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AIMS: To evaluate the effects of testosterone on endothelium-dependent vasodilation and oxidative stress in mesenteric resistance arteries. MAIN METHODS: Spontaneously hypertensive rats (SHR), aged 8 to 10 weeks, were divided into four groups: intact (SHAM), intact treated with testosterone (TTO; 3 mg/kg/day) via subcutaneous route (s.c.), intact treated with testosterone and anastrozole [aromatase enzyme inhibitor (TTO + ANA; 0.1 mg/kg/day, s.c.)] and intact treated with testosterone and finasteride [5 α-reductase enzyme inhibitor (TTO + FIN; 5 mg/kg/day, s.c.)] for four weeks. Concentration-response curves to acetylcholine (ACh, 0.1 nmol/L - 10 µmol/L) were obtained in mesenteric resistance arteries previously contracted with phenylephrine (PE, 3 µmol/L), before and after the use of selective inhibitors. Reactive oxygen species (ROS) levels were assessed in the vessels and the endothelium analyzed by scanning electron microscopy. KEY FINDINGS: TTO group showed a lower participation of nitric oxide (NO), increased oxidative stress, and participation of prostanoids and endothelium-dependent hyperpolarization (EDH), possibly to maintain the vasodilator response. Lower participation of NO and prostanoids, combined to an increased participation of EDH, were observed in the TTO + ANA group, in addition to higher levels of ROS and altered endothelial morphology. The vasodilation to ACh was impaired in TTO + FIN, along increased participation of NO, reduction of prostanoids, and greater EDH-dependent vasodilation. SIGNIFICANCE: Testosterone contributes to endothelial vasodilation by enhancing EDH through an increased participation of epoxyeicosatrienoic acids. While the decrease in NO appears to involve the participation of dihydrotestosterone, 17 ß-estradiol seems to stimulate the action of the NO pathway and prostanoids.
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Hipertensão , Vasodilatação , Ratos , Animais , Espécies Reativas de Oxigênio/metabolismo , Testosterona/farmacologia , Testosterona/metabolismo , Hipertensão/metabolismo , Ratos Endogâmicos SHR , Inibidores Enzimáticos/farmacologia , Acetilcolina/farmacologia , Acetilcolina/metabolismo , Artérias Mesentéricas , Óxido Nítrico/metabolismo , Prostaglandinas/metabolismo , Endotélio Vascular/metabolismoRESUMO
During hypertension an unbalance of short-chain fatty acids (SCFAs) production by intestinal bacteria is described. However, no data evaluate the association of SCFAs and vascular remodeling in hypertension, which is an important hallmark of this disease. Thus, the present study aims to evaluate the correlations between SCFAs availability and the resistance arteries remodeling in hypertension, as well as to identify the possible pathway by which the SCFAs could exert a structural and mechanical influence. Hence, male spontaneously hypertensive rats (SHR) and normotensive Wistar rats had blood pressure measured by tail-cuff plethysmography; fecal SCFAs content assessed by gas chromatography; gene expression of SCFAs-transporters in gut epithelium and SCFAs-sensing receptors on mesenteric resistance arteries (MRA) quantified by PCR; and MRA structural and mechanical parameters analyzed by pressure myograph. Reduced butyrate fecal content was found in SHR, with no changes in propionate and acetate, as well as decreased mRNA levels of SCFAs-transporters (MCT1, MCT4, and SMCT1) in the intestinal epithelium. In addition, lower gene expression of SCFAs-sensing receptors (GPR41, GPR43, and GPR109a, but not Olfr78) was identified in MRAs of SHR, which also shows inward eutrophic remodeling with stiffness. Butyrate content presented a negative correlation with systolic blood pressure and with the structural alterations found on MRAs, while a positive correlation between butyrate content and mechanical parameters was detected. Altogether the present study suggests that lower butyrate content due to ineffective SCFA bioavailability, associated with lower SCFAs-sensing receptors expression, could favor MRA remodeling, increasing peripheral vascular resistance and worsening hypertension prognosis.
