Responses Of Mesenteric Arteries Research Articles

Heart function in magnetic resonance imaging and the mesenteric artery reactivity in rats receiving lead-contaminated drinking water

The aim of this study was to evaluate the effect of lead (Pb)-contaminated drinking water on magnetic resonance imaging (MRI)-estimated cardiac function, vascular reactivity, and serum lipids in rats. For 3months, male Wistar rats, aged 4-6 weeks, were given drinking water with the addition of lead acetate at a concentration of 100ppm Pb (10 rats) or water free from Pb (8 control rats). The cardiac MRI was performed at rest and under β-adrenergic stimulation on a 4.7T scanner using electrocardiogram-triggered gradient echo (FLASH) cine sequence. After 1-2 weeks of the MRI test, experiments were performed ex vivo. After stabilization of perfusion pressure (PP), norepinephrine at doses from 0.01 to 5.0μg was dissolved in Krebs solution, injected in a volume of 100μl, and next infused at a concentration of 0.5μg/ml into the isolated mesenteric artery. In this manner, preconstricted mesenteric bed was used to determine PP changes induced by acetylcholine, given at doses from 0.05 to 5.0μg, before and during the infusion of nitric oxide synthase inhibitor (1.0μg/ml). At the end, dobutamine (5mg), followed by potassium chloride (10.5mg), was injected. Lipid levels were determined enzymatically, blood Pb level was measured by the atomic absorption spectrophotometer. This study showed that Pb impairs the left ventricular systolic and diastolic function. Pb-induced changes in response to resistance of vessels to vasoactive agents may be secondary to the reduced left ventricular ejection fraction. The high-density lipoprotein subfraction 2 (HDL2) is involved in the cardiovascular effect of Pb.

Desensitization of G-protein-coupled receptors induces vascular hypocontractility in response to norepinephrine in the mesenteric arteries of cirrhotic patients and rats

The increased beta-arrestin-2 and its combination with G-protein-coupled receptors (GPCRs) lead to GPCRs desensitization. The latter may be responsible for decreased contractile reactivity in the mesenteric arteries of cirrhotic patients and rats. The present study is to investigate the machinery changes of alpha-adrenergic receptors and G proteins and their roles in the contractility of mesenteric arteries of cirrhotic patients and animal models. Patients with cirrhosis due to hepatitis B and cirrhotic rats induced by CCl4 were studied. Mesenteric artery contractility in response to norepinephrine was determined by a vessel perfusion system. The contractile effect of G protein-coupled receptor kinase-2 (GRK-2) inhibitor on the mesenteric artery was evaluated. The protein expression of the alpha1 adrenergic receptor, G proteins, beta-arrestin-2, GRK-2 as well as the activity of Rho associated coiled-coil forming protein kinase-1 (ROCK-1) were measured by Western blot. In addition, the interaction of alpha1 adrenergic receptor with beta-arrestin-2 was assessed by co-immunoprecipitation. The portal vein pressure of cirrhotic patients and rats was significantly higher than that of controls. The dose-response curve to norepinephrine in mesenteric arteriole was shifted to the right, and EC50 was significantly increased in cirrhotic patients and rats. There were no significant differences in the expressions of the alpha1 adrenergic receptor and G proteins in the cirrhotic group compared with the controls. However, the protein expressions of GRK-2 and beta-arrestin-2 were significantly elevated in cirrhotic patients and rats compared with those of the controls. The interaction of the alpha1 adrenergic receptor and beta-arrestin-2 was significantly aggravated. This interaction was significantly reversed by GRK-2 inhibitor. Both the protein expression and activity of ROCK-1 were significantly decreased in the mesenteric artery in patients with cirrhosis compared with those of the controls, and this phenomenon was not shown in the cirrhotic rats. Norepinephrine significantly increased the activity of ROCK-1 in normal rats but not in cirrhotic ones. Norepinephrine significantly increased ROCK-1 activity in cirrhotic rats when GRK-2 inhibitor was used. beta-arrestin-2 expression and its interaction with GPCRs are significantly upregulated in the mesenteric arteries in patients and rats with cirrhosis. These upregulations result in GPCR desensitization, G-protein dysfunction and ROCK inhibition. These may explain the decreased contractility of the mesenteric artery in response to vasoconstrictors.

