comparable relaxations. CONCLUSIONS: Serotonin, glutamate, L-cysteine, the cytochrome P450, and the cyclooxygenase pathway do not influence the RRF-induced relaxation and the RRF-induced relaxation seems to be resistant to oxidative stress. The mechanism responsible for the enhanced RRF-induced relaxation in the presence of flufenamic acid or carbenoxolone remains elusive and the RRF does not show more effectivity on retinal arteries.

Search for the Source of the Retinal Relaxing Factor.

Purpose/Aim of the study: the retinal relaxing factor (RRF) is an unidentified paracrine factor, which is continuously released from retinal tissue and causes smooth muscle cell relaxation. This study tried to identify the cellular source of the RRF. Furthermore, the possible RRF release by voltage-dependent sodium channel activation and the calcium-dependency of the RRF release were investigated. MATERIALS AND METHODS: mouse femoral arteries were mounted in myograph baths for in vitro isometric tension measurements. The vasorelaxing effect of chicken retinas, which contain no vascular cells, and of solutions incubated with MIO-M1 or primary Müller cell cultures were evaluated. The RRF release of other retinal cells was investigated by using cell type inhibitors. Concentration-response curves of veratridine, a voltage-dependent sodium channel activator, were constructed in the presence or absence of mouse retinal tissue to evaluate the RRF release. The calcium-dependency of the RRF release was investigated by evaluating the vasorelaxing effect of RRF-containing solutions made out of chicken retinas in the absence or presence of calcium. RESULTS: Chicken retinas induced vasorelaxation, whereas solutions incubated with Müller cell cultures did not. Moreover, the gliotoxin DL-α-aminoadipic acid, the microglia inhibitor minocycline, and the tetrodotoxin-resistant voltage-dependent sodium channel 1.8 inhibitor A-803467 could not reduce the RRF-induced relaxation. Concentration-response curves of veratridine were not enlarged in the presence of retinal tissue, and RRF-containing solutions made in the absence of calcium induced a substantial, but reduced vasorelaxation. CONCLUSIONS: the RRF is not released from vascular cells and probably neither from glial cells. The retinal cell type that does release the RRF remains unclear. Veratridine does not stimulate the RRF release in mice, and the RRF release in chickens is calcium-dependent as well as calcium-independent.

Ferulic acid-4-O-sulfate rather than ferulic acid relaxes arteries and lowers blood pressure in mice.

Consumption of foods rich in ferulic acid (FA) such as wholegrain cereals, or FA precursors such as chlorogenic acids in coffee, is inversely correlated with risk of cardiovascular disease and type 2 diabetes. As a result of digestion and phase II metabolism in the gut and liver, FA is converted predominantly into ferulic acid-4-O-sulfate (FA-sul), an abundant plasma metabolite. Although FA-sul is the main metabolite, very little has been reported regarding its bioactivities. We have compared the ex vivo vasorelaxing effect of FA and FA-sul (10-7-3.10-5M) on isolated mouse arteries mounted in tissue myographs. FA-sul, but not FA, elicited a concentration-dependent vasorelaxation of saphenous and femoral arteries and aortae. The FA-sul-mediated vasorelaxation was blunted by 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, a soluble guanylate cyclase (sGC) inhibitor. The role of sGC was confirmed in femoral arteries isolated from sGCα1(-/-) knockout mice. Furthermore, 4-aminopyridine, a specific inhibitor of voltage-dependent potassium channels, significantly decreased FA-sul-mediated effects. In anesthetized mice, intravenous injection of FA-sul decreased mean arterial pressure, whereas FA had no effect, confirming the results obtained ex vivo. FA-sul is probably one of the major metabolites accounting for the blood pressure-lowering effects associated with FA consumption.

The Retinal Relaxing Factor: Update on an Enigmatic Regulator of the Retinal Circulation.

