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Thoracic aortic aneurysm and dissection (TAAD) is caused by the apoptosis and phenotypic transformation of vascular smooth muscle cells (VSMCs). The dysfunction of VSMCs affects their secretion of chemokines such as monocyte chemoattractant protein-1 (MCP-1) to recruit the infiltration of macrophages which release proinflammatory cytokines and matrix metalloproteinases (MMPs) to accelerate the process of TAAD formation.We analyzed the expression levels of nuclear factor erythroid 2-related factor 2 (Nrf2) in aortic tissues of TAAD patients and the β-aminopropionitrile fumarate (BAPN)-induced mouse model, and the levels of Nrf2 were elevated in both aortic lesions. Treatment with the Nrf2 activator oltipraz protects against the formation of BAPN-induced aneurysm and dissection, as demonstrated by a higher survival rate, postponing the time of aortic rupture, and inhibiting aortic luminal dilation. In addition, the thoracic aortas of BAPN-treated mice inhibited the apoptosis and phenotypic transformation of VSMCs. When treated with oltipraz, they had reduced macrophage infiltration proinflammatory cytokines and MMPs. Furthermore, oltipraz treatment promoted the translocation of Nrf2 and downregulated the NLRP3 pathway.Nrf2 plays a crucial role in protecting against TAAD development, and persistent activation of Nrf2 is a promising therapeutic strategy against the progression of TAAD.

Hypertension is one of the most common complications of chronic kidney disease (CKD). Some research has indicated that changes in large artery function especially caused by thromboxane A2 (TXA2) may be a novel factor acting to induce hypertension in CKD. We studied the 5/6 nephrectomy rat model and measured serum levels of creatinine (Cr), calcium (Ca), phosphorus (P), TXA2-stable metabolites (thromboxane B2, TXB2), and caudal artery pressure after nephrectomy. The tension variations in thoracic aortas were measured after stimulating by vasoconstrictor/vasodilator using the cumulative concentration administration method and then tested the expression of TXA2 receptors in the thoracic aortas through western blots. The CKD rats developed uremia, electrolyte imbalances，and hypertension. They also exhibited a significant increase in TXB2 concentration. The aortic rings of CKD rats showed an increased contraction response to U46619 (a TXA2 analogue) and the expression of TXA2 receptors also enhanced. In the meanwhile, the diastolic function decreased in the CKD group. Our results demonstrate that the impairment of artery contractile function caused by the increase of TXA2 receptors on the wall of aortic rings may be involved in hypertension in CKD rats.

Different subtypes of GABA

In the present study, the therapeutic effects of imperatorin on metabolic and vascular alterations and possible underlying mechanisms were investigated in high-fat/high-fructose diet (HFFD)-fed rats. Male Sprague-Dawley rats were fed a high-fat diet plus 15% fructose in drinking water for 16 weeks. HFFD-fed rats were treated with imperatorin (15 or 30 mg/kg/day) for the last 4 weeks. In HFFD-fed rats, imperatorin significantly reduced obesity, hypertension, dyslipidemia, and insulin resistance. Imperatorin markedly improved vascular endothelial function and alleviated changes in vascular morphology. Furthermore, imperatorin treatment significantly increased the plasma levels of the nitric oxide metabolite and adiponectin, and upregulated adiponectin receptor 1 and endothelial nitric oxide synthase (eNOS) protein expression in the thoracic aorta. Imperatorin treatment decreased vascular superoxide anion production and downregulated aortic NADPH oxidase subunit p47phox protein expression. These findings indicated that imperatorin alleviates HFFD-induced metabolic and vascular alterations in rats. The possible underlying mechanism may involve the restoration of adiponectin receptor 1 and eNOS expression and suppression of p47phox expression.

Gestational diabetes mellitus (GDM) affects 5–10% of pregnancies and increases the risk of fetal and maternal adverse outcomes. Interestingly, the vascular response to AngII is decreased by pregnancy while the response is increased by diabetes. It remains unclear how GDM affects vascular tone and how angiotensin II receptors contribute to these changes. In this work, we sought to establish the vascular impact of a hypercaloric diet-induced GDM through changes in AT1 and AT2 receptor's expression. Female rats fed for 7 weeks with standard (SD) or hypercaloric (HD) diet were divided at week 4. Half of the rats of each group were mated to become pregnant and those fed with a HD developed GDM. AngII-induced vasoconstriction was measured in thoracic or abdominal aorta rings using a conventional isolated organ bath and AT1 and AT2 receptors were searched by immunohistochemistry. Experiments where conducted on the pregnant standard diet group (PSD) and the pregnant hypercaloric-gestational diabetes mellitus group (PHD-GDM). Vasoconstriction was reduced in the thoracic aorta (P

Copeptin, a glycosylated peptide fragment derived from the C-terminal region of the precursor of arginine

Abnormal aortic adventitial fibroblasts (AFs) play essential roles in the development of vascular remodeling and disorders. Previous studies revealed that microRNA-122 (miR-122) levels were elevated in the aortic adventitia of hypertensive rats with vascular injury. Here, we aim to evaluate the biological effects and underlying mechanisms of miR-122 in rat AFs. Exposure to angiotensin II (ATII) in rat AFs resulted in decreased levels of sirtuin 6 (SIRT6), elabela (ELA), and angiotensin-converting enzyme 2 (ACE2). Additionally, stimulation with ATII contributed to a decline in autophagic flux and an increase in cellular migration, oxidative stress, and apoptosis, which were exacerbated by the transfection of miR-122-5p mimic but were rescued by miR-122-5p inhibitor, exogenous replenishment of ELA, and recombinant adeno-associated virus expressing SIRT6 (rAAV-SIRT6). Moreover, stimulation with miR-122-5p mimic led to a marked reduction in the levels of SIRT6 and ELA in rat AFs, which were elevated by stimulation with rAAV-SIRT6. Furthermore, miR-122-5p inhibitor-mediated pro-autophagic, anti-oxidant and anti-apoptotic effects in rat AFs were partially suppressed by 3-methyladenine, SIRT6 small interfering RNA (siRNA) and ELA siRNA, which were linked with the downregulation in the protein levels of LC3-II, beclin-1, and ACE2 and the upregulation of p62 expression and bax/bcl-2 ratio. Our findings indicated that miR-122-5p inhibition prevented ATII-mediated loss of autophagy, and the promotion of apoptosis and oxidative stress via activating the SIRT6-ELA-ACE2 signaling. MiR-122-5p may be a novel predictive biomarker of adventitial injury, and targeting the SIRT6-ELA-ACE2 signaling may have the potential therapeutic importance of controlling vascular remodeling and disorders., Highlights • MiR-122-5p inhibitor prevents angiotensin II-mediated loss of SIRT6 and autophagy and promotion of apoptosis. • Treatment with rAAV-SIRT6 inhibits miR-122-5p mimic-mediated loss of ELA and autophagy and promotion of apoptosis. • MiR-122-5p inhibitor-induced beneficial roles in autophagy and apoptosis are suppressed by ELA siRNA. • MiR-122-5p exerts pro-apoptotic and anti-autophagic effects via the modulation of SIRT6-Elabela-ACE2 signaling.

Benzodiazepines (BDZs) and non-BDZ sedative-hypnotics are effective for the management of chronic insomnia; however, they are associated with adverse effects such as headache, dizziness, and palpitations. Furthermore, long-term use of these medications is associated with decreased blood pressure (BP) or depressed baroreflex function. Therefore, here, we assessed whether BDZs and non-BDZs cause vasorelaxation directly. Vasorelaxation in response to 22 BDZs, 2 non-BDZs, and tandospirone was determined by myograph methods using isolated Wistar rat thoracic aortas. All the drugs relaxed phenylephrine-contracted rat aortas in a concentration-dependent manner. Zolpidem and tandospirone caused over 80% relaxation at a concentration of 10 μM; diazepam, estazolam, etizolam, and tofisopam caused 60–70% relaxation; whereas 18 other BDZs (alprazolam, bromazepam, brotizolam, chlordiazepoxide, clobazam, clonazepam, clorazepate, ethyl loflazepate, flunitrazepam, flurazepam, lorazepam, lormetazepam, midazolam, nimetazepam, nitrazepam, oxazepam, temazepam, and triazolam) and zaleplon caused less than 50% relaxation. The relaxation was partially but significantly inhibited to the same extent by a nitric oxide (NO) synthase antagonist and after endothelium removal. Binding assay of gamma-aminobutyric acid type A receptors was performed using [3H]flunitrazepam. No correlation was observed between vasorelaxation at a concentration of 10 μM and the binding affinities for 23 drugs. The study demonstrated that zaleplon, zolpidem, tandospirone, and many BDZs cause vasorelaxation to different extents via endothelial NO-dependent and endothelium-independent pathways. In conclusion, the direct vasodilatory effects of these drugs may be involved in the mechanisms underlying their adverse effects. Additionally, the decreased BP observed in persons who take BDZs or non-BDZs may be partly due to direct vasodilation.