Contribution of Rho kinase to blood pressure elevation and vasoconstrictor responsiveness in type 2 diabetic Goto–Kakizaki rats

The RhoA-Rho kinase (ROCK) pathway contributes to a number of diabetic complications including cardiomyopathy and nephropathy. In this study, we investigated whether it contributes to elevated blood pressure and vascular contractile dysfunction in type 2 diabetes. Blood pressure was measured in Goto-Kakizaki rats, a nonobese model of type 2 diabetes, before and after treatment with the ROCK inhibitor fasudil. Vasoconstrictor responsiveness in the absence and presence of ROCK inhibitors as well as ROCK pathway activity was measured in isolated mesenteric resistance vessels from these animals. Blood pressure was elevated in diabetic rats compared with age-matched Wistar controls, and was normalized by treatment with fasudil. Contractile responses of mesenteric arteries from diabetic rats to phenylephrine and U-46619, as well as relaxant responses to acetylcholine, were unaltered. However, vasoconstrictor responses were more sensitive to ROCK inhibition with either Y-27632 or H-1152 than were responses of control arteries. No differences were found in expression of RhoA, ROCK1, or ROCK2 or in basal ROCK activity between arteries from control and diabetic rats. U-46619 produced a similar magnitude of increase in ROCK activity that was completely blocked by H-1152 in arteries from both groups of animals. These data suggest that ROCK contributes to the increase in blood pressure in type 2 diabetic Goto-Kakizaki rats, and that vasoconstrictor responses of small mesenteric arteries from these animals are more dependent on ROCK than are responses of control arteries.

186 NITRIC OXIDE MEDIATES THE ANTICONTRACTILE EFFECT OF PERIVASCULAR ADIPOSE TISSUE

<h3>Objective</h3> Perivascular adipose tissue (PVAT) exerts an anticontractile effect. Studies using various vasoconstrictor agents suggest that this may be dependent on adrenergic stimulation and the subsequent release of nitric oxide. Moreover, nitric oxide synthase expression has previously been identified in PVAT isolated from human saphenous vein grafts. This study investigated the role of nitric oxide in the anticontractile action of PVAT. <h3>Design and methods</h3> The effect of PVAT on the contractility of isolated rat mesenteric arteries (approx. 250-300 µm internal diameter) was investigated using wire myography. Concentration-response curves to noradrenaline (1x10⁻⁵ - 3x10⁻⁹ mol.l⁻¹) were generated following 30 minute incubation with the nitric oxide synthase inhibitor, L-NMMA (100 µmol.l⁻¹). In addition, a Griess reagent kit was used to determine NO production from the adipose tissue surrounding the mesenteric artery in response to stimulation with the adrenergic agonists (10 µmol.l⁻¹), noradrenaline, phenylephrine or CL-316, 243+/−L-NMMA (100 µmol.l⁻¹). Data were analysed by two-way ANOVA. <h3>Results and conclusions</h3> The vasoconstrictor response to noradrenaline was reduced in the presence of PVAT through an endothelium-dependent mechanism (endo: PVAT vs no PVAT <i>P</i>&lt;0.05, <i>n</i>=5, no endo: PVAT vs no PVAT <i>P</i>=0.965, <i>n</i>=5). The presence of L-NMMA produced an increase in the vasoconstrictor action of noradrenaline only in the presence of <b>both</b> PVAT and endothelium (PVAT: no L-NMMA vs L-NMMA <i>P</i>&lt; 0.001, <i>n</i>= 5, no PVAT: no L-NMMA vs L-NMMA <i>P</i>=0.367, <i>n</i>=5) indicating that nitric oxide release from PVAT contributes to its anticontractile effect. Adrenergic stimulation of adipose tissue (devoid of vessels) with noradrenaline and CL-316,243, but not the α-adrenergic agonist, phenylephrine, was associated with a significant increase in nitric oxide production (noradrenaline and CL-316,243, <i>P</i>&lt;0.05, <i>n</i>=5), which was inhibited by the presence of L-NMMA. This confirms that β-adrenergic activation stimulates nitric oxide production by PVAT which may contribute to the anticontractile effect. Moreover, the similarity in response to noradrenaline and CL-316,243 suggests that β3-adrenoceptors are the main β-adrenergic subtype mediating this response. Overall, our data indicate that the anticontractile effect of PVAT is dependent on nitric oxide release following upstream activation of β-adrenoceptors.