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AIM: Endothelial mechanisms underlying the vascular effects of estrogen modulated by the G protein-coupled estrogen receptor (GPER) are not well understood, especially in gonadal sex hormone deprivation. Thus, we investigated vascular function and endothelial signaling pathways involved in the selective activation of GPER in resistance arteries of gonadectomized rats. METHODS: Gonadectomy was performed in Wistar rats of both sexes. After 21 days, the animals were euthanized. Concentration-response curves were obtained by cumulative additions of G-1 in third-order mesenteric arteries. The vasodilatory effects of G-1 were evaluated before and after endothelium removal or incubation with pharmacological inhibitors. Tissue protein expression was measured by western blotting. Assays with 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM) and 2',7' dichlorodihydrofluorescein-diacetate (H2DCF-DA) were performed in the arteries investigated. Immunolocalization was assessed by immunofluorescence. RESULTS: G-1 induced partially endothelium-dependent relaxation in both sexes. The three isoforms of the enzyme nitric oxide synthase contributed to the production and release of nitric oxide in both gonadectomized groups, but the role of inducible nitric oxide synthase is more expressive in males. The mechanistic pathway by which endothelial nitric oxide synthase is phosphorylated appears to differ between sexes, with the rapid signaling pathway phosphatidylinositol-3-kinase/protein kinase B/endothelial nitric oxide synthase (PI3k-Akt-eNOS) being identified for males and mitogen-activated protein kinase/extracellular signal-regulated kinase/endothelial nitric oxide synthase (MEK-ERK-eNOS) for females. The contribution of hydrogen peroxide as an endothelial relaxation mediator seems to be greater in females. CONCLUSION: These results provide new insights into the effects of estrogen-induced responses via GPER on vascular function in gonadal sex hormone deprivation.
Assuntos
Óxido Nítrico Sintase Tipo III , Proteínas Proto-Oncogênicas c-akt , Animais , Endotélio Vascular , Estrogênios/metabolismo , Estrogênios/farmacologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Proteínas de Ligação ao GTP/metabolismo , Hormônios Esteroides Gonadais/metabolismo , Peróxido de Hidrogênio/metabolismo , Masculino , Artérias Mesentéricas , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/farmacologia , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositóis/metabolismo , Fosfatidilinositóis/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar , Receptores de Estrogênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Caracteres Sexuais , Transdução de Sinais , Vasodilatadores/farmacologiaRESUMO
Increased adiposity in perivascular adipose tissue (PVAT) has been related to vascular dysfunction. High-fat (HF) diet-induced obesity models are often used to analyze the translational impact of obesity, but differences in sex and Western diet type complicate comparisons between studies. The role of PVAT was investigated in small mesenteric arteries (SMAs) of male and female mice fed a HF or a HF plus high-sucrose (HF + HS) diet for 3 or 5 months and compared them to age/sex-matched mice fed a chow diet. Vascular responses of SMAs without (PVAT-) or with PVAT (PVAT+) were evaluated. HF and HF + HS diets increased body weight, adiposity, and fasting glucose and insulin levels without affecting blood pressure and circulating adiponectin levels in both sexes. HF or HF + HS diet impaired PVAT anticontractile effects in SMAs from females but not males. PVAT-mediated endothelial dysfunction in SMAs from female mice after 3 months of a HF + HS diet, whereas in males, this effect was observed only after 5 months of HF + HS diet. However, PVAT did not impact acetylcholine-induced relaxation in SMAs from both sexes fed HF diet. The findings suggest that the addition of sucrose to a HF diet accelerates PVAT dysfunction in both sexes. PVAT dysfunction in response to both diets was observed early in females compared to age-matched males suggesting a susceptibility of the female sex to PVAT-mediated vascular complications in the setting of obesity. The data illustrate the importance of the duration and composition of obesogenic diets for investigating sex-specific treatments and pharmacological targets for obesity-induced vascular complications.