Compounds containing a nitro group may reveal vasodilator properties. Several nitro compounds have a NO2 group in a short aliphatic chain connected to an aromatic group. In this study, we evaluated in rat aorta the effects of two nitro compounds, with emphasis on a putative recruitment of the soluble guanylate cyclase (sGC) pathway to induce vasodilation. Isolated aortic rings were obtained from male Wistar rats to compare the effects induced by 2-nitro-1-phenylethanone (NPeth) or 2-nitro-2-phenyl-propane-1,3-diol (NPprop). In aortic preparations contracted with phenylephrine or KCl, NPeth and NPprop induced vasorelaxant effects that did not depend on the integrity of vascular endothelium. NPeth had a lesser vasorelaxant efficacy than NPprop and only the NPprop effects were inhibited by pretreatment with the sGC inhibitors, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or methylene blue. In an ODQ-preventable manner, NPprop inhibited the contractile component of the phenylephrine-induced response mediated by intracellular Ca2+ release or by extracellular Ca2+ recruitment through receptor- or voltage-operated Ca2+ channels. In contrast, NPprop was inert against the transient contraction induced by caffeine in Ca2+-free medium. In an ODQ-dependent manner, NPprop inhibited the contraction induced by the protein kinase C activator phorbol 12,13-dibutyrate or by the tyrosine phosphatase inhibitor sodium orthovanadate. In silico docking analysis of a sGC homologous protein revealed preferential site for NPprop. In conclusion, the nitro compounds NPeth and NPprop induced vasorelaxation in rat aortic rings. Aliphatic chain substituents selectively interfered in the ability of these compounds to induce vasorelaxant effects, and only NPprop relaxed aortic rings via a sGC pathway.

Coagulopathy is the major cause of organ injury as well as a strong predictor of mortality in septic patients. Systemic inflammatory response and redox imbalance are regarded as the major causes of sepsis-induced coagulopathy. There is growing evidence that a vasodilator hydralazine has beneficial effects on heart failure, hypertension, and ischemia/reperfusion injury via its antioxidant and anti-inflammatory properties. However, the effects of hydralazine on sepsis have not been examined. Therefore, we evaluated the effects of low-dose hydralazine on coagulopathy and multiple organ dysfunction in septic rats induced by endotoxin. Sepsis-induced coagulopathy was established by intravenous injection of rats with lipopolysaccharide (LPS). The changes of blood pressure, heart rate, blood glucose, hemostatic variables, prothrombin time, organ function indices, interleukin-6 (IL-6) concentration, and nitric oxide (NO) level were assessed during the experimental period. In addition, the aortas, lungs, livers, and kidneys were dissected to analyze superoxide levels and protein expressions. LPS induced (i) coagulopathy, multiple organ dysfunction, and circulatory failure successfully, and (ii) excessive superoxide, NO, and IL-6 production, accompanied by the overexpression of iNOS and Wnt5a in animals. Treatment of LPS-induced septic rats with low-dose hydralazine not only improved coagulopathy but also ameliorated multiple organ dysfunction. These could be due to attenuation of the overproduction of superoxide, NO, and IL-6, which were attributed to reduction of the overexpression of iNOS and Wnt5a. Thus, these findings indicate that low-dose hydralazine could be a potential therapy for sepsis-induced coagulopathy and multiple organ dysfunction via its anti-inflammatory and anti-oxidative/nitrosative properties.

This study investigated the vasodilatory effects and acting mechanism of gemigliptin, a dipeptidyl peptidase-4 (DPP-4) inhibitor. Tests were conducted in aortic rings pre-contracted with phenylephrine. Gemigliptin induced dose-dependent vasodilation of the aortic smooth muscle. Several pre-treatment groups were used to investigate the mechanism of action. While pre-treatment with paxilline, a large-conductance Ca

Recent studies reported cardioprotective effects of sodium glucose co-transporter 2 (SGLT2) inhibitors; however, the underlying mechanisms are still obscure. Here, we investigated whether empagliflozin attenuates atherogenesis and endothelial dysfunction in diabetic apolipoprotein E-deficient (ApoE-/-) mice. Male streptozotocin (STZ) - induced diabetic ApoE-/- mice were treated with empagliflozin for 12 or 8 weeks. Empagliflozin lowered blood glucose (P

Vascular calcification is a highly regulated process similar to osteogenesis involving phenotypic change of vascular smooth muscle cells (VSMCs). 25-Hydroxycholesterol (25-HC), one of oxysterols synthesized by the enzyme cholesterol 25-hydroxylase, has been shown to promote bovine calcifying vascular cells (CVC) calcification. However, whether and how 25-HC regulates vascular calcification are not completely understood. In this study, in vitro and ex vivo models of vascular calcification were used to determine whether 25-HC regulates vascular calcification. Alizarin red staining and calcium content assay showed that 25-HC treatment promoted calcification of rat and human VSMCs in a dose-dependent manner. Similarly, ex vivo study further confirmed that 25-HC accelerated calcification of rat aortic rings. In addition, western blot analysis showed that 25-HC significantly up-regulated the expression of endoplasmic reticulum stress (ERS) signaling molecules including ATF4 and CHOP in VSMCs and flow cytometry analysis revealed that 25-HC increased apoptosis of VSMCs. Moreover, knockdown of CHOP by siRNA blocked 25-HC-induced mineral deposition in VSMCs. Collectively, this study for the first time demonstrates that 25-HC promotes vascular calcification via ATF4/CHOP signaling using in vitro and ex vivo models, suggesting that ERS is involved in the regulation of 25-HC-induced vascular calcification.

Certain catecholamine metabolites exert significant pharmacological effects. Herein, we evaluated the pharmacological activities of catecholamine metabolites in the rat thoracic aorta, prostate, and spleen to determine whether these metabolites affect the contractile functions of smooth muscle tissue via direct action on α-adrenoceptors and α-adrenoceptor subtypes. Among the catecholamine metabolites examined, normetadrenaline and metadrenaline (10

The incretin glucagon-like peptide-1 (GLP-1) elicits direct favorable effects on the cardiovascular system. This study aimed to evaluate the acute effects of GLP-1 on improving aortic endothelial dysfunction in diabetic mice. Additionally, we examined whether GLP-1 elucidated the underlying mechanisms. Using the diabetic mouse models induced by nicotinamide and streptozotocin, we investigated the functional changes in the aorta caused by GLP-1. Organ baths were performed for vascular reactivity in isolated aortic rings, and western blotting was used for protein analysis. The diabetic aortas showed enhanced GLP-1-induced relaxation response and nitric oxide (NO) production. However, the pretreatment of GLP-1 did not significantly change the endothelial-dependent relaxation response to acetylcholine and -independent relaxation response to sodium nitroprusside. On the other hand, the GLP-1-induced relaxation response and NO production were abolished by the endothelial NO synthase inhibitor, GLP-1 receptor antagonist, Akt inhibitor, and AMP-activated protein kinase (AMPK) inhibitor. Finally, in diabetic mice, considerable increases in phosphorylation of Akt and AMPK were found in aortas stimulated with GLP-1, both of which were decreased by pretreatment with the AMPK inhibitor. GLP-1 significantly enhanced endothelial-dependent relaxation in diabetic aortas. The effect may be mediated through activation of the AMPK/Akt pathway via a GLP-1 receptor-dependent mechanism.

