Kynurenine Relaxes Arteries of Normotensive Women and Those With Preeclampsia.

[Figure: see text].

Activation of AMPK/proteasome/MLCK degradation signaling axis by telmisartan inhibits VSMC contractility and vessel contraction.

Telmisartan, an angiotensin II type 1 receptor blocker (ARB), is widely used to treat hypertension. Dysfunction of vascular smooth muscle cells (VSMCs) is well-established to contribute to the pathogenesis of various vascular diseases. A growing body of evidence indicates that increased VSMC contractility plays a primary role in the development of pathological artery spasms. Nevertheless, effect of telmisartan on VSMC contractility, and its mechanism of action remain unknown. Here, we investigated the mechanism by which telmisartan inhibits VSMC contractility and vessel contraction in rat VSMCs and endothelium-deprived aortas. Telmisartan inhibited phenylephrine-induced vessel contraction in endothelium-deprived aortas, and decreased myosin light chain kinase (MLCK) levels (without altering corresponding mRNA levels) and myosin light chain (MLC) phosphorylation at Ser19 (p-MLC-Ser19) in VSMCs. MG-132 but not doxycycline significantly restored telmisartan-inhibited MLCK expression and p-MLC-Ser19. Telmisartan induced AMP-activated protein kinase (AMPK) phosphorylation at Thr172 (p-AMPK-Thr172), and compound C or ectopic expression of the dominant negative (dn)-AMPKα1 gene significantly reversed telmisartan-inhibited MLCK expression and p-MLC-Ser19. Of the ARBs tested (including losartan and fimasartan), only telmisartan increased p-AMPK-Thr172, and inhibited MLCK expression and p-MLC-Ser19. GW9662 had no effects on telmisartan-induced changes. Similar to the in vitro results, telmisartan enhanced p-AMPK-Thr172, and inhibited MLCK expression and p-MLC-Ser19 in endothelium-deprived aortas. Furthermore, the telmisartan-inhibited vessel contraction in the aortas was significantly reversed by MG-132 or compound C. In conclusion, we demonstrated that telmisartan inhibits VSMC contractility and vessel contraction by activating AMPK/proteasome/MLCK degradation signaling axis. These results suggest that telmisartan can be used to treat pathological vasospasms.

Mecanismos regulatorios del tono vascular pulmonar neonatal: una perspectiva molecular / Regulatory mechanisms of neonatal pulmonary vascular tone: a molecular perspective

La adaptación al medio extrauterino incluye un aumento considerable de la PaO2, que induce especialmente cambios estructurales y vasoactivos en la circulación pulmonar, que llevarán a una circulación previamente pobremente irrigada, a recibir ∼100% del gasto cardíaco del recién nacido, permitiendo el normal intercambio gaseoso. La regulación local de la circulación arterial pulmonar neonatal basal, es mantenida por un delicado equilibrio entre agentes vasoconstrictores y vasodilatadores. Este equilibrio, permite mantener la circulación pulmonar como un territorio de gran flujo sanguíneo y baja resistencia. La acción de los vasoconstrictores permite la formación de las interacciones entre actina y la cadena liviana de la miosina, esta es inducida en la célula muscular lisa principalmente por dos vías: a) dependiente de calcio, que consiste en aumentar el calcio intracelular, facilitando finalmente la unión de actina y miosina, y b) independiente de calcio, la cual a través de consecutivas fosforilaciones logra sensibilizar a las proteínas involucradas promoviendo la unión de actina y miosina. Estas acciones son mediadas por agonistas generados principalmente en el endotelio pulmonar, como endotelina-1 y tromboxano, o por agonistas provenientes de otros tipos celulares como la serotonina. Los agentes vasodilatadores regulan la respuesta vasoconstrictora, principalmente inhibiendo la señalización que induce la vasocontricción independiente de calcio, a través de la activación de proteínas quinasas que inhibirán la función de la ROCK quinasa, uno de los últimos efectores de la vasocontricción antes de la formación de la unión de actina y miosina. Esta revisión describe estos mecanismos de primordial importancia en las primeras horas de nuestra vida como individuos independientes.