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BACKGROUND: The protective effect of estrogen on the vasculature cannot be explained only by its action through the receptors ERα and ERß. G protein-coupled estrogen receptors (GPER)-which are widely distributed throughout the cardiovascular system-may also be involved in this response. However, little is known about GPER actions in hypertension. Therefore, in this study we evaluated the vascular response mediated by GPER using a specific agonist, G-1, in spontaneously hypertensive rats (SHR). We hypothesized that G-1 would induce a relaxing response in resistance mesenteric arteries from SHR of both sexes. METHODS: G-1 concentration-response curves (1 nM-10 µM) were performed in mesenteric arteries from SHR of both sexes (10-12-weeks-old, weighing 180-250 g). The effects of G-1 were evaluated before and after endothelial removal and incubation for 30 min with the inhibitors L-NAME (300 µM) and indomethacin (10 µM) alone or combined with clotrimazole (0.75 µM) or catalase (1,000 units/mL). GPER immunolocalization was also investigated, and vascular hydrogen peroxide (H2O2) and ROS were evaluated using dichlorofluorescein (DCF) and dihydroethidium (DHE) staining, respectively. RESULTS: GPER activation promoted a similar relaxing response in resistance mesenteric arteries of female and male hypertensive rats, but with the participation of different endothelial mediators. Males appear to be more dependent on the NO pathway, followed by the H2O2 pathway, and females on the endothelium and H2O2 pathway. CONCLUSION: These findings show that the GPER agonist G-1 can induce a relaxing response in mesenteric arteries from hypertensive rats of both sexes in a similar way, albeit with differential participation of endothelial mediators. These results contribute to the understanding of GPER activation on resistance mesenteric arteries in essential hypertension.
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AIM: Previous studies raise cyclooxygenase (COX) activation as a possible mechanism involved in the pathophysiology of ouabain-induced hypertension. We hypothesized that inhibition of COX-2 activity might prevent ouabain-induced vascular dysfunction and worsening of hypertension in spontaneously hypertensive rats (SHR). METHODS: SHR were exposed to ouabain or vehicle and treated or not with the selective COX-2 inhibitor nimesulide for 5 weeks. Systolic blood pressure was measured by plethysmography. Vascular reactivity by wire myograph and protein expression by Western-blot were assessed in mesenteric resistance arteries (MRA) of groups. Thromboxane A2 (TXA2) production by ELISA was evaluated in MRA supernatants of groups. RESULTS: Noradrenaline-induced maximal contraction (Emax) was greater in MRA from SHR receiving ouabain than those of vehicle group. In situ inhibition of COX-2, TXA2 synthase, or TP receptor reduced the Emax to noradrenaline in MRA of ouabain to vehicle levels. TXA2 production was higher in ouabain than in vehicle group. Ouabain enhanced expression of cytoplasmic tyrosine kinase Src (c-Src)/ERK1/2/COX-2/TXA2 synthase/TP receptor in SHR MRA, but did not change NFkB/iKB ratio. Anticontractile effect of nitric oxide (NO) was smaller in MRA from ouabain- than vehicle-treated SHR, as well as eNOS and nNOS expression. Nimesulide co-treatment prevented the ouabain-induced worsening of hypertension and noradrenaline MRA hypercontractility in SHR. CONCLUSION: Ouabain worsen hypertension and induce MRA hypercontractility in SHR associated with upregulated c-Src/ERK1/2/COX-2/TXA2 synthase/TXA2/TP receptor axis. These effects were prevented by COX-2 inhibition.
Assuntos
Hipertensão , Ouabaína , Animais , Pressão Sanguínea , Ciclo-Oxigenase 2/metabolismo , Endotélio Vascular , Hipertensão/induzido quimicamente , Hipertensão/tratamento farmacológico , Hipertensão/prevenção & controle , Artérias Mesentéricas/metabolismo , Ouabaína/farmacologia , Ratos , Ratos Endogâmicos SHR , Vasoconstrição , VasodilataçãoRESUMO
Aim: We investigated the antioxidant protective power of egg white hydrolysate (EWH) against the vascular damage induced by mercury chloride (HgCl2) exposure in resistance arteries. Methods: Male Wistar rats received for 60 days: (I) intramuscular injections (i.m.) of saline and tap water by gavage - Untreated group; (II) 4.6 µg/kg of HgCl2 i.m. for the first dose and subsequent doses of 0.07 µg/kg/day and tap water by gavage - HgCl2 group; (III) saline i.m. and 1 g/kg/day of EWH by gavage - EWH group, or (IV) the combination of the HgCl2 i.m. and EWH by gavage - EWH + HgCl2 group. Blood pressure (BP) was indirectly measured and dose-response curves to acetylcholine (ACh), sodium nitroprusside (SNP), and noradrenaline (NE) were assessed in mesenteric resistance arteries (MRA), as in situ production of superoxide anion, nitric oxide (NO) release, vascular reactive oxygen species (ROS), lipid peroxidation, and antioxidant status. Results: Egg white hydrolysate prevented the elevation in BP and the vascular dysfunction after HgCl2 exposure; restored the NO-mediated endothelial modulation and inhibited the oxidative stress and inflammatory pathways induced by HgCl2. Conclusion: Egg white hydrolysate seems to be a useful functional food to prevent HgCl2-induced vascular toxic effects in MRA.