Pseudoprotodioscin (PPD), a phytoestrogen isolated from Dioscorea nipponica Makino, is recognized to possess anti-inflammatory and antiadipogenic capacities. However, little is known about the antiatherosclerotic effects of PPD and the underlying mechanisms. Here, the contribution of estrogen receptors (ERs) and inflammation to PPD-mediated amelioration of endothelial dysfunction has been fully assessed. PPD administration alleviated atherosclerotic lesions by lowering total cholesterol in ovariectomized apoE-/- mice fed a high-cholesterol diet. Molecular docking analysis suggested a selective interaction of PPD with ERα. Upon PPD treatment, ERα and endothelial nitric oxide synthase (eNOS) protein levels were increased, whereas cell adhesion molecule and monocyte chemoattractant protein-1 (MCP-1) mRNA levels were suppressed in human umbilical vein endothelial cells (HUVECs) after injury caused by oxidized low-density lipoprotein (ox-LDL). These effects could be abolished by an ERα antagonist or a NOS inhibitor. Whereas, PPD can ERα-independently suppress TNFα expression in peritoneal macrophages upon LPS induction. Estrogen deficiency induced inflammatory phenotypes in perivascular adipose tissue (PAT), which could be partially attenuated by PPD. The increased release of adiponectin in PAT after PPD treatment is in accordance with previous reported data showing that adiponectin exerts anti-inflammatory effects in multiple cell types. ERα-dependent antiadipogenic effects of PPD were also detected in PAT-derived stromal cells. The present study reveals a novel mechanism through which PPD exerts estrogenic and anti-inflammatory properties in atherosclerosis-prone mice. Thus, PPD is a promising compound which has potential therapeutic effects on atherosclerotic cardiovascular diseases in postmenopausal women.

Atherosclerosis can cause severe cardiovascular diseases, which is the most common cause of death in the world. It's of great significance to study the prevention and treatment of atherosclerosis. Selenium nanoparticles (SeNPs) has drawn more and more attention due to high biological activity, high bioavailability, strong antioxidant capacity and low toxicity, exhibiting great potential in biomedical application. Thus, this study aimed at explore the anti-atherosclerotic effect of two kinds of SeNPs, bovine serum albumin (BSA) surface-decorated SeNPs and chitosan (CS) surface-decorated SeNPs (CS-SeNPs), in apolipoprotein E deficient (ApoE-/-) mice fed with a high-cholesterol and high-fat diet, and the possible mechanisms. The results demonstrated that both BSA-SeNPs (25, 50 and 100 μg Se/kg body weight/day) and CS-SeNPs (50 μg Se/kg body weight/day) could reduce atherosclerotic lesions in ApoE-/- mice after oral administration for 12 weeks. And these effects might mainly attributed to the ability of BSA-SeNPs and CS-SeNPs to inhibit hyperlipidemia by suppressing hepatic cholesterol and fatty acid metabolism, and alleviate oxidative stress by enhancing antioxidant activity. Moreover, the benefits of BSA-SeNPs were dose-dependent and the medium dose of BSA-SeNPs (50 μg Se/kg body weight/day) was optimal. Generally, BSA-SeNPs with mean size 38.5 nm and negative surface charge showed better anti-atherosclerotic effect than CS-SeNPs with mean size 65.8 nm and positive surface charge. These results suggested that SeNPs could significantly alleviate the formation of atherosclerosis in ApoE-/- mice, possibly by inhibiting hyperlipidemia and oxidative stress, exhibiting a potential to serve as an anti-atherosclerotic agent.

In the present study, we investigated the vasorelaxant effects of alogliptin, an oral antidiabetic drug in the dipeptidyl peptidase-4 (DPP-4) inhibitor class, using phenylephrine (Phe)-induced pre-contracted aortic rings. Alogliptin induced vasorelaxation in a dose-dependent manner. Pre-treatment with the voltage-dependent K+ (Kv) channel inhibitor 4-aminopyridine (4-AP) significantly decreased the vasorelaxant effect of alogliptin, whereas pre-treatment with the inwardly rectifying K+ (Kir) channel inhibitor Ba2+, ATP-sensitive K+ (KATP) channel inhibitor glibenclamide, and large-conductance Ca2+-activated K+ (BKCa) channel inhibitor paxilline did not alter the effects of alogliptin. Although pre-treatment with the Ca2+ channel inhibitor nifedipine did not affect the vasorelaxant effect of alogliptin, pre-treatment with the sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) pump inhibitors thapsigargin and cyclopiazonic acid effectively attenuated the vasorelaxant response of alogliptin. Neither cGMP/protein kinase G (PKG)-related signaling pathway inhibitors (guanylyl cyclase inhibitor ODQ and PKG inhibitor KT 5823) nor cAMP/protein kinase A (PKA)-related signaling pathway inhibitors (adenylyl cyclase inhibitor SQ 22536 and PKA inhibitor KT 5720) reduced the vasorelaxant effect of alogliptin. Similarly, the vasorelaxant effect of alogliptin was not changed by endothelium removal or pre-treatment with the nitric oxide (NO) synthase inhibitor L-NAME or the small- and intermediate-conductance Ca2+-activated K+ (SKCa and IKCa) channel inhibitors apamin and TRAM-34. Based on these results, we suggest that alogliptin induced vasorelaxation in rabbit aortic smooth muscle by activating Kv channels and the SERCA pump independent of other K+ channels, cGMP/PKG-related or cAMP/PKA-related signaling pathways, and the endothelium.

Atherosclerosis is a multifactorial chronic disease, initiated by an endothelial dysfunction. Adenosine and its analogs can change a variety of inflammatory diseases and has shown important effects at different disease models. Inosine is a stable analogous of adenosine, but its effects in inflammatory diseases, like atherosclerosis, have not yet been studied. The aim of this study was to evaluate the pharmacological properties of inosine, administered sub chronically in a hypercholesterolemic model. Male Wistar rats were divided into four groups: control group (C) and control + inosine (C + INO) received standard chow, hypercholesterolemic diet group (HCD) and HCD + inosine (HCD + INO) were fed a hypercholesterolemic diet. At 31st experimentation day, the treatment with inosine was performed for C + INO and HCD + INO groups once daily in the last 15 days. We observed that the hypercholesterolemic diet promoted an increase in lipid levels and inflammatory cytokines production, while inosine treatment strongly decreased these effects. Additionally, HCD group presented a decrease in maximum relaxation acetylcholine induced and an increase in contractile response phenylephrine induced when compared to the control group, as well as it has presented an enhancement in collagen and ADP-induced platelet aggregation. On the other hand, inosine treatment promoted a decrease in contractile response to phenylephrine, evoked an improvement in endothelium-dependent vasorelaxant response and presented antiplatelet properties. Moreover, inosine activated eNOS and reduced p38 MAPK/NF-κB pathway in aortic tissues. Taken together, the present results indicate inosine as a potential drug for the treatment of cardiovascular disorders such as atherosclerosis.

Alamandine attenuates hypertension and cardiac remodeling in spontaneously hypertensive rats (SHRs). We examined whether alamandine attenuates vascular remodeling in mice, and regulates angiotensin II (Ang II)-induced fibrosis in rat vascular smooth muscle cells (VSMCs). Alamandine attenuated hypertension in mice induced by Ang II. Ang II increased the fibrosis of thoracic aorta in mice, which was attenuated by alamandine treatment. Increased levels of collagen I, transforming growth factor-β (TGF-β), and connective tissue growth factor (CTGF) levels in thoracic aortas after Ang II treatment in mice were inhibited by alamandine. Ang II-stimulated collagen I, TGF-β, and CTGF level increases were inhibited by alamandine in rat VSMCs. This could be reversed by Mas-related G protein-coupled receptor, member D (MrgD) antagonist D-Pro7-Ang-(1-7) but not Mas receptor antagonist A779. MrgD expression was increased in the thoracic aortas of mice or VSMCs treatment with Ang II. Ang II increased p-p38 and cAMP levels in rat VSMCs, and alamandine blocked Ang II-induced these increases. Cyclic adenosine monophosphate (cAMP) reversed the inhibitory effects of alamandine on the Ang II-induced increases in collagen I, TGF-β, and CTGF levels. These results demonstrate alamandine attenuates vascular fibrosis by stimulating MrgD expression and decreases arterial fibrosis by blocking p-p38 expression. Alamandine/MrgD axis is a potential target for the treatment of vascular remodeling.