The extrauterine-milieu adaptation includes a considerable increase in PaO2, that specifically induces structural and vasoactive changes at pulmonary circulation. Such changes transform a poor irrigated circulation into a circulation that receive ∼100% of neonatal cardiac output, supporting the normal alveolar-capillary gas exchange. Local regulation of basal neonatal pulmonary circulation is maintaining by a delicate equilibrium between vasoconstrictor and vasodilator agents. This equilibrium, allows to maintain the pulmonary circulation as an hemodynamic system with a high blood flow and a low vascular resistance. Vasocontrictors action allows actin and light-chain myosin interaction. Two main pathways induced this effect in smooth muscle cell: a) a calcium dependent pathway, that increases intracellular calcium, facilitating actin - myosin binding, and b) the independent calcium pathway, which achieves through consecutive phosphorylation reactions sensitize the proteins involved, promoting the binding of actin and light-chain myosin. These actions are mediated by agonists produced mainly in the pulmonary endothelium, such as endothelin-1 and thromboxane, or by agonists from other cell types such as serotonin. Vasodilator agents regulate the vasoconstrictor response, mainly by inhibiting signals that induce calcium-independent vasoconstriction, through activation of protein kinases, which in turn will inhibit the function of ROCK kinase, one of the last effectors of vasoconstriction before formation of the actin and light-chain myosin binding. This review will focus on describing these mechanisms of primal importance in the first hours of our lives as independent individuals.

TRPA1 and TRPV1 contribute to propofol-mediated antagonism of U46619-induced constriction in murine coronary arteries.

BACKGROUND: Transient receptor potential (TRP) ion channels have emerged as key components contributing to vasoreactivity. Propofol, an anesthetic is associated with adverse side effects including hypotension and acute pain upon infusion. Our objective was to determine the extent to which TRPA1 and/or TRPV1 ion channels are involved in mediating propofol-induced vasorelaxation of mouse coronary arterioles in vitro and elucidate the potential cellular signal transduction pathway by which this occurs. METHODS: Hearts were excised from anesthetized mice and coronary arterioles were dissected from control C57Bl/6J, TRPA1-/-, TRPV1-/- and double-knockout mice (TRPAV-/-). Isolated microvessels were cannulated and secured in a temperature-controlled chamber and allowed to equilibrate for 1 hr. Vasoreactivity studies were performed in microvessels pre-constricted with U46619 to assess the dose-dependent relaxation effects of propofol on coronary microvascular tone. RESULTS: Propofol-induced relaxation was unaffected in vessels obtained from TRPV1-/- mice, markedly attenuated in pre-constricted vessels obtained from TRPA1-/- mice and abolished in vessels obtained from TRPAV-/- mice. Furthermore, NOS inhibition with L-NAME or endothelium denuding abolished the proporfol-induced depressor response in pre-constricted vessels obtained from all mice. In the absence of L-NAME, BKCa inhibition with penitrem A markedly attenuated propofol-mediated relaxation in vessels obtained from wild-type mice and to a lesser extent in vessels obtained from TRPV1-/-, mice with no effect in vessels obtained from TRPA1-/- or TRPAV-/- mice. CONCLUSIONS: TRPA1 and TRPV1 appear to contribute to the propofol-mediated antagonism of U46619-induced constriction in murine coronary microvessels that involves activation of NOS and BKCa.

A peptide vaccine targeting angiotensin II attenuates the cardiac dysfunction induced by myocardial infarction.

A peptide vaccine targeting angiotensin II (Ang II) was recently developed as a novel treatment for hypertension to resolve the problem of noncompliance with pharmacotherapy. Ang II plays a crucial role in the pathogenesis of cardiac remodeling after myocardial infarction (MI), which causes heart failure. In the present study, we examined whether the Ang II vaccine is effective in preventing heart failure. The injection of the Ang II vaccine in a rat model of MI attenuated cardiac dysfunction in association with an elevation in the serum anti-Ang II antibody titer. Furthermore, any detrimental effects of the Ang II vaccine were not observed in the rats that underwent sham operations. Treatment with immunized serum from Ang II vaccine-injected rats significantly suppressed post-MI cardiac dysfunction in MI rats and Ang II-induced remodeling-associated signaling in cardiac fibroblasts. Thus, our present study demonstrates that the Ang II vaccine may provide a promising novel therapeutic strategy for preventing heart failure.

Increased hydrogen peroxide impairs angiotensin II contractions of afferent arterioles in mice after renal ischaemia-reperfusion injury.

AIM: Renal ischaemia-reperfusion injury (IRI) increases angiotensin II (Ang II) and reactive oxygen species (ROS) that are potent modulators of vascular function. However, the roles of individual ROS and their interaction with Ang II are not clear. Here we tested the hypothesis that IRI modulates renal afferent arteriolar responses to Ang II via increasing superoxide (O2-) or hydrogen peroxide (H2 O2 ). METHODS: Renal afferent arterioles were isolated and perfused from C57BL/6 mice 24 h after IRI or sham surgery. Responses to Ang II or noradrenaline were assessed by measuring arteriolar diameter. Production of H2 O2 and O2- was assessed in afferent arterioles and renal cortex. Activity of SOD and catalase, and mRNA expressions of Ang II receptors were assessed in pre-glomerular arterioles and renal cortex. RESULTS: Afferent arterioles from mice after IRI had a reduced maximal contraction to Ang II (-27±2 vs. -42±1%, P < 0.001), but retained a normal contraction to noradrenaline. Arterioles after IRI had a 38% increase in H2 O2 (P < 0.001) and a 45% decrease in catalase activity (P < 0.01). Contractions were reduced in normal arterioles after incubation with H2 O2 (-22±2 vs. -42±1%, P < 0.05) similar to the effects of IRI. However, the impaired contractions were normalized by incubation with PEG catalase despite a reduced AT1 R expression. CONCLUSIONS: Renal IRI in mice selectively impairs afferent arteriolar responses to Ang II because of H2 O2 accumulation that is caused by a reduced catalase activity. This could serve to buffer the effect of Ang II after IRI and may be a protective mechanism.