The retinal circulation is regulated by different local factors and might include the retinal relaxing factor (RRF). This factor is found to be continuously released by the retina and relaxes smooth muscle cells. This review describes the current knowledge about the RRF. Despite many research efforts, the cellular source, identity, mechanism, and physiological role of the RRF remain largely unknown. Thus far, it seems that the RRF is a hydrophilic, thermostable, diffusible chemical messenger, which characteristics do not correspond with most well-known endogenous vasorelaxants. The RRF-induced relaxation seems to rely on activation of the inward rectifier K+ channels and the Rho kinase Ca2+ sensitization mechanism. Voltage-dependent K+ channels and plasma membrane Ca2+-ATPase might also be involved, whereas the involvement of cyclooxygenase is still a point of discussion. Furthermore, it appears that the RRF is involved in other relaxation pathways, namely those of hypoxia, adenosine, and adenosine triphosphate, hydrogen sulfide, γ-aminobutyric acid, and dorzolamide.

Inhibition of Cyclic GMP Export by Multidrug Resistance Protein 4: A New Strategy to Treat Erectile Dysfunction?

BACKGROUND: Intracellular cyclic guanosine monophosphate (cGMP) concentrations are regulated by degradation enzymes (phosphodiesterases) and by active transport across the plasma membrane by multidrug resistance proteins (MRPs) 4 and 5. AIM: To evaluate the functional effect of MRP-4 inhibition and the role of MRP-4-mediated cGMP export in mouse corpora cavernosa. METHODS: Isometric tension of mouse corpora cavernosa was measured after cumulative addition of MK-571, an inhibitor of MRP-4, or sildenafil, a phosphodiesterase type 5 inhibitor. In addition, the effect of MRP-4 inhibition on cGMP-independent and cGMP-dependent relaxations was studied. In vivo intracavernosal pressure and mean arterial pressure measurements were performed after intracavernosal injection of MK-571. The effect of MRP-4 inhibition on cGMP content was determined using an enzyme immunoassay kit. OUTCOMES: Measurement of the effect of MK-571 on cGMP content, relaxant responses of mouse corpora cavernosa to cGMP-independent and cGMP-dependent vasodilating substances, and determination of the ratio of intracavernosal pressure to mean arterial pressure after intracavernosal injection of MK-571. RESULTS: MK-571 and sildenafil relaxed the corpora cavernosa concentration dependently, with sildenafil being the more potent relaxing compound. Furthermore, MK-571 enhanced relaxing responses to cGMP-dependent substances, such as sodium nitroprusside, sildenafil, acetylcholine, and electrical field stimulation, with the latter even under in vitro diabetic conditions. In contrast, cGMP-independent relaxations were not altered by MRP-4 inhibition. Intracavernosal administration of MK-571 significantly increased intracavernosal pressure, with minimal effect on mean arterial pressure. The cGMP analysis showed that MRP-4 inhibition was accompanied by increased cGMP levels. CLINICAL TRANSLATION: MRP-4, at least when targeted locally in the penis or when combined with a phosphodiesterase type 5 inhibitor, might be a valuable alternative strategy for the treatment of (diabetic) erectile dysfunction. STRENGTHS AND LIMITATIONS: This study is the first to demonstrate an in vitro direct relaxant and an in vivo pro-erectile effect of the MRP-4 inhibitor, MK-571, on mouse corpora cavernosa. However, the functional effect of MRP-5-mediated export in mouse corpora cavernosa was not explored, which has been suggested to play the predominant role in cGMP export. CONCLUSION: Inhibition of MRP-4 increases basal and stimulated levels of cGMP, leading to corpora cavernosa relaxation and penile erection. Therefore, in addition to degradation of cGMP, export of cGMP by MRP-4 could contribute substantially to regulating cGMP levels in mouse corpora cavernosa. Boydens C, Pauwels B, Vanden Daele L, Van de Voorde J. Inhibition of Cyclic GMP Export by Multidrug Resistance Protein 4: A New Strategy to Treat Erectile Dysfunction? J Sex Med 2017;14:502-509.