Morin (3,5,7,2′,4′-pentahydroxyflavone) is a yellow coloured natural flavonoid found in plants of the Moraceae family. This favonoid is easily sources from readily available fruits, vegetables and eve certain beverages. Among the sources that was identified, it is clear that morin is most abundantly found in almond, old fustic, Indian guava, and Osage orange. Multiple studies have suggested that morin has multiple therapeutic actions and possess potential to be a functional potent drug. Previous studies demonstrated that morin is capable of resolving deoxycorticosterone acetate-salt-induced hypertension and possess strong vasorelaxant properties. However, the exact mechanisms remains unknown. Therefore, this study is designed to investigate the in vitro mechanism of morin-induced vasorelaxant effects. The underlying mechanisms of morin's vasorelaxant activities were evaluated on thoracic aortic rings isolated from Sprague-Dawley rats. Results from the study demonstrated morin causing vasodilatory reaction in phenylephrine and potassium chloride pre-contracted endothelium-intact aortic rings with the effect being significantly affected in endothelium-denuded aortic rings. Pre-incubation of the aortic rings with ODQ (selective cGMP-independent sGC inhibitor), indomethacin (nonselective COX inhibitor), L-NAME (endothelial nitric oxide inhibitor), propranolol (β2-adrenegic receptors blocker), and atropine (muscarinic receptors blocker) significantly reduced the vasorelaxant effect of morin. It was also found to be able to reduce the intracellular calcium level by blocking VOCC and calcium intake from the extracellular environment and the intracellular release of calcium from the sarcoplasmic reticulum. The present study showed that the vasorelaxant effect of morin potentially involves the NO/sGC, muscarinic receptors, β2-adrenegic receptors, and calcium channels.

The bile acids (BAs) and their conjugates have vascular activities and the serum levels of BAs and their conjugates are increased in liver diseases. In the present study, we examined the in vitro vasoactivities of BAs conjugates taurochenodeoxycholate (TCDC) (5-80 µM), glycochenodeoxycholate (GCDC) (20-150 µM) and tauroursodeoxycholate (TUDC) (20-150 µM) in rat mesenteric arteries and thoracic aorta. The isometric tension of rat mesenteric arteries and thoracic aorta was recorded by using multi-wire myograph systems. TCDC induced significant concentration-dependent relaxation in endothelium-intact but not endothelium-denuded rat mesenteric arteries pre-contracted with phenylephrine (PE). TCDC also showed vasorelaxant effects on high K(+) induced contraction in rat mesenteric arteries. L-NAME treatment inhibited TCDC-induced relaxation in mesenteric arteries pre-contracted with PE. Acute treatment with TCDC increased protein expression of P-eNOS (ser1177) in human umbilical vein endothelial cells. GCDC dose-dependently relaxed PE-induced vasoconstriction in both endotheium-intact and endothelium-denuded rat mesenteric arteries, but GCDC showed no effect on high K(+)-induced vasoconstriction. Both GCDC and TCDC showed no apparent relaxation on PE and high K(+)-induced vasoconstriction in rat thoracic aorta. TUDC showed no effect on PE and high K(+)-induced vasoconstriction in rat mesenteric arteries and thoracic aorta. The study demonstrates that TCDC relaxes rat mesenteric arteries through activating eNOS. TCDC might be the major BAs conjugate for vasorelaxation in vivo.

Ezetimibe is as an inhibitor of NPC1L1 protein, which has a key role in cholesterol absorption. The aim of this study was to evaluate the influence of ezetimibe on the plasma lipid profile, atherosclerotic lesions, and cardiomyocyte ultrastructure in an animal model of atherosclerosis with intermittent hypoxia. Apolipoprotein E-knockout mice received a high-fat diet for 30 days. Then animals were exposed to intermittent hypoxia for 10 days or were maintained under normoxic conditions. In the ezetimibe group, ezetimibe (5 mg/kg/day) was added to the diet. Under normoxic conditions, the total cholesterol level was significantly lower in the ezetimibe group (63.6±6.6 mg/dl) than in the control group (116.3±16.9 mg/dl, P

New preventive strategies for atherosclerosis are needed. In this study, we tested whether a new therapeutic approach consisting of low-dose treatment with a statin and sartan combination could prevent atherogenic diet-induced impairment of the arterial wall in guinea pigs. Twenty-five Dunkin-Hartley guinea pigs were randomly assigned to five experimental groups: 1) normal diet; 2) atherogenic diet (AD); 3) AD + a low-dose atorvastatin and valsartan combination (5mg/kg/day and 2.4mg/kg/day, respectively); 4) AD + low-dose atorvastatin (5mg/kg/day); 5) AD + low-dose valsartan (2.4mg/kg/day). After 8 weeks of treatment, the animals were killed, blood samples collected and thoracic and abdominal aortas isolated. The atherogenic diet significantly impaired maximal thoracic aorta endothelium-dependent relaxation by 40.1% relative to the normal diet. The low-dose combination, compared to the separate drugs, completely preserved thoracic aorta endothelium-dependent relaxation at the level of the group receiving normal diet. This substantial effect was associated with a significant change in the expression of NOS3 (R=0.93; P=0.0002) and IL1b (R=-0.79; P=0.003) genes. In addition, treatment with the low-dose combination or the separate drugs also prevented atherosclerotic plaque formation. We found that treatment with the low-dose atorvastatin and valsartan combination has the capability to completely protect the arterial wall from atherogenic diet-induced damage in the guinea pig model. Further studies evaluating this new therapeutic approach are desirable.

This study investigates the possible protective effects of levocetirizine against fructose-induced insulin resistance, hepatic steatosis and vascular dysfunction, in comparison to pioglitazone, a standard insulin sensitizer. Male Sprague Dawley rats (150-200 g) were divided into 4 groups. Three groups were fed on high fructose diets (HFD) containing 60% w/w fructose, while the fourth control group was fed on standard laboratory food for 8 weeks. AUCOGTT, AUCITT, fasting glucose, HOMA-IR, hepatic glutathione (GSH) and malondialdehyde (MDA) levels, serum total cholesterol, LDL-C, C-reactive protein (CRP) level and lactate dehydrogenase (LDH) activity and liver steatosis scores were significantly higher in HFD group compared to control group. Moreover, body weight gain, food intake, feeding efficiency, HOMA-β, Emax and pEC50 of acetylcholine-induced relaxations of aortic rings and hepatic superoxide dismutase (SOD) activity were significantly lower in HFD group than in control group. Treatment with levocetirizine caused significant decreases in AUCOGTT, AUCITT, HOMA-IR, hepatic GSH and MDA levels and serum CRP level and LDH activity and significant increases in hepatic SOD activity and HOMA-β when compared with the HFD group. Although levocetirizine failed to alter TC and LDL-C levels, it produced a significant increase in HDL-C level relative to control group. Levocetirizine was also able to improve acetylcholine-induced relaxations of aortic rings, indicating a protective effect against insulin resistance-induced endothelial damage comparable to that offered by pioglitazone. Moreover, levocetirizine substantially attenuated insulin resistance-associated liver macrovesicular steatosis. These findings demonstrate that levocetirizine ameliorates insulin resistance, improves glucose tolerance and attenuates insulin resistance-linked hepatic steatosis and vascular damage.

The aim of present work was to investigate the underlying mechanism of vasculature-protecting effects of exercise training in aged rats. Experiment 1: aged rats were given moderate-intensity exercise for 12 weeks. Exercise training suppressed advanced glycation evidenced by reduced activity of aldose reductase, increased activity of glyoxalase 1, reduced levels of methylglyoxal and N(ε)-(carboxymethyl) lysine, and decreased expression of receptor for advanced glycation end products (RAGE) in aged aortas. Experiment 2: aged rats were given moderate-intensity exercise for 12 weeks or treated with FPS-ZM1, an inhibitor of RAGE. Exercise training attenuated aortic stiffening with age marked by reduced collagen levels, increased elastin levels and reduced pulse wave velocity (PWV), and prevented aging-related endothelial dysfunction marked by restored endothelium-mediated vascular relaxation of aortas in response to acetylcholine. Exercise training in aging aortas reduced formation of malondialdehyde, 3-nitrotyrosin and reactive oxygen species, increased GSH/GSSG ratio, suppressed activation of NFκB, and reduced levels of IL-6 and chemokine (C-C motif) ligand 2. Similar effects were demonstrated in aged rats treated with FPS-ZM1. Collectively, exercise suppressed advanced glycation in the aortas of aged rats, which, at least in part, explained the vasculature-protecting effects of exercise training in aged population.