The Labdane Ent-3-Acetoxy-Labda-8(17), 13-Dien-15-Oic Decreases Blood Pressure In Hypertensive Rats / O Labdano Ácido Ent-3-Acetóxi-Labda-8(17), 13-Dieno-15-Óico Reduz Pressão Arterial Em Ratos Hipertensos

Abstract Background: Labdane-type diterpenes induce lower blood pressure via relaxation of vascular smooth muscle; however, there are no studies describing the effects of labdanes in hypertensive rats. Objective: The present study was designed to investigate the cardiovascular actions of the labdane-type diterpene ent-3-acetoxy-labda-8(17), 13-dien-15-oic acid (labda-15-oic acid) in two-kidney 1 clip (2K-1C) renal hypertension. Methods: Vascular reactivity experiments were performed in aortic rings isolated from 2K-1C and normotensive (2K) male Wistar rats. Nitrate/nitrite (NOx) measurement was performed in aortas by colorimetric assay. Blood pressure measurements were performed in conscious rats. Results: Labda-15-oic acid (0.1-300 µmol/l) and forskolin (0.1 nmol/l - 1 µmol/l) relaxed endothelium-intact and endothelium-denuded aortas from both 2K-1C and 2K rats. Labda-15-oic acid was more effective at inducing relaxation in endothelium-intact aortas from 2K pre-contracted with phenylephrine when compared to the endothelium-denuded ones. Forskolin was more potent than labda-15-oic acid at inducing vascular relaxation in arteries from both 2K and 2K-1C rats. Labda-15-oic acid-induced increase in NOx levels was lower in arteries from 2K-1C rats when compared to 2K rats. Intravenous administration of labda-15-oic acid (0.3-3 mg/kg) or forskolin (0.1-1 mg/kg) induced hypotension in conscious 2K-1C and 2K rats. Conclusion: The present findings show that labda-15-oic acid induces vascular relaxation and hypotension in hypertensive rats.

Resumo Fundamento: Diterpenos do tipo labdano induzem uma queda da pressão arterial por meio do relaxamento do músculo liso vascular; todavia, não há estudos que descrevam os efeitos de labdanos em ratos hipertensos. Objetivo: O presente estudo foi desenvolvido para investigar as ações cardiovasculares do labdano ácido ent-3-acetóxi-labda-8(17),13-dieno-15-óico (labda-15-óico) na hipertensão renal dois rins-1 clipe (2R-1C). Métodos: Foram feitos experimentos de reatividade vascular em anéis aórticos isolados de ratos machos 2R-1C e normotensos (2R). A medição de Nitrato/Nitrito (NOx) foi feita nas aortas por meio de ensaio colorimétrico. As medidas de pressão arterial foram feitas em ratos conscientes. Resultados: O ácido labda-15-óico (0,1 - 300 µmol/l) e a forscolina (0,1 nmol/l - 1 µmol/l) relaxaram as aortas com endotélio intacto e as aortas sem endotélio dos ratos 2R-1C e 2R. O labda-15-óico mostrou-se mais eficaz na indução do relaxamento em aortas com endotélio intacto de 2R pré-contraídas com fenilefrina em comparação àquelas sem endotélio. A forscolina mostrou-se mais potente do que o ácido labda-15-óico na indução do relaxamento vascular nas artérias tanto de ratos 2R-1C quanto de ratos 2R. O aumento dos níveis de NOx induzido pelo ácido labda-15-óico foi menor nas artérias de ratos 2R-1C em comparação a ratos 2R. A administração intravenosa de ácido labda-15-óico (0,3-3 mg/kg) ou forscolina (0,1-1 mg/kg) induziu hipertensão em ratos 2R-1C e 2R conscientes. Conclusão: Os presentes resultados mostram que o labda-15-óico induz relaxamento vascular e hipotensão em ratos hipertensos.

The Labdane Ent-3-Acetoxy-Labda-8(17), 13-Dien-15-Oic Decreases Blood Pressure In Hypertensive Rats.