Impaired endothelium‐dependent vasorelaxation contributes to hypertension in leukocyte‐specific protein 1 (LSP1)‐deficient mice

LSP1 is an intracellular, Ca2+‐and F‐actin‐binding protein expressed in leukocytes and endothelial cells. Adult male LSP1‐deficient (Lsp1−/−) mice had higher blood pressure than their WT counterparts. By Western blot, LSP1 was found present in aorta and mesenteric artery tissues in WT mice but not in Lsp1−/− mice. The role of LSP1 in regulating arterial vascular tone was studied. The contraction of aorta and mesenteric arteries in response to phenylephrine and angiotensin II was similar in WT and Lsp1−/− mice. The relaxation of aorta artery was more sensitive to sodium nitroprusside and methacholine in Lsp1−/− mice than in WT mice. The phenylephrine‐constricted mesenteric arteries showed identical relaxation curve to sodium nitroprusside in these two strains of mice, but had no relaxation to methacholine in Lsp1−/− mice or in denuded WT mesenteric arteries. Methacholine‐induced mesenteric arterial relaxation in WT mice was suppressed partially by the pretreatment of L‐NAME or the combination of apamin + TRAM‐34, abolished completely by the combination of L‐NAME + apamin + TRAM‐34, but not changed by indomethacin. Our results demonstrate the defects in endothelium‐dependent vasorelaxation in resistance arteries in mice deficient in LSP1 revealing an important role for LSP1 in endothelium‐dependent vasodilation in small resistance arteries and blood pressure regulation.

Short‐term estrogen depletion does not promote hypertension in mice

Our lab has previously demonstrated that estrogen (E2) downregulates the expression and function of voltage‐gated, L‐type Ca2+ channels (CaL) in coronary arteries in vitro. In this study we hypothesized that the in vivo depletion of E2 would generate an increase in Ca2+‐dependent arterial tone. Mice (C57BL/6) underwent an ovariectomy (OVX) or sham surgery at 8‐wks of age. Biotelemetry measured a drop in heart rate (p=0.002) and blood pressure (p=0.09) in OVX mice within 2‐wks post‐surgery suggesting a baroreceptor reflex compensation. Mice were sacrificed 4‐wks post‐surgery and the mesenteric arteries isolated. The OVX mice demonstrated atrophied uteri and low E2 plasma levels. In the mesenteric arteries, Western blots demonstrated a slight (p=0.12) increase in CaL expression from OVX mice. Mesenteric arterial reactivity experiments indicated no difference between groups in their spontaneous tone and response to the CaL agonist, FPL64176 (3×10−7 to 1×10−6). This suggests that, while E2 depletion alone does not promote hypertension, other underlying pathologies in conjunction with E2 depletion may lead to hypertension and other cardiovascular diseases. Support: NIGMM of the NIH, Grant #P20 GM103429–11

Mesenteric vascular remodeling in different mouse strains

Inflammatory Bowel Disease (IBD) is a category of inflammatory conditions of the colon and small intestine associated with oxidative stress (OS). Hyperhomocysteinemia (HHcy) is known as the cause of mesenteric vascular remodeling, inducing hypertension and acute abdomen pain. Patients with IBD often have HHcy compare to healthy people. We followed alterations of homocysteine (Hcy) level and mesenteric artery remodeling comparing four different strains of mice: C5BJ/L6 wild type (WT) as a control; cystathione beta synthase heterozygote knockout (CBS+/−) with increased level of Hcy; C3H, which is known as the most resistant to (OS) and double‐transgenic CBS/C3H mice. We suggested that Hcy level will be decreased in CBS/C3H compared to CBS, there will be amelioration of hemodynamics in mesenteric artery of CBS/C3H in comparison with CBS mice and attenuation of mesenteric vascular remodeling in CBS/C3H versus CBS. We used blood flow and mesenteric artery reactivity measurements, immunohistochemistry, electron microscopy of mitochondria, measurement of Dpr‐1 level (mediates mitochondrial fission during OS), measurement of Hcy level in plasma, Masson trichrome, PCR and Western blot (eNOS/iNOS, DDAH‐2, SAHH, CBS, CSE, 3MST, MTHFR, DNMT). Our preliminary data suggests that mesenteric vascular remodeling and HHcy are attenuated and hemodynamic of mesenteric artery is ameliorated in CBS/C3H versus CBS mice.