Erectile Dysfunction in Heme-Deficient Nitric Oxide-Unresponsive Soluble Guanylate Cyclase Knock-In Mice.

INTRODUCTION: The nitric oxide (NO), soluble guanylate cyclase (sGC), and cyclic guanosine monophosphate (cGMP) pathway is the leading pathway in penile erection. AIM: To assess erectile function in a mouse model in which sGC is deficient in heme (apo-sGC) and unresponsive to NO. METHODS: Mutant mice (sGCß1ki/ki) that express an sGC enzyme that retains basal activity but fails to respond to NO because of heme deficiency (apo-sGC) were used. Isolated corpora cavernosa from sGCß1ki/ki and wild-type mice were mounted in vitro for isometric tension recordings in response to sGC-dependent and -independent vasorelaxant agents. In addition, the erectile effects of some of these agents were tested in vivo at intracavernosal injection. MAIN OUTCOME MEASURES: In vitro and in vivo recordings of erectile responses in sGCß1ki/ki and wild-type mice after stimulation with sGC-dependent and -independent vasorelaxant agents. RESULTS: NO-induced responses were abolished in sGCß1ki/ki mice in vitro and in vivo. The ability of the heme-dependent, NO-independent sGC stimulator BAY 41-2272 to relax the corpora cavernosa was markedly attenuated in sGCß1ki/ki mice. In contrast, the relaxation response to the heme- and NO-independent sGC activator BAY 58-2667 was significantly enhanced in sGCß1ki/ki mice. The relaxing effect of sGC-independent vasorelaxant agents was similar in wild-type and sGCß1ki/ki mice, illustrating that the observed alterations in vasorelaxation are limited to NO-sGC-cGMP-mediated processes. CONCLUSION: Our results suggest that sGC is the sole target of NO in erectile physiology. Furthermore, this study provides indirect evidence that, in addition to sGCα1ß1, sGCα2ß1 is important for erectile function. In addition, the significant relaxation observed in sGCß1ki/ki mice with the cumulative addition of the sGC activator BAY 58-2667 indicates that sGC activators might offer value in treating erectile dysfunction.

At the cross-point of connexins, calcium, and ATP: blocking hemichannels inhibits vasoconstriction of rat small mesenteric arteries.

AIMS: Connexins form gap-junctions (GJs) that directly connect cells, thereby coordinating vascular cell function and controlling vessel diameter and blood flow. GJs are composed of two hemichannels contributed by each of the connecting cells. Hemichannels also exist as non-junctional channels that, when open, lead to the entry/loss of ions and the escape of ATP. Here we investigated cross-talk between hemichannels and Ca2+/purinergic signalling in controlling blood vessel contraction. We hypothesized that hemichannel Ca2+ entry and ATP release contributes to smooth muscle cell (SMC) Ca2+ dynamics, thereby influencing vessel contractility. We applied several peptide modulators of hemichannel function and inhibitors of Ca2+ and ATP signalling to investigate their influence on SMC Ca2+ dynamics and vessel contractility. METHODS AND RESULTS: Confocal Ca2+ imaging studies on small mesenteric arteries (SMAs) from rat demonstrated that norepinephrine-induced SMC Ca2+ oscillations were inhibited by blocking IP3 receptors with xestospongin-C and by interfering with hemichannel function, most notably by the specific Cx43 hemichannel blocking peptide TAT-L2 and by TAT-CT9 that promotes Cx43 hemichannel opening. Evidence for hemichannel involvement in SMC function was supported by the fact that TAT-CT9 significantly increased SMC resting cytoplasmic Ca2+ concentration, indicating it facilitated Ca2+ entry, and by the observation that norepinephrine-triggered vessel ATP release was blocked by TAT-L2. Myograph tension measurements on isolated SMAs showed significant inhibition of norepinephrine-triggered contractility by the ATP receptor antagonist suramin, but the strongest effect was observed with TAT-L2 that gave ∼80% inhibition at 37 °C. TAT-L2 inhibition of vessel contraction was significantly reduced in conditional Cx43 knockout animals, indicating the effect was Cx43 hemichannel-dependent. Computational modelling suggested these results could be explained by the opening of a single hemichannel per SMC. CONCLUSIONS: These results indicate that Cx43 hemichannels contribute to SMC Ca2+ dynamics and contractility, by facilitating Ca2+ entry, ATP release, and purinergic signalling.