BACKGROUND: Labdane-type diterpenes induce lower blood pressure via relaxation of vascular smooth muscle; however, there are no studies describing the effects of labdanes in hypertensive rats. OBJECTIVE: The present study was designed to investigate the cardiovascular actions of the labdane-type diterpene ent-3-acetoxy-labda-8(17), 13-dien-15-oic acid (labda-15-oic acid) in two-kidney 1 clip (2K-1C) renal hypertension. METHODS: Vascular reactivity experiments were performed in aortic rings isolated from 2K-1C and normotensive (2K) male Wistar rats. Nitrate/nitrite (NOx) measurement was performed in aortas by colorimetric assay. Blood pressure measurements were performed in conscious rats. RESULTS: Labda-15-oic acid (0.1-300 µmol/l) and forskolin (0.1 nmol/l - 1 µmol/l) relaxed endothelium-intact and endothelium-denuded aortas from both 2K-1C and 2K rats. Labda-15-oic acid was more effective at inducing relaxation in endothelium-intact aortas from 2K pre-contracted with phenylephrine when compared to the endothelium-denuded ones. Forskolin was more potent than labda-15-oic acid at inducing vascular relaxation in arteries from both 2K and 2K-1C rats. Labda-15-oic acid-induced increase in NOx levels was lower in arteries from 2K-1C rats when compared to 2K rats. Intravenous administration of labda-15-oic acid (0.3-3 mg/kg) or forskolin (0.1-1 mg/kg) induced hypotension in conscious 2K-1C and 2K rats. CONCLUSION: The present findings show that labda-15-oic acid induces vascular relaxation and hypotension in hypertensive rats.

Effects of cyclopiazonic acid and dexamethasone on serotonin-induced calcium responses in vascular smooth muscle cells.

We previously observed that sarcoendoplasmic reticulum Ca(2+) ATPase (SERCA) blockade by cyclopiazonic acid (CPA) significantly potentiates serotonin (5-hydroxytryptamine (5-HT))-induced vascular contractions. Furthermore, 5-HT receptor antagonist methysergide partially inhibited CPA-potentiated 5-HT contractions. In the present study, we further investigated whether SERCA inhibition potentiates 5-HT-induced Ca(2+) responses along with attenuating the receptor antagonism by store-operated Ca(2+) (SOC) entry and protein kinase C (PKC)-mediated mechanisms. The effects of dexamethasone that was previously shown to induce SOC entry and enhance 5-HT responses were also tested. For this purpose, intracellular Ca(2+) levels were monitored in A7r5 embryonic rat vascular smooth muscle cells by spectrofluorometry using the fluorescent indicator fura-2. The results showed that CPA, although not dexamethasone, significantly potentiated 5-HT-induced Ca(2+) elevations. Ketanserin partially decreased 5-HT-induced and CPA-potentiated Ca(2+) elevations whereas both PKC inhibitor D-sphingosine and SOC entry blocker 2-aminoethoxydiphenyl borate (2-APB) abolished the remaining responses. The data suggests that diminished antagonistic effect on 5-HT-induced Ca(2+) elevations in the presence of SERCA inhibition is induced by SOC entry and PKC activation.

Pharmacological evidence that NaHS inhibits the vasopressor responses induced by stimulation of the preganglionic sympathetic outflow in pithed rats.

It has been reported that i.v. administration of NaHS, a donor of H2S, elicited dose-dependent hypotension although the mechanisms are not completely understood. In this regard, several mechanisms could be involved including the inhibition of the vasopressor sympathetic outflow. Thus, this study was designed to determine the potential capability of NaHS to mediate inhibition of the vasopressor responses induced by preganglionic sympathetic stimulation. For this purpose, Wistar rats were anaesthetised, pithed and cannulated for drug administration. In animals pre-treated with gallamine, the effect of i.v. infusion of NaHS (310 and 560µg/kgmin) or its vehicle (phosphate buffer) was determined on the vasopressor responses induced by: (1) sympathetic stimulation (0.03-10Hz); (2) i.v. bolus injections of exogenous noradrenaline (0.03-3µg/kg); or (3) methoxamine (1-100µg/kg). The vasopressor responses induced by preganglionic sympathetic stimulation were dose-dependently inhibited by i.v. infusion of NaHS (310 and 560µg/kgmin), but not by vehicle, particularly at high frequencies. In marked contrast, the vasopressor responses to exogenous noradrenaline or methoxamine were not inhibited by the above doses of NaHS or its vehicle. The above results, taken together, demonstrate that NaHS inhibited the vasopressor responses induced by preganglionic sympathetic outflow by a prejunctional mechanism. This is the first evidence demonstrating this effect by NaHS that may contribute, at least in part, to the hypotension induced by NaHS.

Mechanisms Involved in Thromboxane A2 -induced Vasoconstriction of Rat Intracavernous Small Penile Arteries.