Taste-sensing type 2 receptors (TAS2Rs) have been implicated in extraoral functions. Airway smooth muscle expresses TAS2Rs and is strongly relaxed by TAS2R agonists. We hypothesised that TAS2R agonists might affect vascular smooth muscle as well. Moreover, the general pharmacological profile of TAS2R agonists, which are used to investigate the functions of TAS2R׳s, are undefined. The aim of this study was to pharmacologically characterise the effects of five prototype TAS2R agonists in vascular smooth muscle. Responses to the TAS2R agonists were investigated in guinea-pig aorta and taenia coli, mouse aorta (wild-type and caveolin-1(-/-) mice) and human pulmonary arteries. Chloroquine, denatonium, dextromethorphan, noscapine and quinine, agonists for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, induced strong endothelium-independent relaxations (responses between 82-96% of maximal relaxations) in phenylephrine pre-contracted guinea-pig aorta that persisted in the presence of L-type Ca(2+) and KCa1.1-channel blockers. Experiments in guinea-pig taenia coli revealed that denatonium and quinine also inhibited relaxations to phenylephrine, indicating antagonism of α-adrenoceptors. Only chloroquine and noscapine mediated relaxations when the guinea pig aorta was pre-contracted by U-46619 or PGF2α. Relaxations to chloroquine and noscapine after U-46619 pre-contractions were however markedly impaired in aortae from caveolin-1(-/-) mice. Chloroquine and noscapine mediated relaxations of human pulmonary arteries that expressed also mRNA for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, at levels similar to that of the α1A adrenoceptor. Notwithstanding whether TAS2Rs are involved or not, TAS2R agonists have profound effects on vascular smooth muscle. Chloroquine and noscapine are of special interest as their effects cannot be accounted for by conventional pathways. (Less)

Vascular calcification increases the risk of morbidity and mortality in patients with cardiovascular diseases, chronic kidney diseases, and diabetes. However, viable therapeutic methods to target vascular calcification are limited. Aloe-emodin (AE), an anthraquinone is a natural compound found in the leaves of Aloe-vera. In this study, we investigated the underlying mechanism of AE in the calcification of vascular smooth muscle cells (VSMCs) and murine thoracic aorta. We demonstrate that AE repressed not only the phenotypes of Ca2+ induced calcification but also level of calcium in VSMCs. AE has no effect on cell viability in VSMC cells. Alizarin red, von Kossa stainings and calcium quantification showed that Ca2+ induced vascular calcification is significantly decreased by AE in a concentration-dependent manner. In contrast, AE attenuated Ca2+ induced calcification through inhibiting osteoblast differentiation genes such as SMAD4, collagen 1α, osteopontin (OPN), Runt-related transcription factor (RUNX-2) and Osterix. AE also suppressed Ca2+ induced osteoblast-related protein expression including collagen 1α, bone morphogenic protein 2 (BMP-2), RUNX-2 and smooth muscle actin (SMA). Furthermore, Alizarin red, von Kossa stainings and calcium quantification showed that AE significantly inhibited the calcification of ex vivo ring formation in murine thoracic aorta, and markedly inhibited vitamin D3 induced medial aorta calcification in vivo. Taken together, our findings suggest that AE may have therapeutic potential for the prevention of vascular calcification program.

Mepivacaine is an aminoamide local anesthetic that produces vasoconstriction in vivo and in vitro . The goals of this in vitro study were to determine whether mepivacaine-induced contraction involves calcium sensitization in isolated endothelium-denuded aortas, and to investigate the specific protein kinases involved. The effects of mepivacaine and potassium chloride on intracellular calcium concentrations ([Ca 2+ ] i ) and tension in the presence or absence of Y-27632 or GF 109203X were measured simultaneously using the acetoxymethyl ester of fura-2-loaded aortic strips. Cumulative mepivacaine concentration–response curves were generated in the presence or absence of the following inhibitors: Rho kinase inhibitor Y-27632, protein kinase C (PKC) inhibitor GF 109203X, extracellular signal-regulated kinase (ERK) inhibitor PD 98059, c- Jun NH 2 -terminal kinase (JNK) inhibitor SP600125, and p38 mitogen-activated protein kinase (MAPK) inhibitor SB 203580. Phosphorylation of PKC and MAPK, and membrane translocation of Rho kinase were detected in vascular smooth muscle cells by Western blotting. The slope of the mepivacaine-induced [Ca 2+ ] i -tension curve was higher than that of the KCl-induced [Ca 2+ ] i -tension curve. Pretreatment with Y-27632 or GF 109203X shifted the mepivacaine-induced [Ca 2+ ] i -tension curve to the lower right. Pretreatment with Y-27632, GF 109203X, PD 98059, or SP600125 attenuated mepivacaine-induced contraction in a concentration-dependent manner. Y-27632 and GF 109203X attenuated mepivacaine-induced Rho kinase membrane translocation and PKC phosphorylation, respectively. PD 98059 and SP600125 attenuated mepivacaine-induced ERK and JNK phosphorylation, respectively. Taken together, these results indicate that mepivacaine-induced contraction involves increased calcium sensitization mediated by Rho kinase and PKC. Such contraction mainly involves activation of ERK- and JNK-mediated pathways.

Little is known of the interactions between diabetes and sex hormones on vascular function. The objectives of this study were to investigate whether there were sex differences in rat aortic endothelial function one week after the induction of streptozotocin (STZ)-diabetes, and to examine the potential roles of superoxide and nitric oxide (NO) in this sex-specific effect. Endothelium-dependent vasodilatation to acetylcholine (ACh) was measured in rat aortic rings before and after treatment with MnTMPyP (25µM), a superoxide dismutase. Contractile responses to phenylephrine (PE) were generated before and after treatment with l-NAME (200μM), a nitric oxide synthase (NOS) inhibitor. The mRNA expression of NADPH oxidase (Nox) and endothelial nitric oxide synthase (eNOS) were also determined. We demonstrated that (1) STZ-diabetes impaired endothelium-dependent vasodilatation to ACh to a greater extent in female than male aortae, (2) inhibition of superoxide enhanced sensitivity to ACh only in diabetic females, and (3) Nox1 and Nox4 mRNA expression were significantly elevated only in aortic tissue of diabetic females. Furthermore, incubation of aortic rings with l-NAME potentiated PE responses in all groups, but aortae from control females showed a greater potentiation of the PE response after NOS inhibition compared with others. STZ-diabetes reduced the extent of PE potentiation after l-NAME and the aortic eNOS mRNA expression in females to the same levels as seen in males. These data suggest that a decrease in NO, resulting from either decreased eNOS or elevated superoxide, may partially contribute to the predisposition of the female aorta to injury early in diabetes.

Circulating levels of anandamide are increased in diabetes, and cannabidiol ameliorates a number of pathologies associated with diabetes. The aim of the present study was to examine how exposure to anandamide or cannabidiol might affect endothelial dysfunction associated with Zucker Diabetic Fatty rats. Age-matched Zucker Diabetic Fatty and Zucker lean rats were killed by cervical dislocation and their arteries mounted on a myograph at 37 °C. Arteries were incubated for 2h with anandamide, cannabidiol or vehicle, contracted, and cumulative concentration-response curves to acetylcholine were constructed. Anandamide (10 µM, 2h) significantly improved the vasorelaxant responses to acetylcholine in aortae and femoral arteries from Zucker Diabetic Fatty rats but not Zucker lean rats. By contrast, anandamide (1 µM, 2h) significantly blunted acetylcholine-induced vasorelaxation in third-order mesenteric arteries (G3) from Zucker Diabetic Fatty rats. Cannabidiol incubation (10 µM, 2h) improved acetylcholine responses in the arteries of Zucker Diabetic Fatty rats (aorta and femoral) and Zucker lean (aorta, femoral and G3 mesenteric), and this effect was greater in the Zucker Diabetic Fatty rat. These studies suggest that increased circulating endocannabinoids may alter vascular function both positively and negatively in type 2 diabetes, and that part of the beneficial effect of cannabidiol in diabetes may be due to improved endothelium-dependent vasorelaxation.

The development of hepatocellular carcinomas from malignant hepatocytes is frequently associated with intra- and peritumoral accumulation of connective tissue arising from activated hepatic stellate cells (HSC). Inhibition of receptor tyrosine kinase (RTK) signaling showed promise in the treatment of hepatocellular carcinoma. However, there is a lack of knowledge about the effects of RTK inhibitors on the tumor supportive cells. We performed in vitro experiments to study whether Sunitinib, a platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) RTKs' inhibitor, could block both activated HSC functions and angiogenesis and thus prevent the progression of cirrhotic liver to hepatocellular carcinoma. In immortalized human activated HSC LX-2, treatment with Sunitinib 100 nM blocked collagen synthesis by 47%, as assessed by Sirius Red staining, attenuated HSC contraction by 65%, and reduced cell migration by 28% as evaluated using a Boyden's chamber, without affecting cell viability, measured by Trypan blue staining, and apoptosis, measured by propidium iodide (PI) incorporation assay. Our data revealed that Sunitinib treatment blocked the transdifferentiation of primary human HSC (hHSC) to activated myofibroblast-like cells by 65% without affecting hHSC apoptosis and migration. In in vitro angiogenic assays, Sunitinib 100 nM reduced endothelial cells (EC) ring formation by 46% and tube formation by 68%, and decreased vascular sprouting in aorta ring assay and angiogenesis in vascular bed of chick embryo. In conclusion, the present study demonstrates that the RTK inhibitor Sunitinib blocks the activation of HSC and angiogenesis suggesting its potential as a drug candidate in pathological conditions like liver fibrosis and hepatocellular carcinoma.