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Fetal undernutrition programs hypertension and cardiovascular diseases, and resistance artery remodeling may be a contributing factor. We aimed to assess if fetal undernutrition induces resistance artery remodeling and the relationship with hypertension. Sprague-Dawley dams were fed ad libitum (Control) or with 50% of control intake between days 11 and 21 of gestation (maternal undernutrition, MUN). In six-month-old male and female offspring we assessed blood pressure (anesthetized and tail-cuff); mesenteric resistance artery (MRA) structure and mechanics (pressure myography), cellular and internal elastic lamina (IEL) organization (confocal microscopy) and plasma MMP-2 and MMP-9 activity (zymography). Systolic blood pressure (SBP, tail-cuff) and plasma MMP activity were assessed in 18-month-old rats. At the age of six months MUN males exhibited significantly higher blood pressure (anesthetized or tail-cuff) and plasma MMP-9 activity, while MUN females did not exhibit significant differences, compared to sex-matched controls. MRA from 6-month-old MUN males and females showed a smaller diameter, reduced adventitial, smooth muscle cell density and IEL fenestra area, and a leftward shift of stress-strain curves. At the age of eighteen months SBP and MMP-9 activity were higher in both MUN males and females, compared to sex-matched controls. These data suggest that fetal undernutrition induces MRA inward eutrophic remodeling and stiffness in both sexes, independent of blood pressure level. Resistance artery structural and mechanical alterations can participate in the development of hypertension in aged females and may contribute to adverse cardiovascular events associated with low birth weight in both sexes.
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Mechanisms underlying the vasorelaxant effects of the synthetic nitro compound, trans-4-methoxy-ß-nitrostyrene (T4MN) were studied in isolated small resistance arteries from spontaneously hypertensive rats (SHRs). T4MN caused vasorelaxation in endothelium-intact third-order branches of the mesenteric artery pre-contracted with noradrenaline (NA). This effect was unchanged by indomethacin and atropine but was significantly reduced by endothelium removal, L-NAME, LY294002, glybenclamide, TEA, apamin, TRAM 34, or by the association of apamin and TRAM 34. Pretreatment with the sGC inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) reduced the T4MN-induced relaxation in endothelium-intact, but not in denuded preparations. Incubation of small resistance arteries with T4MN increased nitric oxide (NO) production, an effect that was blocked by L-NAME. In Ca2+-free medium, T4MN inhibits the contractions induced by (i) NA, (ii) exogenous calcium through receptor- or voltage-operated Ca2+ channels and (iii) those evoked by Ca2+ influx through stores-operated Ca2+ channels activated by thapsigargin-induced Ca2+ store depletion. In contrast, T4MN was inert against the transient contraction induced by caffeine in Ca2+-free medium. In conclusion, T4MN induced effective vasorelaxant effects in isolated small resistance arteries from SHRs. This vasorelaxation seems to be mediated partly by an endothelium-dependent mechanism involving activation of Akt/eNOS/NO pathway and partly by an endothelium-independent mechanism through activation of sGC/cGMP/PKG pathway in vascular smooth muscle, leading to inhibition of Ca2+ influx from the extracellular milieu and IP3-sensitive intracellular Ca2+ release as well as activation of potassium channels.