Hydrogen sulphide–mediated vasodilatation involves the release of neurotransmitters from sensory nerves in pressurized mesenteric small arteries isolated from rats

Hydrogen sulphide (H(2)S) is a gas that has recently been shown to have biological activity. In the majority of blood vessels studied so far, H(2)S has been shown to cause vasorelaxation, although contractile responses have been reported. In the present study, we have made a pharmacological assessment of the effects of H(2)S in mesenteric small arteries isolated from rats. Rat mesenteric small arteries were studied using pressure myography. In pressurised arteries, responses were obtained to the H(2)S donor, sodium hydrogen sulphide (NaHS), in the absence and presence of the NOS inhibitor L-NAME, raised extracellular potassium, the K(ATP) channel inhibitor glibenclamide, the Cl- channel blockers DIDS, NPPB and A9C, the TRPV1 receptor desensitizing agent, capsaicin, the CGRP antagonist, olcegepant, the TRPV1 channel blocker capsazepine and the TRPA1 channel blocker HC-030031. NaHS produced a vasodilator response in rat mesenteric small arteries held at 90 mmHg. Responses to NaHS were not reproducible. Neither, glibenclamide nor, L-NAME inhibited responses to NaHS. DIDS abolished vasodilator responses to NaHS, but these were unaffected by the chloride channel blockers, NPPB and A9C. Responses to NaHS were attenuated after capsaicin pre-treatment, by a CGRP receptor antagonist and an inhibitor of TRPA1 channels. In small arteries isolated from the rat mesentery, NaHS caused a vasodilatation. This response was not reproducible in vitro, since it was mediated by the release of sensory neurotransmitters in a capsaicin-like action. This release was mediated by a H(2)S-induced activation of TRPA1 channels.

Vasorelaxant and antihypertensive effects of ZCM298, a dihydropyridine derivative, are through inhibiting extracellular calcium influx

BackgroundZCM298 is a novel 1,4-dihydropyridine derivative. The aim of the study was to investigate its vasodilation and hypotension, and the related mechanisms. MethodsThe isometric tension of artery ring segments was recorded using an in vitro myography system. The blood pressure of spontaneously hypertensive rats (SHRs) was measured in vivo using a non-invasive tail cuff blood pressure system. Changes in the intracellular calcium concentration ([Ca2+]i) in the mesenteric artery were surveyed using real-time confocal microscopy. Regional cerebral blood flow (rCBF) in the pia mater was monitored by laser-Doppler flowmetry (LDP). ResultsZCM298 (10−9–10−4 M) relaxed rat mesenteric artery obviously and concentration-dependently, which was not affected by the removal of the endothelium. ZCM298 shifted the concentration-contractile curves of mesenteric arteries in response to phenylephrine, U46619, KCl and CaCl2 towards the right in a non-parallel manner. The potency of ZCM298 on relaxing basilar artery was much higher than on mesenteric artery. ZCM298 did not depress the phenylephrine-induced vasoconstriction; however, it inhibited the contraction caused by the addition of CaCl2 in Ca2+-free solution. ZCM298 (10−6 M) inhibited the increase of [Ca2+]i induced by KCl in the artery. ZCM298 improved the rCBF in the pia mater of rats at 0.03 and 0.06mg/kg. ZCM298 depressed the systolic and diastolic blood pressure of SHRs in a dose-dependent manner. ConclusionsZCM298 relaxes arteries probably through inhibiting extracellular calcium influx and decreases the blood pressure of SHRs. ZCM298 is more potent in the basilar artery than in the mesenteric artery and improves rCBF in the pia mater of rats.