1-Nitro-2-phenylethene (NPe) induces a more potent vasorelaxant effect in rat aorta than its structural analog 1-nitro-2-phenylethane, but mediated through a different mechanism, independent of soluble guanylate cyclase (sGC) stimulation. We hypothesized that introducing an electron donor into the aromatic moiety might stabilize NPe, enhancing its potency and/or interaction with sGC. Therefore, trans-4-methoxy-β-nitrostyrene (T4MN) was synthesized, and mechanisms underlying its vasorelaxant effects were studied in rat aortic ring preparations. In endothelium-intact preparations, T4MN fully relaxed contractions induced by phenylephrine (PHE) with a potency similar to that of its parent drug, NPe. This vasorelaxant effect that was unchanged by endothelium removal, pretreatment with L-NAME, indomethacin, or MDL-12,330 A, but was significantly reduced by tetraethylammonium, 4-aminopyridine, methyl blue, or ODQ. Under Ca 2+ -free conditions, T4MN did not alter contractions evoked by caffeine, but significantly reduced, in an ODQ-preventable manner, those induced by either PHE or extracellular Ca 2+ restoration following depletion of intracellular Ca 2+ stores in thapsigargin-treated aortic preparations. Under the same conditions, T4MN also reduced contractions induced by protein kinase C activator phorbol-12,13-dibutyrate with a potency similar to that evoked by this nitroderivative against PHE-induced contractions. In conclusion, T4MN induces potent vasorelaxation in rat aorta by stimulating the sGC-cGMP pathway through a NO-independent mechanism. Introduction of a methoxy group into the aromatic moiety apparently stabilizes NPe, thereby enhancing its interaction with sGC.

Many studies have shown that anti-inflammatory agents are effective in the treatment of endometriosis. β-Caryophyllene exerted a potent anti-inflammatory effect in vivo. However, its effect on endometriosis has not been investigated. This study aims at investigating the effect of β-caryophyllene on endometriosis and on fertility and reproduction in adult female rats. Autologous fragments of the endometrium were implantated in the peritoneal cavity in adult female rats. The growth of the endometriotic implants that developed after four weeks was recorded. Treatment started then with β-caryophyllene (10 mg/kg or 30 mg/kg) or vehicle (control) for 21 days and the growth of the endometriotic implants was measured again. In fertility studies, female rats that received β-caryophyllene or vehicle were mated and reproductive functions were observed including number and viability of implants, number of corpora lutea, length of pregnancy and outcome of litter. β-Caryophyllene (10 mg/kg) suppressed the growth of endometriotic implants by 52.5% compared with controls. Also β-caryophyllene produced apoptosis in luminal epithelim of the cyst as well as in endothelial cells of blood vessels. Ultrstructural studies revealed the presence of active mast cells and eosinophils in both control and β-caryophyllene-treated rat cysts. No statistically significant difference was observed in any studied parameter between control and β-caryophyllene-treated groups in fertility study. Therapy with β-caryophyllene may present a promising novel, non-toxic therapeutic option for patients with endometriosis.

As a Ca2+-permeable cationic channel, canonical transient receptor potential 5 channel (TRPC5) has been known to be involved in various functions of cells. Although TRPC5 is abundantly expressed in vascular endothelium, it remains unknown whether TRPC5 plays a role in regulating endothelial senescence. In the present study, we investigated the involvement of TRPC5 in the senescence of mouse aortic endothelial cells (MAECs). Meanwhile, the regulatory role of TRPC5 in the endothelial dysfunction in aged mice was also studied. Results showed that the endothelium-dependent relaxations were significantly promoted but the endothelium-dependent contractions were markedly attenuated in aortic rings from aged mice with TRPC5 gene knocked out compared with those from aged C57BL/6J mice. Meanwhile, the nuclear telomerase activity and nitric oxide generation were notably enhanced but the reactive oxygen species production was significantly inhibited in aged mice or in hydrogen peroxide-induced senescent MAECs lacking the TRPC5 gene. In addition, by performing the senescence-associated β-galactosidase staining the oxidative stress-induced premature senescence was markedly depressed in MAECs with TRPC5 gene deficient. Expressions of endothelial nitric oxide synthase and silent information regulator protein 1 (SIRT1) were significantly up-regulated but those of P53 and P21 were markedly down-regulated both in aged mice and in hydrogen peroxide-treated MAECs in the absence of the TRPC5 gene. These results of our study uncover novel functions of TRPC5 in regulating vascular aging.

This study examined whether coenzyme Q

Esmolol produces early regression of left ventricular hypertrophy and improves coronary artery remodeling, although the impact of short-term treatment with this beta-blocker on remodeling in large arteries has not yet been studied. We hypothesized that even a short (48 h) course of esmolol might alter remodeling of the aorta in the spontaneously hypertensive rat (SHR). Fourteen-month-old male SHRs were treated intravenously with vehicle (SHR, n=8) or esmolol (SHR-E, n=8) (300 μg/kg/min). Age-matched, vehicle-treated male Wistar-Kyoto rats (WKY, n=8) served as controls. After 48 h, we studied the structure, volume density of elastic fibers, and passive mechanical properties of the aorta. Determination of asymmetrical dimethylarginine concentrations and total protein carbonyls in the aorta were analyzed. Esmolol significantly attenuated abnormal aortic wall thickness, cross-sectional area, wall-to-lumen ratio, volume density of elastic fibers, and wall stiffness. The protective effect of esmolol could be related to a decrease in asymmetrical dimethylarginine levels after down-regulation by oxidative stress. These findings could play a key role in the selection of antihypertensive therapy in patients with hypertension and aortic remodeling.

Experiments were designed to determine if Wy14643 ([[4-chloro-6-[(2,3-dimethylphenyl)amino]-2-pyrimidinyl]thio]-acetic acid), a preferential agonist at peroxisome proliferator-activated receptors (PPAR) α , improves vascular function in hypertension, and if so, the mechanism(s) involved. Isometric tension was measured in isolated thoracic aorta of spontaneously hypertensive rats (SHR). Wy14643-induced relaxations in SHR aortic rings were greater than those induced by fenofibrate or rosiglitazone (PPARα or PPAR γ agonists, respectively) and were larger in rings with endothelium than those without. Both MK886 [(1-[(4-chlorophenyl)methyl]-3-1,1-dimethylethyl)thio]-( α , α -dimethyl-5-1-methylethyl)-1H-indole-2-propanoic acid (PPAR α antagonist) and GW9662 (2-chloro-5-nitrobenzanilide) (PPAR γ antagonists) inhibited Wy14643-induced relaxations. The inhibitory effect of MK886 was more pronounced in rings with endothelium than those without. In SHR aortic rings with endothelium, L-NAME ( N ω -nitro-L-arginine methyl ester, nitric oxide synthase inhibitor), ODQ (1H-1,2,4)oxadiazolo[4,3-a]quinoxalin-1-one, soluble guanylyl cyclase inhibitor) and compound C [adenosine monophosphate-activated protein kinase (AMPK) inhibitor] reduced Wy14643-induced relaxations. Western blotting studies indicated that Wy14643 and fenofibrate, but not rosiglitazone, increased the phosphorylation of endothelial nitric oxide synthase and AMPK; these effects were abolished by compound C but not L-NAME. Endothelium-dependent contractions evoked by acetylcholine in quiescent SHR aorta in the presence of L-NAME were reduced by Wy14643 and fenofibrate but not by rosiglitazone. MK886, but not GW9662, prevented this effect. Wy14643 and fenofibrate inhibited acetylcholine-induced prostanoid release to the same extent. These findings suggest that PPARα agonists induce nitric oxide-mediated relaxation through activation of AMPK and reduce the release of endothelium-dependent contracting factors. Because also of the ability to activate smooth muscle PPARγ to induce relaxation, Wy14643 offers additional protection against vascular dysfunction of spontaneous hypertension.