Assuntos
Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiologia , Estirenos/farmacologia , Vasodilatação/efeitos dos fármacos , Animais , Pressão Sanguínea/efeitos dos fármacos , Canais de Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Receptores Muscarínicos/metabolismo , Receptores de Prostaglandina/metabolismo , Vasoconstrição/efeitos dos fármacosRESUMO
We aimed to investigate the effect of linoleic acid (LA) treatment on the blood pressure and function of mesenteric resistance arteries (MRA) in spontaneous hypertensive rats (SHR). Male SHR were treated daily with LA (15 mg/kg) or vehicle (control) for 15 days. Compared with controls, LA treatment decreased blood pressure and showed the following in MRA: (1) increased lumen and external diameter, (2) decreased wall:lumen ratio and wall thickness, (3) decreased stiffness and (4) less collagen deposition. LA treatment reduced the contractile response to phenylephrine, although there were no changes observed in MRA in regard to the acetylcholine or sodium nitroprusside responses. Incubation with L-NAME left-shifted the reactivity to phenylephrine only in the MRA treated group, suggesting that LA treatment can improve NO bioavailability. This result was accompanied by an increase "in situ" NO production. Incubation with tiron decreased vascular reactivity to phenylephrine in MRA in LA rats, which was accompanied by decreased superoxide anion production. Moreover, incubation with indomethacin (non-selective COX inhibitor, 10 µM), NS 398 (COX-2 specific inhibitor, 1 µM), furegrelate (TXA2 synthase inhibitor, 1 µM), SQ 29.548 (TP receptor antagonist, 1 µM) and SC 19220 (EP1 receptor antagonist, 10 µM) reduced the vasoconstrictor responses to phenylephrine in MRA in the treated group. These results were accompanied by a reduction in COX-2 protein expression. In conclusion, these findings show that LA treatment decreases blood pressure. In addition, the improvement of endothelial dysfunction and structural changes in this hypertension model may be responsible for the reduction in blood pressure.
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Hipertensão/fisiopatologia , Ácido Linoleico/farmacologia , Artérias Mesentéricas/efeitos dos fármacos , Animais , Anti-Hipertensivos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Hipertensão/tratamento farmacológico , Masculino , Artérias Mesentéricas/fisiopatologia , NG-Nitroarginina Metil Éster/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Fenilefrina/farmacologia , Prostaglandinas/metabolismo , Ratos Endogâmicos SHR , Remodelação Vascular/efeitos dos fármacos , Vasoconstrição/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
AIM: An imbalance between antioxidant and pro-oxidant factors, with a predominance of the latter, characterises oxidative stress and is indicative of a loss of vascular function. The beneficial vascular effects of oestrogen may be related to its ability to stimulate the G protein-coupled oestrogen receptor (GPER) and produce antioxidant activity. This study evaluated the GPER-dependent relaxation response in the mesenteric resistance arteries of female and male rats and measured the contributions of pro-oxidant and antioxidant enzymes in this response. MAIN METHODS: The relaxation response was characterised in third-order mesenteric arteries using concentration-response curves of the selective GPER agonist G-1 (1â¯nM-10⯵M), target protein levels were measured using Western blots, and vascular superoxide anion (O2-) and hydrogen peroxide (H2O2) levels were measured using dihydroethidium (DHE) and dichlorofluorescein (DCF) staining, respectively. KEY FINDINGS: The GPER agonist induced concentration-dependent vasorelaxation without showing differences between sexes. However, GPER expression was greater in male rats. No sex differences were detected in the expression of antioxidant proteins (catalase, SOD-1, and SOD-2). The basal vascular production of O2- and H2O2 was similar in the studied groups, and stimulation with G-1 maintained this response. SIGNIFICANCE: Together, our results show that the expression of GPER is greater in male mesenteric arteries, despite of the lack of a difference in vascular response. Nevertheless, antioxidant enzyme expression levels and the generation rates of pro-oxidants were similar between the studied groups. These results offer a new perspective for understanding GPER expression and functionality in resistance arteries.