Vasorelaxing Effect of Resveratrol on Bovine Retinal Arteries.

PURPOSE: Resveratrol is a red wine polyphenol that causes vasorelaxation, which could be of interest in the treatment or prevention of eye diseases with an impaired blood flow. In this study, the vasorelaxant capacity of resveratrol (cis and trans) on bovine retinal arteries, its vasorelaxing mechanism, and its influence on the relaxation induced by the retinal relaxing factor (RRF) were examined. METHODS: Isolated bovine retinal arteries were mounted into wire myographs for isometric tension measurements. Concentration-response curves of cis- and trans-resveratrol and concentration-response curves of resveratrol in the absence or presence of the endothelium or different inhibitors were constructed. Relaxations elicited by the RRF with and without resveratrol incubation were also compared. RESULTS: Both resveratrol isomers caused a similar strong concentration-dependent relaxation. Removal of the endothelium or blocking endothelium-dependent pathways did not change the relaxation. Also, K+ channel blockers did not reduce the relaxation, except the 120 mM K+ Krebs Ringer bicarbonate solution. Phorbol 12-myristate 13-acetate and phorbol 12,13-dibutyrate blocked the relaxation partially and so did the inhibition of heme oxygenase-1. Blocking adenylyl cyclase, AMP-activated protein kinase, estrogen receptors, sirtuin 1, or sarco/endoplasmic reticulum Ca2+ ATPase did not have an effect. The relaxation caused by the RRF was not altered by resveratrol incubation. CONCLUSIONS: Cis- and trans-resveratrol relax bovine retinal arteries similarly and concentration dependently. The main relaxation mechanism remains unclear, but K+ channels, carbon monoxide, and the myosin phosphatase pathway may be involved. Resveratrol does not have an influence on the RRF.

Protective effect of resveratrol and quercetin on in vitro-induced diabetic mouse corpus cavernosum.

BACKGROUND: Hyperglycemia and increased levels of methylglyoxal (MGO) can trigger the development of vascular complications in diabetes. Resveratrol and quercetin are red wine polyphenols with known beneficial cardiovascular properties, including an antioxidant capacity. This study evaluated whether resveratrol and/or quercetin could prevent in vitro-induced diabetic changes in neurogenic and vascular relaxant responses of mouse arteries and corpora cavernosa. METHODS: Isometric tension of isolated aorta, mesenteric arteries and corpora cavernosa was measured using organ bath systems. Diabetic conditions were mimicked in vitro by co-incubating the tissues for 2 h with high glucose (HG, 30 mM) and MGO (120 µM). RESULTS: The presence of HG and MGO significantly blunted acetylcholine (Ach)-induced relaxations in corpora cavernosa and mesenteric arteries but not in aorta. Electrical field stimulated (EFS) responses of corpora cavernosa were also significantly inhibited by these diabetic conditions. In corpora cavernosa 2 h co-incubation with resveratrol (30 µM) or quercetin (30 µM) significantly attenuated HG and MGO-induced deficits in Ach- and EFS-responses. CONCLUSIONS: Our study demonstrates that in mouse arteries, HG and MGO rather affect endothelium derived hyperpolarizing factor-mediated than nitric oxide (NO)-mediated relaxations. In corpora cavernosa HG and MGO interfere with NO release. Resveratrol and quercetin protect mouse corpora cavernosa from diabetic-induced damage to NO-mediated relaxant responses. This might rely on their antioxidant capacity.