Tyrosine Phosphorylation Modulates the Vascular Responses of Mesenteric Arteries from Human Colorectal Tumors

The aim of this study was to analyze whether tyrosine phosphorylation in tumoral arteries may modulate their vascular response. To do this, mesenteric arteries supplying blood flow to colorectal tumors or to normal intestine were obtained during surgery and prepared for isometric tension recording in an organ bath. Increasing tyrosine phosphorylation with the phosphatase inhibitor, sodium orthovanadate produced arterial contraction which was lower in tumoral than in control arteries, whereas it reduced the contraction to noradrenaline in tumoral but not in control arteries and reduced the relaxation to bradykinin in control but not in tumoral arteries. Protein expression of VEGF-A and of the VEGF receptor FLT1 was similar in control and tumoral arteries, but expression of the VEGF receptor KDR was increased in tumoral compared with control arteries. This suggests that tyrosine phosphorylation may produce inhibition of the contraction in tumoral mesenteric arteries, which may increase blood flow to the tumor when tyrosine phosphorylation is increased by stimulation of VEGF receptors.

Endosomal proteolysis regulates calcitonin gene‐related peptide responses in mesenteric arteries

Calcitonin gene-related peptide (CGRP) is a potent vasodilator, implicated in the pathogenesis of migraine. CGRP activates a receptor complex comprising, calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1). In vitro studies indicate recycling of CLR●RAMP1 is regulated by degradation of CGRP in early endosomes by endothelin-converting enzyme-1 (ECE-1). However, it is not known if ECE-1 regulates the resensitization of CGRP-induced responses in functional arterial tissue. CLR, ECE-1a-d and RAMP1 expression in rat mesenteric artery smooth muscle cells (RMA-SMCs) and mesenteric arteries was analysed by RT-PCR and by immunofluorescence and confocal microscopy. CGRP-induced signalling in cells was examined by measuring cAMP production and ERK activation. CGRP-induced relaxation of arteries was measured by isometric wire myography. ECE-1 was inhibited using the specific inhibitor, SM-19712. RMA-SMCs and arteries contained mRNA for CLR, ECE-1a-d and RAMP1. ECE-1 was present in early endosomes of RMA-SMCs and in the smooth muscle layer of arteries. CGRP induced endothelium-independent relaxation of arteries. ECE-1 inhibition had no effect on initial CGRP-induced responses but reduced cAMP generation in RMA-SMCs and vasodilation in mesenteric arteries responses to subsequent CGRP challenges. ECE-1 regulated the resensitization of responses to CGRP in RMA-SMCs and mesenteric arteries. CGRP-induced relaxation did not involve endothelium-derived pathways. This is the first report of ECE-1 regulating CGRP responses in SMCs and arteries. ECE-1 inhibitors may attenuate an important vasodilatory pathway, implicated in primary headaches and may represent a new therapeutic approach for the treatment of migraine.

Altered Reactivity of Tertiary Mesenteric Arteries following Acute Myocardial Ischemia

Background: It is unknown if cardiac ischemia has any deleterious effect on the contractile properties of nonischemic, peripheral vascular beds. Thus, the objective of the present study was to determine whether acute myocardial ischemia results in peripheral vascular dysfunction. Methods and Results: This study characterized force maintenance and the sensitivity to acetylcholine (ACh)-mediated smooth muscle (SM) relaxation of tertiary (3rd) mesenteric arteries from Sprague-Dawley rats following 30 min of myocardial ischemia. Both the phosphorylation of nonmuscle (NM) light chain (LC) and SM-LCs as well as the expression of myosin phosphatase targeting subunit 1 (MYPT1) were also determined. Our data demonstrate that acute myocardial ischemia resulted in vascular dysfunction of 3rd mesenteric vessels, characterized by decreases in force maintenance, ACh- and cGMP-mediated SM relaxation, the phosphorylation of NM-LCs and SM-LCs, and MYPT1 expression. Ischemia was also associated with an increase in protein polyubiquitination, suggesting that during ischemia MYPT1 is targeted for degradation or proteolysis. Conclusion: Acute myocardial ischemia produces peripheral vascular dysfunction; the changes in LC phosphorylation and MYPT1 expression result in a decrease in both tone and the sensitivity to NO-mediated SM relaxation of the peripheral vasculature.