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Antioxidantes/metabolismo , Endotélio Vascular/metabolismo , Artérias Mesentéricas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Vasodilatação/fisiologia , Animais , Endotélio Vascular/citologia , Feminino , Masculino , Artérias Mesentéricas/citologia , Ratos , Ratos Wistar , Fatores Sexuais , Transdução de SinaisRESUMO
AIM: The action of oestrogen has traditionally been attributed to the activation of nuclear receptors (ERα and ERß). A third receptor, the G protein-coupled oestrogen receptor (GPER), has been described as mediator of the rapid action of oestrogen. Based on the possible protective role of oestrogen in the cardiovascular system, the present study was designed to determine whether selective GPER activation induces relaxation of mesenteric resistance arteries in both sexes and which signalling pathways are involved. MAIN METHODS: Third-order mesenteric arteries were isolated, and concentration-response curves were plotted following the cumulative addition of the selective GPER agonist G-1 (1nM-10µM) following induction of contraction with phenylephrine (3µM). The vasodilatory effects of G-1 were assessed before and after removal of the endothelium or incubation for 30min with nitric oxide synthase (Nω-nitro-L-arginine methyl ester - L-NAME, 300µM) and cyclooxygenase (indomethacin - INDO, 10µM) inhibitors alone or combined, PI3K-Akt pathway inhibitor (LY-294,002, 2.5µM) or a potassium channel blocker (tetraethylammonium - TEA, 5mM). GPER immunolocalisation was also performed on the investigated arteries. KEY FINDINGS: The tested GPER agonist induced concentration-dependent relaxation of the mesenteric resistance arteries without differences related to sex that were partially endothelium dependent, mainly mediated by the PI3K-Akt-eNOS pathway and attenuated by nonspecific potassium channel blockade. In addition, the endothelial GPER immunolocalisation was stronger among females. SIGNIFICANCE: This evidence provides a new perspective for understanding the mechanisms involved in the vascular responses triggered by oestrogen via GPER in both sexes.
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Ciclopentanos/farmacologia , Estrogênios/metabolismo , Artérias Mesentéricas/efeitos dos fármacos , Canais de Potássio/efeitos dos fármacos , Quinolinas/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Animais , Ciclopentanos/administração & dosagem , Relação Dose-Resposta a Droga , Feminino , Masculino , Artérias Mesentéricas/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Quinolinas/administração & dosagem , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
Endothelial dysfunction is a common problem associated with hypertension and is considered a precursor to the development of micro- and macro-vascular complications. The present study investigated the involvement of nNOS (neuronal nitric oxide synthase) and H2O2 (hydrogen peroxide) in the impaired endothelium-dependent vasodilation of the mesenteric arteries of DOCA (deoxycorticosterone acetate)-salt-hypertensive mice. Myograph studies were used to investigate the endothelium-dependent vasodilator effect of ACh (acetylcholine). The expression and phosphorylation of nNOS and eNOS (endothelial nitric oxide synthase) were studied by Western blot analysis. Immunofluorescence was used to examine the localization of nNOS and eNOS in the endothelial layer of the mesenteric artery. The vasodilator effect of ACh is strongly impaired in mesenteric arteries of DOCA-salt-hypertensive mice. Non-selective inhibition of NOS sharply reduced the effect of ACh in both DOCA-salt-hypertensive and sham mice. Selective inhibition of nNOS and catalase led to a higher reduction in the effect of ACh in sham than in DOCA-salt-hypertensive mice. Production of H2O2 induced by ACh was significantly reduced in vessels from DOCA-salt-hypertensive mice, and it was blunted after nNOS inhibition. The expression of both eNOS and nNOS was considerably lower in DOCA-salt-hypertensive mice, whereas phosphorylation of their inhibitory sites was increased. The presence of nNOS was confirmed in the endothelial layer of mesenteric arteries from both sham and DOCA-salt-hypertensive mice. These results demonstrate that endothelial dysfunction in the mesenteric arteries of DOCA-salt-hypertensive mice is associated with reduced expression and functioning of nNOS and impaired production of nNOS-derived H2O2 Such findings offer a new perspective for the understanding of endothelial dysfunction in hypertension.