Ruthenium-based nitric oxide-donating and carbon monoxide-donating molecules.

OBJECTIVES: Over the past few years, the use of metallocomplexes for medical purposes has considerably grown. Because of its favourable characteristics, ruthenium has taken a significant place in this expanding field of research. Several ruthenium-containing metal compounds have been developed as delivery agents of physiological important molecules such as nitric oxide (NO) and carbon monoxide (CO). KEY FINDINGS: This review focuses on the (vaso)relaxant capacity of ruthenium-based NO-donating and CO-donating molecules in view of their potential usefulness in the treatment of cardiovascular diseases and erectile dysfunction. SUMMARY: Ruthenium seems to be a valuable candidate for the design of NO-donating and CO-donating molecules. To date, ruthenium remains of interest in drug research as the search for new alternatives is still necessary.

Effect of resveratrol and orchidectomy on the vasorelaxing influence of perivascular adipose tissue.

Perivascular adipose tissue (PVAT) releases several adipo(cyto)kines. Some are vasoactive substances that elicit a net beneficial anticontractile effect. Resveratrol and testosterone are known to modulate adipo(cyto)kine release from adipose tissue and could therefore influence the anticontractile effect of PVAT. In vitro tension measurements were performed using thoracic aorta segments with and without adipose tissue from sham-operated or orchidectomized male Swiss mice. Concentration-response curves to norepinephrine (NOR) were constructed in the presence and absence of resveratrol (10 µM, 15 min) or the relaxant effect of resveratrol (10-100 µM) was investigated after inducing tone with NOR (5 µM). Aortas with PVAT displayed significantly attenuated contractions to NOR compared with aortas without PVAT. In aortas without PVAT, resveratrol (10 µM) significantly decreased NOR responses and elicited concentration-dependent (10-100 µM) relaxations. However, in aortas with adherent PVAT, resveratrol (10 µM) neither decreased NOR responses, nor did resveratrol (10-100 µM) induce arterial relaxations. The anticontractile effect of PVAT was less pronounced in the presence of resveratrol and unaltered by orchidectomy. Orchidectomy did not influence contractions induced by NOR. Orchidectomy does not modulate the anticontractile capacity of PVAT, while resveratrol decreases the vasorelaxing influence of PVAT. The positive effects associated with resveratrol addition are neutralized by the presence of PVAT. This is thought to result from a dual effect of resveratrol: (1) inhibition of the influence of vasodilatory adipo(cyto)kines and (2) a direct relaxant effect on the vascular smooth muscle. Overall, the beneficial relaxing effect of resveratrol is lost in mice thoracic aorta surrounded by PVAT.

Cardiovascular and pharmacological implications of haem-deficient NO-unresponsive soluble guanylate cyclase knock-in mice.

Oxidative stress, a central mediator of cardiovascular disease, results in loss of the prosthetic haem group of soluble guanylate cyclase (sGC), preventing its activation by nitric oxide (NO). Here we introduce Apo-sGC mice expressing haem-free sGC. Apo-sGC mice are viable and develop hypertension. The haemodynamic effects of NO are abolished, but those of the sGC activator cinaciguat are enhanced in apo-sGC mice, suggesting that the effects of NO on smooth muscle relaxation, blood pressure regulation and inhibition of platelet aggregation require sGC activation by NO. Tumour necrosis factor (TNF)-induced hypotension and mortality are preserved in apo-sGC mice, indicating that pathways other than sGC signalling mediate the cardiovascular collapse in shock. Apo-sGC mice allow for differentiation between sGC-dependent and -independent NO effects and between haem-dependent and -independent sGC effects. Apo-sGC mice represent a unique experimental platform to study the in vivo consequences of sGC oxidation and the therapeutic potential of sGC activators.