Diabetes is associated with erectile dysfunction and with hypercontractility in erectile tissue and this is in part ascribed to increased formation of thromboxane. Rho kinase (ROCK) is a key regulator of calcium sensitization and contraction in vascular smooth muscle. This study investigated the role of calcium and ROCK in contraction evoked by activation of the thromboxane receptors. Rat intracavernous penile arteries were mounted for isometric tension and intracellular calcium ([Ca2+ ]i ) recording and corpus cavernosum for measurements of MYPT1 phosphorylation. In penile arteries, U46619 by activation of thromboxane receptors concentration dependently increased calcium and contraction. U46619-induced calcium influx was blocked by nifedipine, a blocker of L-type calcium channels, and by 2-aminoethoxydiphenyl borate, a blocker of transient receptor potential (TRP) channels. Inhibitors of ROCK, Y27632 and glycyl-H1152P, concentration dependently reduced U46619-induced contraction, but only Y27632 reduced [Ca2+ ]i levels in the penile arteries activated with either high extracellular potassium or U46619. MYPT-Thr850 phosphorylation in corpus cavernous strips was increased in response to U46619 through activation of TP receptors and was found to be a direct result of phosphorylation by ROCK. Y27632 induced less relaxation in mesenteric arteries, H1152P induced equipotent relaxations, and a protein kinase C inhibitor, Ro-318220, failed to relax intracavernous penile arteries, but induced full relaxation in rat mesenteric arteries. Our findings suggest that U46619 contraction depends on Ca2+ influx through L-type and TRP channels, and ROCK-dependent mechanisms in penile arteries. Inhibition of the ROCK pathway is a potential approach for the treatment of erectile dysfunction associated with hypertension and diabetes.

3,3'-Diindolylmethane attenuates cardiac H9c2 cell hypertrophy through 5'-adenosine monophosphate-activated protein kinase-α.

3,3'-Diindolylmethane (DIM) is the major product of the acid-catalyzed condensation of indole-3-carbinol (I3C), a component of extracts of Brassica food plants. Numerous studies have suggested that DIM has several beneficial biological activities, including elimination of free radicals, antioxidant and anti-angiogenic effects and activation of apoptosis of various tumor cells. In the present study, an in vitro model was established, using 1 µM angiotensin II (Ang II) in cultured rat cardiac H9c2 cells, to observe the effects of DIM on cardiac hypertrophy. Following 24 h stimulation with DIM (1, 5, and 10 µM) with or without Ang II, cells were characterized by immunofluorescence to analyze cardiac α-actinin expression. Cardiomyocyte hypertrophy and molecular markers of cardiac hypertrophy were assessed by quantitative polymerase chain reaction. Atrial natriuretic peptide, brain natriuretic peptide and myosin heavy chain ß mRNA expression were induced by Ang II in H9c2 cells treated with the optimal concentration of DIM for 6, 12, and 24 h. The levels of phosphorylated and total proteins of the 5' AMP-activated protein kinase α (AMPKα)/mitogen-activated protein kinase (MAPK)/mechanistic target of rapamycin (mTOR) signaling pathways in H9c2 cells treated with DIM for 0, 15, 30, and 60 min induced by Ang II were determined by western blot analysis. The results showed that DIM attenuated cellular hypertrophy in vitro, enhanced the phosphorylation of AMPKα and inhibited the MAPK­mTOR signaling pathway in response to hypertrophic stimuli.

Vasodilatory effect of a novel Rho-kinase inhibitor, DL0805-2, on the rat mesenteric artery and its potential mechanisms.

PURPOSE: In the present study, we investigated the vasodilatory effect of a novel scaffold Rho-kinase inhibitor, DL0805-2, on isolated rat arterial rings including mesenteric, ventral tail, and renal arteries. We also examined the potential mechanisms of its vasodilatory action using mesenteric artery rings. METHODS: A DMT multiwire myograph system was used to test the tension of isolated small arteries. Several drugs were employed to verify the underlying mechanisms. RESULTS: DL0805-2 (10(-7)-10(-4) M) inhibited KCl (60 mM)-induced vasoconstriction in three types of small artery rings (pEC50: 5.84 ± 0.03, 5.39 ± 0.03, and 5.67 ± 0.02 for mesenteric, renal, and ventral tail artery rings, respectively). Pre-incubation with DL0805-2 (1, 3, or 10 µM) attenuated KCl (10-60 mM) and angiotensin II (AngII; 10(-6) M)-induced vasoconstriction in mesenteric artery rings. The relaxant effect on the rat mesenteric artery was partially endothelium-dependent (pEC50: 6.02 ± 0.05 for endothelium-intact and 5.72 ± 0.06 for endothelium-denuded). The influx and release of Ca(2+) were inhibited by DL0805-2. In addition, the increased phosphorylation levels of myosin light chain (MLC) and myosin-binding subunit of myosin phosphatase (MYPT1) induced by AngII were blocked by DL0805-2. However, DL0805-2 had little effect on K(+) channels. CONCLUSIONS: The present results demonstrate that DL0805-2 has a vasorelaxant effect on isolated rat small arteries and may exert its action through the endothelium, Ca(2+) channels, and the Rho/ROCK pathway.