Bioavailability of nitric oxide (NO) is mostly studied in isolated blood vessels. We investigated changes in basal and receptor-stimulated endothelial NO synthase (eNOS) activity after isolation of wild-type and Marfan mouse aorta. Starting 1h after dissection, basal NO release was assessed at hourly intervals by its ability to suppress isometric contractions in aortic segments. Relaxation induced by acetylcholine or α(2)-adrenoceptor agonist 5-bromo-N-(4,5-dihydro-1H-imidazol-2-yl)-6-quinoxalinamine (UK14304) was used to study stimulated NOS activity. One hour after dissection, phenylephrine- or prostaglandin F(2α)-induced force attained only 17 ± 4% or 31 ± 7% of maximum tension in the presence of N(Ω)-nitro-l-arginine-methylesther (l-NAME), and contractions increased to 63 ± 6% and 82 ± 11%, respectively at 5h. In contrast, acetylcholine or UK14304 relaxation curves changed minimally. l-NAME and eNOS-deficiency abolished basal NO production, unlike inhibitors of neuronal (N(Ω)-propyl-l-arginine) or inducible (1400W) NOS. Acetylcholine-induced relaxation was abolished by l-NAME, strongly suppressed by eNOS-deficiency and attenuated by N(Ω)-propyl-l-arginine. In a bioassay based on diethylamine NONOate concentration-response curves the suppression of contractile forces was interpolated into NO equivalents. This showed exponential decay of basal NO, which occurred three times faster in aortas from mice with Marfan syndrome, while acetylcholine-induced relaxation remained unaltered. Immunoblotting showed unchanged eNOS expression, or phosphorylation at Ser1177, Ser617 or Thr495 between 1h and 4h, but Akt phosphorylation declined gradually. The dramatic loss of basal NO release after tissue isolation shows that timing is crucial when studying NO responses. The preservation of receptor-induced relaxation implies differential regulation of basal and stimulated eNOS activity, and phosphoinositide-3-kinase/Akt signalling seems specifically associated with basal eNOS activity.

Omentin is a recently identified adipocytokine and its effect in vasculature is largely unknown. Here we examined the effects of omentin on smooth muscle cells (SMCs) inflammatory states. Western blotting was performed to analyze inflammatory signal transduction in cultured SMCs. Phosphorylation of nuclear factor-κB (NF-κB), p38 and JNK, and expression of vascular cell adhesion molecule (VCAM)-1 and cyclooxygenase-2 were not induced by omentin (50-300ng/ml, 20min or 24h). On the other hand, tumor necrosis factor-α (TNF-α; 10ng/ml, 20min)-induced phosphorylation of p38 and JNK was significantly inhibited by omentin pretreatment in a concentration-dependent manner (50-300ng/ml, 30min). TNF-α (24h)-induced expression of VCAM-1 was also significantly inhibited by omentin pretreatment in a concentration-dependent manner. Both inhibitor of p38 (SB203580) and JNK (SP600125) significantly inhibited TNF-α-induced VCAM-1 expression. Omentin (300ng/ml, 30min) inhibited TNF-α (1h)-induced nicotinamide adenine dinucleotide phosphate oxidase activity as determined by lucigenin assay. An antioxidant drug, N-acetyl-l-cysteine significantly inhibited TNF-α-induced phosphorylation of p38 and JNK. Furthermore, omentin (300ng/ml, 30min) significantly inhibited TNF-α (24h)-induced monocytic cells adhesion to SMCs. In rat isolated thoracic aorta, omentin (300ng/ml, 30min) inhibited TNF-α (24h)-induced VCAM-1 expression. The present results demonstrate for the first time that omentin plays an anti-inflammatory role by preventing the TNF-α-induced VCAM-1 expression in SMCs. It is suggested that omentin inhibits TNF-α-induced VCAM-1 expression via preventing the activation of p38 and JNK at least in part through inhibition of superoxide production.

In the present study, we investigated the vasculoprotective effect of sigma-1 receptor stimulation with fluvoxamine on pressure overload hypertrophy-induced vascular injury in the thoracic aorta and defined mechanisms underlying that activity. Wistar rats underwent bilateral ovariectomy, and two weeks later were further treated with abdominal aortic stenosis. To confirm the vasculoprotective role of sigma-1 receptor signaling, we treated rats with the agonist fluvoxamine (at 0.5 and 1.0 mg/kg) and with the antagonist NE-100 (at 1.0mg/kg) for 4 weeks orally once a day after the onset of aortic banding. Interestingly, sigma-1 receptor expression in the thoracic aorta decreased significantly 4 weeks after pressure overload-induced hypertrophy in vehicle treated ovariectomized rats. Fluvoxamine administration significantly attenuated pressure overload-induced vascular injury with concomitant increase in receptor expression and subsequent decrease in IP3 receptor expression. Fluvoxamine treatment also significantly restored pressure overload-induced impaired Akt phosphorylation and stimulated eNOS protein expression as well as Akt-mediated eNOS phosphorylation (Ser1177). Fluvoxamine's vasculoprotective effect was nullified by co-administration of a sigma-1 receptor antagonist. No changes in phosphorylation of ERK1/2 or PKCα in the aorta were observed following pressure overload and after fluvoxamine treatment. Our findings confirm, for the first time, a potential role for sigma-1 receptor expression and signaling in the thoracic aorta in attenuating hypertrophy-induced vascular injury in ovariectomized rats. Thus, we demonstrate, for the first time, a potential role in the thoracic aorta for sigma-1 receptor expression and signaling via Akt-eNOS in attenuating hypertrophy-induced vascular injury in ovariectomized rats.

Treatment of diabetes complications remains a substantial challenge. The aim of this study was to explore the ability of the soy isoflavone genistein in attenuating the signs that follow diabetes onset: nociceptive hypersensitivity, oxidative and inflammatory state, nerve growth factor (NGF) decrease and vascular dysfunctions. Genistein (3 and 6 mg/kg) was administered to C57BL/6J streptozotocin diabetic mice from the 2nd till the 5th week after disease induction. The hind paw withdrawal threshold to mechanical stimulation (tactile allodynia) was evaluated by a von Frey filament. The oxidative stress was assessed measuring: reactive oxygen species by fluorimetric analysis, both the lipoperoxide content, as malondialdehyde, the antioxidant enzymatic activities spectrophotometrically and the glutathione content spectrofluorimetrically. Proinflammatory cytokines and NGF were measured in the sciatic nerve by enzyme-linked immunosorbent assay. Aortic inducible (iNOS) and endothelial nitric oxide synthase (eNOS) protein content was measured by western immunoblotting. Genistein relieved diabetic peripheral painful neuropathy, reverted the proinflammatory cytokine and reactive oxygen species overproduction, and restored the NGF content in diabetic sciatic nerve. Furthermore it restored the GSH content and the GSH and GSSG ratio, improved the antioxidant enzymes activities, decreased reactive oxygen species and lipoperoxide level in the brain and liver. Finally it restored the iNOS and eNOS content and the superoxide dismutase activity in thoracic aorta. Hyperglycaemia and weight decrease were not affected. Genistein is able to reverse a diabetes established condition of allodynia, oxidative stress and inflammation, ameliorates NGF content and the vascular dysfunction, thus suggesting its possible therapeutic use for diabetes complications.

Adipose tissue releases an "adipocyte-derived relaxing factor" (ADRF) lowering tone of isolated arteries. The potential influence of hypoxia on the vasorelaxing properties of adipose tissue was investigated. Aortas from male Swiss mice with or without adherent adipose tissue were mounted in a wire myograph for isometric tension recording. Hypoxia (bubbling with 95% N(2), 5% CO(2)) relaxed precontracted (norephinephrine, 5 microM) aorta with adipose tissue while only a minimal vasorelaxing effect was observed in arteries without adipose tissue. This effect was also seen after precontraction with prostaglandin F(2alpha) (30 microM) or U-46619 (10 nM). Precontraction with 60 or 120 mM K(+), incubation with tetraethylammoniumchloride (3 mM) or glibenclamide (30 microM) significantly impaired the hypoxic response. Glibenclamide (30 microM) enhanced the vasorelaxing effect of NaHS (except at high concentrations of NaHS). Lactate (10 nM to 1 mM) had no effect on preparations with or without adipose tissue. 8-(p-sulfophenyl)theophylline (0.1 mM), zinc protoporphyrin IX (10 microM), 1 H-[1, 2, 4]oxadiazolo[4,3-A]quinoxalin-1-one (10 microM) and removal of the endothelium did not influence the hypoxic relaxation. Our findings indicate that hypoxia has a relaxing influence on mice aorta that is dependent on the presence of adherent adipose tissue. This relaxation is partly mediated by opening K(ATP) channels and independent of the endothelium and soluble guanylyl cyclase. Neither lactate, adenosine, CO nor H(2)S seems to be involved in this hypoxic response. However, the involvement of the as yet unidentified "adipocyte-derived relaxing factor" (ADRF) cannot be excluded.