The influence of ruthenium on vascular tone.

OBJECTIVES: Over the past few years, ruthenium has been under attention for development of organometallic drugs with various therapeutic applications. Because of its favourable characteristics, ruthenium is perfectly suitable for drug design. Ruthenium-containing complexes exert a wide range of biological effects. However, so far, the influence of ruthenium itself on vascular tone has never been studied. METHODS: The effect of ruthenium was analysed through organ bath studies measuring isometric tension on mice thoracic aorta. After obtaining a stable contraction plateau, cumulative concentration-response curves of the ruthenium-compounds (RuCl3 , Ruthenium Red, [RuCl2 (CO)3 ]2 and RuCl2 (DMSO)4 ) (30-600 µmol/l) were performed. The effect of RuCl3 after contraction with different contractile agents was evaluated. Furthermore, the influence of ruthenium-containing molecules on endogenous (acetylcholine) and exogenous (sodium nitroprusside) NO-mediated relaxations was determined. KEY FINDINGS: All studied ruthenium compounds elicit, to some extent, a decrease of the contraction level. Looking further into the underlying mechanism, we found that RuCl3 relaxes aortic rings only when contracted with norepinephrine. This RuCl3 -induced relaxation can be prevented by the antioxidants ascorbic acid and N-acetyl L-cysteine. In addition, ruthenium compounds may diminish acetylcholine- or sodium nitroprusside-induced relaxations. CONCLUSIONS: Ruthenium-containing molecules can influence vascular tone induced by norepinephrine due to oxidative inactivation. Moreover, they can undermine NO-mediated responses. This should be considered when developing ruthenium-containing drugs.

Oximes induce erection and are resistant to oxidative stress.

INTRODUCTION: Because of their nitric oxide (NO)-donating capacities, oxime derivatives have shown to offer some therapeutic perspective for the treatment of erectile dysfunction (ED) as well as cardiovascular diseases. However, to date the in vivo effect of these oximes on erectile function remains unknown. In many disease states oxidative stress occurs, impairing NO-mediated relaxations. Hence the influence of oxidative stress on oxime-induced effects is also of interest. AIMS: This study aimed to evaluate the in vivo effect of formaldoxime (FAL) and formamidoxime (FAM) on blood pressure and intracavernosal pressure (ICP); and to examine the role of soluble guanylyl cyclase (sGC) and the influence of oxidative stress on the FAL and FAM responses. METHODS: Blood pressure and ICP were monitored in vivo after resp. intravenous or intracavernosal injection of FAL and FAM. Moreover isometric tension was measured in vitro on isolated mice corpora cavernosa (CC), thoracic aorta, and femoral artery in organ baths. The role of sGC was investigated using transgenic mice lacking the alpha 1 subunit of sGC. MAIN OUTCOME MEASURES: Mean arterial pressure (MAP) and ICP were measured after FAL/FAM injection. In vitro relaxation of CC strips was evaluated in response to addition of FAL/FAM. RESULTS: In vivo both FAL and FAM elicit a dose-dependent lowering of blood pressure (maximal ΔMAP: 33.66 ± 4.07 mm Hg [FAL] and 20.43 ± 2.06 mm Hg [FAM] ) as well as an increase of ICP (maximal increase of ICP/MAP: 70.29 ± 2.88% [FAL] and 52.91 ± 8.61% [FAM] ). The FAL/FAM effect is significantly lower in knockout vs. wild-type mice. Oxidative stress has an inhibitory effect on corporal NO-mediated relaxations induced by electrical field stimulation, acetylcholine, and sodium nitroprusside whereas the responses to 8-(4-chlorophenylthio)-guanosine 3',5'-cyclic monophosphate sodium salt, FAL and FAM were not influenced. CONCLUSIONS: Oximes induce erection which is mediated by sGC. The oxime-induced relaxations are resistant to oxidative stress, which increases their therapeutic potential for the treatment of ED.