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Endotélio Vascular/metabolismo , Peróxido de Hidrogênio/metabolismo , Hipertensão/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Óxido Nítrico/metabolismo , Animais , Acetato de Desoxicorticosterona/farmacologia , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Hipertensão/fisiopatologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/metabolismo , Camundongos , Vasodilatadores/farmacologiaRESUMO
BACKGROUND: Hancornia speciosa Gomes is an herb traditionally used in Brazil for blood pressure control. PURPOSE: The present work investigated the antihypertensive effect of an extract from Hancornia speciosa leaves (SFH) and analyzed its underlying mechanisms of action. METHODS: Hypertension was induced in mice by surgical removal of a kidney and by subcutaneous administration of a pellet with deoxycorticosterone. Vasodilatation was measured in mesenteric arteries with a wire myograph. Nitrites were measured by fluorescence with 2,3-diaminonaphthalene and H2O2 was measured with carbon microsensors. RESULTS: SFH (0.03, 0.1 or 1 mg/kg; po) induced a dose-dependent, long-lasting reduction in the systolic blood pressure in conscious DOCA-salt hypertensive mice (DOCA). Administration of SFH produced a significant increase in the plasmatic level of nitrites. The systemic inhibition of nitric oxide synthase by L-NAME (20 mg/kg) reduced its antihypertensive effect. SFH also induced a concentration-dependent vasodilatation of mesenteric resistance arteries contracted with phenylephrine, which was more potent in arteries from DOCA mice. Removal of the endothelium or pretreatment with L-NAME or catalase reduced the vasodilator response for SFH. The nitrite production induced by SFH was significantly bigger in mesenteric arteries from DOCA than in SHAM mice. However, the production of H2O2 induced by SFH was twice higher in DOCA mice. CONCLUSION: Altogether, our results point to an antihypertensive effect of SFH due to a reduction in peripheral resistance through the production of NO and by a mechanism involving an increased production of H2O2 in the mesenteric arteries from hypertensive mice. These findings are further evidence to support the use of Hancornia speciosa by traditional medicine as an antihypertensive drug.
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Anti-Hipertensivos/farmacologia , Apocynaceae/química , Hipertensão/tratamento farmacológico , Extratos Vegetais/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Brasil , Desoxicorticosterona , Peróxido de Hidrogênio/metabolismo , Hipertensão/fisiopatologia , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Camundongos , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Nitritos/metabolismo , Folhas de Planta/química , Vasodilatação/efeitos dos fármacos , Vasodilatadores/farmacologiaRESUMO
NADPH oxidases derived reactive oxygen species (ROS) play an important role in vascular function and remodeling in hypertension through redox signaling processes. Previous studies demonstrated that protein disulfide isomerase (PDI) regulates Nox1 expression and ROS generation in cultured vascular smooth muscle cells. However, the role of PDI in conductance and resistance arteries during hypertension development remains unknown. The aim of the present study was to investigate PDI expression and NADPH oxidase dependent ROS generation during hypertension development. Mesenteric resistance arteries (MRA) and thoracic aorta were isolated from 6, 8, and 12 week-old spontaneously hypertensive (SHR) and Wistar rats. ROS production (dihydroethidium fluorescence), PDI (WB, imunofluorescence), Nox1 and NOX4 (RT-PCR) expression were evaluated. Results show a progressive increase in ROS generation in MRA and aorta from 8 to 12 week-old SHR. This effect was associated with a concomitant increase in PDI and Nox1 expression only in MRA. Therefore, suggesting a positive correlation between PDI and Nox1 expression during the development of hypertension in MRA. In order to investigate if this effect was due to an increase in arterial blood pressure, pre hypertensive SHR were treated with losartan (20 mg/kg/day for 30 days), an AT1 receptor antagonist. Losartan decreased blood pressure and ROS generation in both vascular beds. However, only in SHR MRA losartan treatment lowered PDI and Nox1 expression to control levels. In MRA PDI inhibition (bacitracin, 0.5 mM) decreased Ang II redox signaling (p-ERK 1/2). Altogether, our results suggest that PDI plays a role in triggering oxidative stress and vascular dysfunction in resistance but not in conductance arteries, increasing Nox1 expression and activity. Therefore, PDI could be a new player in oxidative stress and functional alterations in resistance arteries during the establishment of hypertension.