Vascular dysfunction in young, mid-aged and aged mice with latent cytomegalovirus infections

Human cytomegalovirus (HCMV) is associated with vascular diseases in both immunosuppressed and immunocompetent individuals. CMV infections cycle between active and latent phases throughout life. We and others have shown vascular dysfunction during active mouse CMV (mCMV) infections. Few studies have examined changes in physiology during latent CMV infections, particularly vascular responses or whether the negative effects of aging on vascular function and fertility will be exacerbated under these conditions. We measured vascular responses in intact mesenteric and uterine arteries dissected from young, mid-aged, and aged latently mCMV-infected (mCMV genomes are present but infectious virus is undetectable) and age-matched uninfected mice using a pressure myograph. We tested responses to the α(1)-adrenergic agonist phenylephrine, the nitric oxide donor sodium nitroprusside, and the endothelium-dependent vasodilator methacholine. In young latently mCMV-infected mice, vasoconstriction was increased and vasodilation was decreased in mesenteric arteries, whereas both vasoconstriction and vasodilation were increased in uterine arteries compared with those in age-matched uninfected mice. In reproductively active mid-aged latently infected mice, mesenteric arteries showed little change, whereas uterine arteries showed greatly increased vasoconstriction. These vascular effects may have contributed to the decreased reproductive success observed in mid-aged latently mCMV-infected compared with age-matched uninfected mice (16.7 vs. 46.7%, respectively). In aged latently infected mice, vasodilation is increased in mesenteric and uterine arteries likely to compensate for increased vasoconstriction to mediators other than phenylephrine. The novel results of this study show that even when active mCMV infections become undetectable, vascular dysfunction continues and differs with age and artery origin.

Altered sensitivity to nitric oxide donors, induced by intravascular infusion of quantum dots, in murine mesenteric arteries

Quantum dots (QDs) are utilised in imaging diagnostics, tissue engineering and medical therapeutics, however, their influence on vascular function is not ascertained. Here, we examined small mesenteric arterial responses after acute intravascular exposure to QDs. Incubation in mercaptoundecanoic acid (MUA)-coated QDs (at 15μg/mL) had no influence on endothelial-dependent dilator responses (Acetylcholine; Ach) but led to an attenuated relaxation to the nitric oxide donor, sodium nitroprusside (SNP). Conversely, incubation in POSS-PCU coated QDs (at 15μg/mL) led to attenuated Ach responses (10−11–10−3M; n=5, P<0.05), but had no influence on SNP-induced relaxation. At lower concentrations of POSS-PCU coated QDs (5μg/mL), Ach responses were preserved. We demonstrate that acute exposure to QDs, can attenuate vasodilation but not vasoconstriction, and is dependent on their surface coatings. Our findings have implications in QD use for imaging diagnostics in disease states, where SNP based drugs are used in therapeutic intervention. From the Clinical EditorIn this paper, the influence of quantum dots on vascular function is investigated---an important aspect to consider with the growing utility of quantum dots in imaging diagnostics, tissue engineering and medical therapeutics.

185 Mesenteric Artery Reactivity and Small Intestine Morphology in a Chicken Model of Hypoxia-Induced Fetal Growth Restriction

Background and aimsInfants with intrauterine growth retardation are prone to intestinal dysfunction which is manifested by feeding intolerance and, in the most severe cases, necrotizing enterocolitis. The morphological and molecular...