Relaxant and antioxidant capacity of the red wine polyphenols, resveratrol and quercetin, on isolated mice corpora cavernosa.

INTRODUCTION: The red wine polyphenols resveratrol and quercetin are known for their vasorelaxant and antioxidant capacity, which is assumed to rely on the activation of the nitric oxide (NO)/soluble guanylyl cyclase (sGC) pathway. Vasodilators as well as antioxidants can regulate penile erection and be beneficial for the treatment of erectile dysfunction (ED). AIMS: The goal of this study was to evaluate the NO/sGC dependency of the relaxant effect of resveratrol and quercetin on mice aorta and corpora cavernosa (CC), as well as to explore their influence on oxidative stress-induced ED. METHODS: Isolated mice aorta and CC were mounted for isometric tension recordings into organ baths. Cumulative concentration-response curves were constructed for resveratrol and quercetin in the absence/presence of inhibitors of the NO/sGC pathway. In addition, in CC the effect of resveratrol and quercetin was studied on NO-mediated relaxations using acetylcholine (Ach), sodium nitroprusside (SNP), and electrical field stimulation (EFS). In certain experiments, corporal tissues were exposed to oxidative stress using palmitic acid (PA, 0.5 mM). MAIN OUTCOME MEASURES: Corporal responses to resveratrol and quercetin were measured in the presence/absence of inhibitors of different molecular pathways. The effect of resveratrol and quercetin incubation on Ach-, SNP-, or EFS-mediated responses was explored in the presence/absence of PA. RESULTS: While both polyphenols are potent vasodilators of mice aorta, only resveratrol relaxes mice CC. The relaxation response to resveratrol on aorta was diminished in sGCα1 (-/-) mice, but not on CC. The polyphenols did not influence Ach-, SNP-, or EFS-mediated relaxations as such. Resveratrol, but not quercetin, was able to significantly reverse PA-induced decrease of EFS relaxations. CONCLUSION: The red wine compound resveratrol, but not quercetin, relaxes isolated mice CC concentration-dependently through mechanisms independent of the NO/sGC pathway. Resveratrol is a more potent antioxidant than quercetin, being able to restore decreased neuronal NO responses in mice CC.

Perivascular adipose tissue, inflammation and vascular dysfunction in obesity.

Adipocytes are no longer considered just as cells related to storage of energy and thermoregulation. Now we know that they release a huge number of paracrine and endocrine biologically active molecules. This is also the case for perivascular adipose tissue (PVAT) that surrounds almost all blood vessels in the organism. PVAT secretes the so-called adipo(cyto)kines that, because of its proximity, can easily influence vascular smooth muscle cells. The role of PVAT on vascular function can be both protective and deleterious. Normal healthy PVAT, as present in lean subjects, helps to keep the blood vessels dilated as its presence diminishes the effect of vasocontractile agents. Obesity is associated with an increased mass in PVAT. Excessive adipocyte hypertrophy may result in "adiposopathy" in which PVAT attracts macrophages and becomes a more inflammatory phenotype. This leads to a change in profile of the released adipo(cyto)kines, resulting in a decreased vasorelaxing effect of PVAT, which may be linked to obesity-induced hypertension. It also results in smooth muscle cell migration and proliferation and the development of atherosclerotic lesions. The increased knowledge of PVAT function brings up new targets that can be useful to develop novel therapeutic and preventive strategies for obesity-related cardiovascular diseases. This mini-review presents a general overview of the actual knowledge on the role of PVAT on vascular function and dysfunction in obesity.

NO-donating oximes relax corpora cavernosa through mechanisms other than those involved in arterial relaxation.