Reactive oxygen species, such as peroxynitrite, induce oxidative stress and DNA injury leading to endothelial dysfunction. It has been proposed, that elevated intracellular cyclic GMP (cGMP)-levels may contribute to an effective cytoprotection against nitro-oxidative stress. We investigated the dose-dependent effects of vardenafil, an inhibitor of phosphodiesterase-5, on endothelial dysfunction induced by peroxynitrite. In organ bath experiments, we investigated the endothelium-dependent (acetylcholine) and -independent (sodium nitroprusside, SNP) vasorelaxation of isolated aortic rings of rats. Endothelial dysfunction was induced by peroxynitrite. In the treatment groups, rats received low doses (0.01-5 microg/kg) or high doses (5-300 microg/kg) of vardenafil. DNA strand breaks were assessed by the TUNEL method. Immunohistochemical analysis was performed for cGMP and nitrotyrosine. Exposure to peroxynitrite resulted in an impairment of endothelium-dependent vasorelaxation of aortic rings. Pre-treatment with lower doses of vardenafil led to an improvement of endothelial function as reflected by the higher maximal vasorelaxation (R(max)) to acetylcholine. Interestingly, at higher doses, R(max) to acetylcholine was attenuated leading to U-shaped dose-response curves. The endothelium-independent vasorelaxation to SNP under peroxynitrite stress showed a significant left-shift of the SNP concentration-response curves in the vardenafil groups without any alterations of the R(max). Vardenafil-pre-treatment significantly reduced DNA-breakage, reduced nitrosative stress, and increased cGMP score in the aortic wall. Our working hypothesis is that improvement of endothelial function could be mainly due to the cytoprotection of endothelium by vardenafil. This work supports the view that acute PDE5-inhibition might be advantageous in the treatment of endothelial dysfunction induced by disturbed NO-cGMP pathway due to nitro-oxidative stress.

Apigenin is shown to have cardiovascular effects, but the effects of apigenin on aortas injured by exogenous oxidants are unknown. The objective of this study was to investigate the effect of apigenin on endothelium-dependent vasorelaxation in isolated rat aortic rings exposed to superoxide anion produced by pyrogallol, and its mechanism. The male Sprague-Dawley rat thoracic aorta was rapidly dissected out and the effect of apigenin on tension of aortic rings pretreated with 500 microM pyrogallol, inducing oxidative stress injury, was measured. The activity of nitric oxide synthase (NOS), the level of nitric oxide (NO) and the inhibition of superoxide anion in aortic tissues were measured. We found that pretreatment with pyrogallol concentration-dependently decreased acetylcholine-induced endothelium-dependent vasorelaxation. Apigenin (0.5-72.0 microM) evoked a concentration-dependent relaxation in aortas (pD(2): 5.304+/-0.049), which was weakened by L-NAME (the maximal relaxation fell from 87.6+/-6.7% to 37.1+/-8.8%, P

In Wistar rats, maternal exposure to nicotine was shown to impair the inhibitory function of perivascular adipose tissue on vascular contractility in the aorta of the offspring. It is not known whether an impairment of perivascular adipose tissue function occurs in smaller arteries, and whether the control of blood pressure is affected. Here we studied the blood pressure effects and the alteration of perivascular adipose tissue function in mesenteric arteries of the offspring born to Wistar-Kyoto rat (WKY) dams exposed to nicotine. Nulliparous female WKY rats were given either nicotine bitartrate (1 mg/kg/day) or saline (vehicle) by subcutaneous injection 2 weeks prior to mating, during pregnancy and until weaning. Blood pressure of the offspring and functional studies with mesenteric arteries were conducted. Tissue samples (thoracic aorta, mesenteric arteries, and kidneys) were collected for morphological and immunohistochemical examinations. Blood pressure increased from 14 weeks of age onwards in the offspring born to nicotine-exposed dams. Nicotine-exposed offspring showed a significant increase in the number of brown adipocytes in aortic perivascular adipose tissue relative to control offspring. In mesenteric arteries from control offspring, contractile responses induced by phenylephrine, serotonin, and 9,11-dideoxy-11alpha, 9alpha-epoxymethanoprostaglandin F(2)alpha (U44619) were significantly attenuated in the presence of perivascular adipose tissue, an effect not observed in the nicotine-exposed tissues. Endothelium-dependent relaxation responses to carbachol, kidney weight, the total number of nephrons and glomerulus' size were comparable in nicotine and saline groups. We conclude that fetal and neonatal exposure to nicotine caused blood pressure elevation. Alterations in perivascular adipose tissue composition and modulatory function are some of the mechanisms associated with this blood pressure increase.

WOS: 000258628400032, PubMed ID: 18538317, The pathophysiology of pre-eclampsia is still unknown thus effective primary prevention is not possible at the stage. The present study was conducted to research the smooth muscle responses in the pre-eclampsia model with suramin treated rats and the effect of phosphodiesterase-5 (PDE5) inhibitor on these responses. Rats of three groups; control, suramin and suramin+sildenafil were given intraperitoneal injections of saline, suramin or sildenafil citrate. Suramin injections caused increased blood pressure, protein in urine and caused fetal growth retardation.The use of sildenafil citrate straightened significantly both blood pressure and average fetus weight, but did not reach to control values. At the end of pregnancy, thoracic aorta rings were exposed to contractile and relaxant agents. KCl contraction responses, sodium nitroprusside and papaverine relaxation responses were similar in three groups. Contraction responses of phenylephrine, increased significantly in suramin group. Relaxation responses of acethylcholine and bradykinin decreased in suramin group. The use of sildenafil citrate partially straightened both relaxation and contraction responses, but did not reach to control values. In all groups in the presence of L-nitromonomethylarginine (L-NAME), 1H-(1, 2, 4) oxadiazole (4, 3-a) guinoxalin-1-one (ODQ) and indomethacin decreased the relaxation responses of acetylcholine and bradykinin. The cyclic guanosine monophosphate (cGMP) content of thoracic aorta tissue was determined by radioimmunoassay technique. The content of cGMP in suramin group decreased and use of sildenafil citrate increased the cGMP content but did not reach to control values. We conclude that in pre-eclampsia, the increase of contraction responses, the decrease of relaxation responses and the decrease of cGMP content can depend on insufficiency about synthesis or release of relaxant factors which was released from the vessel endothelium. The results in this study show that in pre-eclampsia: PDE5 inhibitors enhance endothelial function and may be used for protection. Further studies are needed to clear the efficiency and safety of PDE5 inhibitors. (c) 2008 Elsevier B.V. All rights reserved., Cumhuriyet University Scientific Research [T-282], This study was supported by Cumhuriyet University Scientific Research Project T-282 (CUBAP, Sivas, Turkey).

Although antihypertensive drugs confer improvement in endothelial dysfunction and protection from atherogenesis in hypertension, different classes of antihypertensive drugs may elicit different degrees of vasculoprotective effects. We have investigated the effects of a long-acting calcium antagonist, benidipine, and an angiotensin AT(1) receptor antagonist, losartan, on the vascular damage observed in OLETF rats, an animal model of metabolic syndrome. At 34 weeks of age, OLETF rats were treated with either benidipine (3 mg/kg/day, per os) or losartan (25 mg/kg/day, per os) for 8 weeks. The extent of blood pressure reduction, restoration endothelium-dependent aortic relaxation, and elevation of serum nitrite/nitrate concentration did not differ significantly between benidipine- and losartan-treated OLETF rats. Benidipine and losartan also reduced the aortic expression of transforming growth factor-beta1 mRNA and thickening of the vascular wall to a similar extent. Increased cardiac fibrosis was also inhibited by both benidipine and losartan. These data suggest that, when used in an antihypertensive dose, benidipine is as effective as losartan in restoring vascular endothelial function and in suppressing of cardiovascular remodeling in an animal model of metabolic syndrome.