Novel vasocontractile role of the P2Y14 receptor: characterization of its signalling in porcine isolated pancreatic arteries.

BACKGROUND AND PURPOSE: The P2Y14 receptor is the newest member of the P2Y receptor family; it is G(i/o) protein-coupled and is activated by UDP and selectively by UDP-glucose and MRS2690 (2-thiouridine-5'-diphosphoglucose) (7-10-fold more potent than UDP-glucose). This study investigated whether P2Y14 receptors were functionally expressed in porcine isolated pancreatic arteries. EXPERIMENTAL APPROACH: Pancreatic arteries were prepared for isometric tension recording and UDP-glucose, UDP and MRS2690 were applied cumulatively after preconstriction with U46619, a TxA2 mimetic. Levels of phosphorylated myosin light chain 2 (MLC2) were assessed with Western blotting. cAMP concentrations were assessed using a competitive enzyme immunoassay kit. KEY RESULTS: Concentration-dependent contractions with a rank order of potency of MRS2690 (10-fold) > UDP-glucose ≥ UDP were recorded. These contractions were reduced by PPTN {4-[4-(piperidin-4-yl)phenyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthoic acid}, a selective antagonist of P2Y14 receptors, which did not affect responses to UTP. Contraction to UDP-glucose was not affected by MRS2578, a P2Y6 receptor selective antagonist. Raising cAMP levels and forskolin, in the presence of U46619, enhanced contractions to UDP-glucose. In addition, UDP-glucose and MRS2690 inhibited forskolin-stimulated cAMP levels. Removal of the endothelium and inhibition of endothelium-derived contractile agents (TxA2, PGF(2α) and endothelin-1) inhibited contractions to UDP glucose. Y-27632, nifedipine and thapsigargin also reduced contractions to the agonists. UDP-glucose and MRS2690 increased MLC2 phosphorylation, which was blocked by PPTN. CONCLUSIONS AND IMPLICATIONS: P2Y14 receptors play a novel vasocontractile role in porcine pancreatic arteries, mediating contraction via cAMP-dependent mechanisms, elevation of intracellular Ca²âº levels, activation of RhoA/ROCK signalling and MLC2, along with release of TxA2, PGF(2α) and endothelin-1.

Selective and potent inhibitory effect of docosahexaenoic acid (DHA) on U46619-induced contraction in rat aorta.

Inhibitory effects of docosahexaenoic acid (DHA) on blood vessel contractions induced by various constrictor stimulants were investigated in the rat thoracic aorta. The inhibitory effects of DHA were also compared with those of eicosapentaenoic acid (EPA) and linoleic acid (LA). DHA exhibited a strong inhibitory effect on the sustained contractions induced by U46619, a TXA(2) mimetic. This inhibitory effect of DHA was not affected by removal of the endothelium or by treatment with either indomethacin or N(ω)-nitro-l-arginine. DHA also significantly diminished PGF(2α)-induced contraction but did not show any appreciable inhibitory effects on the contractions to both phenylephrine (PE) and high-KCl. Similarly, EPA exhibited significant inhibitory effects against the contractions induced by both U46619 and PGF(2α) without substantially affecting either PE- or high-KCl-induced contractions. However, both DHA and EPA generated more potent inhibitions against contractions induced by U46619 than those by PGF(2α). In contrast, LA did not show significant inhibitory effects against any contractions, including those induced by U46619. The present findings suggest that DHA and EPA elicit more selective inhibition against blood vessel contractions that are mediated through stimulation of prostanoid receptors than those through α-adrenoceptor stimulation or membrane depolarization. Although DHA and EPA have similar inhibitory potencies against prostanoid receptor-mediated contractions, they had a more potent inhibition against TXA(2) receptor (TP receptor)-mediated contractions than against PGF(2α) receptor (FP receptor)-mediated responses. Selective inhibition by either DHA or EPA of prostanoid receptor-mediated blood vessel contractions may partly underlie the mechanisms by which these ω-3 polyunsaturated fatty acids exert their circulatory-protective effects.

Nifedipine inhibits angiotensin II-induced cardiac fibrosis via downregulating Nox4-derived ROS generation and suppressing ERK1/2, JNK signaling pathways.