INTRODUCTION: Erectile dysfunction (ED), as well as many cardiovascular diseases, is associated with impaired nitric oxide (NO) bioavailability. Recently, oxime derivatives have emerged as vasodilators due to their NO-donating capacities. However, whether these oximes offer therapeutic perspectives as an alternative NO delivery strategy for the treatment of ED is unexplored. AIMS: This study aims to analyze the influence of formaldoxime (FAL), formamidoxime (FAM), and cinnamaldoxime (CAOx) on corporal tension and to elucidate the underlying molecular mechanisms. METHODS: Organ bath studies were carried out measuring isometric tension on isolated mice corpora cavernosa (CC), thoracic aorta, and femoral artery. After contraction with norepinephrine (NOR), cumulative concentration-response curves of FAL, FAM, and CAOx (100 nmol/L-1 mmol/L) were performed. MAIN OUTCOME MEASURES: FAL-/FAM-induced relaxations were evaluated in the absence/presence of various inhibitors of different molecular pathways. RESULTS: FAL, FAM, and CAOx relax isolated CC as well as aorta and femoral artery from mice. ODQ (soluble guanylyl cyclase-inhibitor), diphenyliodonium chloride (nonselective flavoprotein inhibitor), and 7-ethoxyresorufin (inhibitor of CYP450 1A1 and NADPH-dependent reductases) substantially blocked the FAL-/FAM-induced relaxation in the arteries but not in CC. Only a small inhibition of the FAM response in CC was observed with ODQ. CONCLUSIONS: This study shows for the first time that NO-donating oximes relax mice CC. Therefore, oximes are a new group of molecules with potential for the treatment of ED. However, the underlying mechanism(s) of the FAL-/FAM-induced corporal relaxation clearly differ(s) from the one(s) involved in arterial vasorelaxation.

Protein-bound uremic toxins stimulate crosstalk between leukocytes and vessel wall.

Leukocyte activation and endothelial damage both contribute to cardiovascular disease, a major cause of morbidity and mortality in CKD. Experimental in vitro data link several protein-bound uremic retention solutes to the modulation of inflammatory stimuli, including endothelium and leukocyte responses and cardiovascular damage, corroborating observational in vivo data. However, the impact of these uremic toxins on the crosstalk between endothelium and leukocytes has not been assessed. This study evaluated the effects of acute and continuous exposure to uremic levels of indoxylsulfate (IS), p-cresylsulfate (pCS), and p-cresylglucuronide (pCG) on the recruitment of circulating leukocytes in the rat peritoneal vascular bed using intravital microscopy. Superfusion with IS induced strong leukocyte adhesion, enhanced extravasation, and interrupted blood flow, whereas pCS caused a rapid increase in leukocyte rolling. Superfusion with pCS and pCG combined caused impaired blood flow and vascular leakage but did not further enhance leukocyte rolling over pCS alone. Intravenous infusion with IS confirmed the superfusion results and caused shedding of heparan sulfate, pointing to disruption of the glycocalyx as the mechanism likely mediating IS-induced flow stagnation. These results provide the first clear in vivo evidence that IS, pCS, and pCG exert proinflammatory effects that contribute to vascular damage by stimulating crosstalk between leukocytes and vessels.

Adipocytokines in relation to cardiovascular disease.

Adipose tissue can be considered as a huge gland producing paracrine and endocrine hormones, the adipo(cyto)kines. There is growing evidence that these adipo(cyto)kines may link obesity to cardiovascular diseases. The excessive adipocyte hypertrophy in obesity induces hypoxia in adipose tissue. This leads to adiposopathy, the process that converts "healthy" adipose tissue to "sick" adipose tissue. This is accompanied by a change in profile of adipo(cyto)kines released, with less production of the "healthy" adipo(cyto)kines such as adiponectin and omentin and more release of the "unhealthy" adipo(cyto)kines, ultimately leading to the development of cardiovascular diseases. The present review provides a concise and general overview of the actual concepts of the role of adipo(cyto)kines in endothelial dysfunction, hypertension, atherosclerosis and heart diseases. The knowledge of these concepts may lead to new tools to improve health in the next generations.