Sex differences in mesenteric endothelial function of streptozotocin-induced diabetic rats: a shift in the relative importance of EDRFs

Several studies suggest that diabetes affects male and female vascular beds differently. However, the mechanisms underlying the interaction of sex and diabetes remain to be investigated. This study investigates whether there are 1) sex differences in the development of abnormal vascular responses and 2) changes in the relative contributions of endothelium-derived relaxing factors in modulating vascular reactivity of mesenteric arteries taken from streptozotocin (STZ)-induced diabetic rats at early and intermediate stages of the disease (1 and 8 wk, respectively). We also investigated the mesenteric expression of the mRNAs for endothelial nitric oxide (NO) synthase (eNOS) and NADPH oxidase (Nox) in STZ-induced diabetes in both sexes. Vascular responses to acetylcholine (ACh) in mesenteric arterial rings precontracted with phenylephrine were measured before and after pretreatment with indomethacin (cyclooxygenase inhibitor), N(ω)-nitro-L-arginine methyl ester (NOS inhibitor), or barium chloride (K(ir) blocker) plus ouabain (Na(+)-K(+)-ATPase inhibitor). We demonstrated that ACh-induced relaxations were significantly impaired in mesenteric arteries from both male and female diabetic rats at 1 and 8 wk. However, at 8 wk the extent of impairment was significantly greater in diabetic females than diabetic males. Our data also showed that in females, the levels of eNOS, Nox2, and Nox4 mRNA expression and the relative importance of NO to the regulation of vascular reactivity were substantially enhanced, whereas the importance of endothelium-derived hyperpolarizing factor (EDHF) was significantly reduced at both 1 and 8 wk after the induction of diabetes. This study reveals the predisposition of female rat mesenteric arteries to vascular injury after the induction of diabetes may be due to a shift away from a putative EDHF, initially the major vasodilatory factor, toward a greater reliance on NO.

Abstract 138: Altered Vascular Reactivity in Mice Overexpressing Adipocyte Mineralocorticoid Receptors - Role of Rho Kinase.

Aldosterone (aldo) plays a role in cardiovascular diseases, including hypertension and obesity. We previously demonstrated that adipocyte-derived factors regulate vascular function and cell signaling in cultured vascular smooth muscle cells. Moreover, adipocytes are able to produce aldo, which influences vascular reactivity. Plasma levels of aldo are positively correlated with obesity and hypertension. However, the pathophysiological role of aldo and mineralocorticoid receptors (MR) in adipose tissue and its interactions with the vasculature remains elusive. In our study, we investigated molecular mechanisms whereby activation of MR, in adipocytes, leads to release of vascular reactive factors and regulation of vascular tone, using a conditional transgenic mouse model that overexpresses MR only in the adipocytes. Vascular reactivity of resistance mesenteric arteries to acetylcholine (Ach), sodium nitroprusside and phenylephrine (Phe), in the absence or presence of fat conditioned medium (Fcm) from control and adipocyte overexpressing MR (DT) mice, was performed by myography. In basal conditions, endothelial dysfunction was not observed in DT or control mice. However, in the presence of Fcm from DT mice, relaxation to Ach was impaired in control mice (Ach 10 -6 M: 77.5±9.6% no Fcm vs. 49.8±7.5% Fcm, p&lt;0.05), an effect blocked by N-acetyl-cysteine (anti-oxidant) (Ach 10 -6 M: 82.2±6.6%). Resistance arteries from DT mice had decreased Phe-induced contraction, compared to control mice (Phe 10 -5 M: 2.7±0.2 mN/mm CT vs. 1.7±0.2 mN/mm DT, p&lt;0.05). Phosphorylation of ezrin, a marker of Rho kinase activation, measured by immunoblotting, was decreased in white and brown adipose tissues of DT (CT: 3.1±0.7 vs. DT: 0.6±0.1, arbitrary units, p&lt;0.05). In conclusion, MR in adipocytes may play an important role in the regulation of vascular function, and may be involved in vascular oxidative stress. MR in adipocytes is also important to the anti-contractile properties of the adipose tissue through downregulation of Rho kinase signaling. Our study identiy novel mechanisms linking vascular and adipose biology through adipocyte MRs.