Nifedipine, a classic L-type dihydropyridine calcium channel blocker (CCB), has been reported to possess multiple cardioprotective properties. However, little is known about the effects of nifedipine on cardiac fibrosis induced by angiotensinII (AngII) and the detailed molecular mechanisms. In this study, we found that nifedipine attenuated AngII-induced cardiac fibrosis in vitro via inhibiting the proliferation, differentiation of cardiac fibroblasts and antagonizing the upregulation of extracellular matrix (ECM) protein fibronectin (FN) and the pro-fibrotic cytokine connective tissue growth factor (CTGF). Furthermore, nifedipine suppressed the upregulation of NAD(P)H oxidase 4 (Nox4) and the production of reactive oxygen species (ROS) induced by AngII. In addition, it markedly inhibited the phosphorylation of extracellular signal-regulate kinases 1/2 (ERK1/2) and c-Jun NH(2)-terminal kinase (JNK) stimulated by AngII. However, nifedipine exhibited no effect on the variation of intracellular Ca2+ concentration ([Ca2+]i). These results suggested that (1) nifedipine inhibited cardiac fibrosis induced by AngII; (2) the anti-fibrotic effects of nifedipine may be mediated by interfering with the production of ROS and the activation of ERK1/2 and JNK signaling pathways; (3) the classic calcium channel blocking action of nifedipine may not be involved in the anti-fibrotic activities.

Vasorelaxant effects of the extracts and some flavonoids from the buds of Coreopsis tinctoria.

CONTEXT: The buds of Coreopsis tinctoria Nutt (Compositae) are used in the treatment of hypertension in the Uyghur folk medicine in China. OBJECTIVE: To investigate vasorelaxant properties of extracts and some flavonoids from C. tinctoria (CT) and their underlying mechanisms in isolated rat thoracic aortic rings. MATERIALS AND METHODS: Vasorelaxant effects of ethanol extracts of CT (CTA) and its flavonoids as well as water-ethanol eluates from CTA by AB-8 resins (CTAA∼CTAF) were evaluated on rat aortic rings pre-contracted with phenylephrine (PE, 1 µM) or high KCl (60 µM). We evaluated the effect of CTA, CTAD and CTAE on PE-induced contraction in a Ca²âº-free medium and a dose-effect curve of Ca²âº in pre-contracted ring with high KCl. RESULTS: Endothelial removal did not modify the effect of CTAD and CTAE (3.00 g/L) neither on PE-pre-contracted rings (164.78 ± 21.44 and 191.47 ± 16.75%) nor on KCl-pre-contracted rings (75.68 ± 10.76 and 125.14 ± 17.41%) compared with intact-endothelium rings pre-contracted with high KCl (100.49 ± 17.30 and 110.81 ± 16.33%). CTAD and CTAE (3.00 g/L) down-regulated the dose-effect curve of Ca²âº in pre-contraction with high KCl, and inhibited the pre-contraction with PE in a Ca²âº-free medium (p < 0.05). Seven flavonoids were obtained from CTAD, of which luteolin (5) and quercetin (6) were found to be the most effective relaxation in rings precontracted with PE (EC50: 0.006 and 0.039 g/L, p < 0.05) or high KCl (EC50: 0.023 and 0.045 g/L, p < 0.05). DISCUSSION AND CONCLUSION: These data demonstrated the vasorelaxant effect of CT, and its mechanism is likely due to an inhibitory effect on calcium movements through cell membranes.

Nitric-oxide synthase knockout modulates Ca²âº-sensing receptor expression and signaling in mouse mesenteric arteries.

Extracellular calcium (Ca²âº(e))-induced relaxation of isolated, phenylephrine (PE)-contracted mesenteric arteries is dependent on an intact perivascular sensory nerve network that expresses the Ca²âº-sensing receptor (CaSR). Activation of the receptor stimulates an endocannabinoid vasodilator pathway, which is dependent on cytochrome P450 and phospholipase A2 but largely independent of the endothelium. In the present study, we determined the role of nitric oxide (NO) in perivascular nerve CaSR-mediated relaxation of PE-contracted mesenteric resistance arteries isolated from mice. Using automated wire myography, we studied the effects of NO synthase (NOS) gene knockout (NOS(-/-)) and pharmacologic inhibition of NOS on Ca²âº(e)-induced relaxation of PE-contracted arteries. Endothelial NOS knockout (eNOS(-/-)) upregulates but neuronal NOS knockout (nNOS(-/-)) downregulates CaSR expression. NOS(-/-) reduced maximum Ca²âº(e)-induced relaxation with no change in EC50 values, with eNOS(-/-) having the largest effect. The responses of vessels to calindol and Calhex 231 indicate that the CaSR mediates relaxation. L-N5-(1-iminoethyl)-ornithine reduced Ca²âº(e)-induced relaxation of PE-contracted arteries from C57BL/6 control mice by ≈38% but had a smaller effect in vessels from eNOS(-/-) mice. 7-Nitroindazole had no significant effect on relaxation of arteries from NOS(-/-) mice, but both N(G)-nitro-L-arginine methylester and N(G)-monomethyl-L-arginine significantly reduced the relaxation maxima in all groups. Interestingly, the nNOS-selective inhibitor S-methyl-L-thiocitrulline significantly increased the EC50 value by ≈60% in tissues from C57BL/6 mice but reduced the maximum response by ≈80% in those from nNOS(-/-) mice. Ca²âº-activated big potassium channels play a major role in the process, as demonstrated by the effect of iberiotoxin. We conclude that CaSR signaling in mesenteric arteries stimulates eNOS and NO production that regulates Ca²âº(e)-induced relaxation.

Low-dose angiotensin II infusion restores vascular function in cerebral arteries of high salt-fed rats by increasing copper/zinc superoxide dimutase expression.

BACKGROUND: This study examined the vasoprotective role of circulating angiotensin II (ANG II) levels in the cerebral circulation of high salt (HS)-fed (SS.BN-(D13hmgc41-13hmgc23)/Mcwi) (Ren1-BN) congenic rats, which carry a normally functioning renin allele from the Brown Norway (BN) rat on the Dahl salt-sensitive genetic background. METHODS: Ren1-BN rats were placed on an HS (4.0% NaCl) diet for 3 days. The vasodilator response to acetylcholine (ACh; 10(-10) - 10(-6) mol/L) was assessed in isolated middle cerebral arteries (MCAs), and Western blots were performed to assess the expression of the antioxidant enzymes copper (Cu)/zinc (Zn) superoxide dismutase (SOD) and manganese (Mn) SOD in cerebral resistance vessels. A separate group of HS-fed animals were infused with either a subpressor dose of ANG II (100ng/kg/min) or saline vehicle via osmotic minipump for 3 days. RESULTS: HS diet eliminated acetylcholine (ACh)-induced dilation in the MCAs of the congenic rats. Western blot analysis of antioxidant enzymes showed that Cu/Zn SOD and Mn SOD expression were significantly reduced in the cerebral resistance arteries of the HS-fed rats compared with control animals fed a normal salt diet. Infusion of ANG II restored the vasodilator response to ACh in the MCAs and increased Cu/Zn SOD (but not Mn SOD) expression compared with saline-infused animals. CONCLUSIONS: These results indicate that prevention of salt-induced ANG II suppression prevents vascular dysfunction in the cerebral circulation by preventing the downregulation of Cu/Zn SOD and vascular oxidant stress that normally occurs with HS diet.

Angiotensin-(1-7) reduces the perfusion pressure response to angiotensin II and methoxamine via an endothelial nitric oxide-mediated pathway in cirrhotic rat liver.

Recent studies have shown that, in cirrhosis, portal angiotensin-(1-7) [Ang-(1-7)] levels are increased and hepatic expression of angiotensin converting enzyme 2 (ACE2) and the Mas receptor are upregulated, but the effects of Ang-(1-7) on hepatic hemodynamics in cirrhosis have not been studied. This study investigated the effects of Ang-(1-7) on vasoconstrictor-induced perfusion pressure increases in cirrhotic rat livers. Ang II or the alpha 1 agonist methoxamine (MTX) were injected in the presence or absence of Ang-(1-7), and the perfusion pressure response was recorded. Denudation of vascular endothelial cells with sodium deoxycholate was used to investigate the contribution of endothelium to the effects of Ang-(1-7). Ang-(1-7) alone had no effect on perfusion pressure. However, it reduced the maximal vasoconstriction response and area under the pressure response curve to Ang II and MTX by >50% (P < 0.05). This effect of Ang-(1-7) was not blocked by Mas receptor inhibition with A779 or by Ang II type 1 and type 2 receptor and bradykinin B(2) receptor blockade and was not reproduced by the Mas receptor agonist AVE0991. D-Pro(7)-Ang-(1-7), a novel Ang-(1-7) receptor antagonist, completely abolished the vasodilatory effects of Ang-(1-7), as did inhibition of endothelial nitric oxide synthase (eNOS) with N(G)-nitro-L-arginine methyl-ester, guanylate cyclase blockade with ODQ and endothelium denudation. The functional inhibition by D-Pro(7)-Ang-(1-7) was accompanied by significant (P < 0.05) inhibition of eNOS phosphorylation. This study shows that Ang-(1-7) significantly inhibits intrahepatic vasoconstriction in response to key mediators of increased vascular and sinusoidal tone in cirrhosis via a receptor population present on the vascular endothelium that is sensitive to D-Pro(7)-Ang-(1-7) and causes activation of eNOS and guanylate cyclase-dependent NO signaling pathways.