260 CHRONIC IMPAIRMENT OF ENDOTHELIAL CONNEXIN40 ENHANCES ARTERIAL CONSTRICTION AND REDUCES ARTERIAL DISTENSIBILITY

Background: Synchronization of cellular activity is essential for cardiovascular function, such as the heart beat or vasocontractility. This is achieved through gap junctions and their constituent connexin (Cx) proteins. When Cx40, which highly expressed in the heart, juxtaglomerular apparatus and vascular endothelium, is globally deleted or mutated, significant cardiovascular pathology is observed in both animals and humans. However, the precise function of Cx40 in these different tissues remains unknown. We therefore aimed to determine the impact on cardiovascular function of endothelially expressed Cx40. Methods: We generated transgenic mice in which Cx40 function in the vascular endothelium was reduced through expression of a mutant form, ΔCx40, under control of the endothelium specific promoter, Tie2. We assessed active and passive properties of small mesenteric arteries by measuring the myogenic response to intraluminal pressure and determining vascular elasticity. The involvement of nitric oxide and endothelium derived hyperpolarising factor (EDHF) was determined pharmacologically, while blood pressure and renin secretion were also measured. Results: The myogenic response of mesenteric arteries from ΔCx40 mice was activated at significantly lower intraluminal pressures than in wildtype mice (EC50: Wildtype, 41.7± 0.3mmHg, n=13; ΔCx40, 33.2±0.4 mmHg, n=13; P<0.001). This occurred due to loss of contribution from EDHF. Transgenic vessels also exhibited reduced distensibility (P<0.05) without hypertrophic remodelling of the vascular wall. A modest but significant increase in blood pressure was observed in these mice without change in renin secretion. Conclusions: Life-long impairment in endothelial Cx40 enhances vasoconstriction, reduces vasodilator capacity and arterial distensibility and elevates blood pressure.

Increased Microparticle Production and Impaired Microvascular Endothelial Function in Aldosterone-Salt-Treated Rats: Protective Effects of Polyphenols

We aimed to characterize circulating microparticles in association with arterial stiffness, inflammation and endothelial dysfunction in aldosterone-salt-induced hypertension in rats and to investigate the preventive effects of red wine polyphenols. Uninephrectomized male Sprague-Dawley rats were treated with aldosterone-salt (1 µg.h−1), with or without administration of either red wine polyphenols, Provinols™ (20 mg.kg−1.day−1), or spironolactone (30 mg.kg−1.day−1) for 4 weeks. Microparticles, arterial stiffness, nitric oxide (NO) spin trapping, and mesenteric arterial function were measured. Aldosterone-salt rats showed increased microparticle levels, including those originating from platelets, endothelium and erythrocytes. Hypertension resulted in enhanced aortic stiffness accompanied by increased circulating and aortic NO levels and an upregulation of aortic inducible NO-synthase, NFκB, superoxide anions and nitrotyrosine. Flow-induced dilatation was reduced in mesenteric arteries. These effects were prevented by spironolactone. Provinols™ did not reduce arterial stiffness or systolic hypertension but had effects similar to those of spironolactone on endothelial function assessed by flow-mediated vasodilatation, microparticle generation, aortic NO levels and oxidative stress and apoptosis in the vessel wall. Neither the contractile response nor endothelium-dependent relaxation in mesenteric arteries differed between groups. The in vivo effects of Provinols™ were not mediated by mineralocorticoid receptors or changes in shear stress. In conclusion, vascular remodelling and endothelial dysfunction in aldosterone-salt-mediated hypertension are associated with increased circulating microparticles. Polyphenols prevent the enhanced release of microparticles, macrovascular inflammation and oxidative stress, and microvascular endothelial dysfunction independently of blood pressure, shear stress and mineralocorticoid receptor activation in a model of hyperaldosteronism.

Western-style diet modulates contractile responses to phenylephrine differently in mesenteric arteries from senescence-accelerated prone (SAMP8) and resistant (SAMR1) mice

The influence of two known cardiovascular risk factors, aging and consumption of a high-fat diet, on vascular mesenteric artery reactivity was examined in a mouse model of accelerated senescence (SAM). Five-month-old SAM prone (SAMP8) and resistant (SAMR1) female mice were fed a Western-type high-fat diet (WD; 8 weeks). Mesenteric arteries were dissected, and vascular reactivity, protein and messenger RNA expression, superoxide anion (O 2 (·-) ) and hydrogen peroxide formation were evaluated by wire myography, immunofluorescence, RT-qPCR, ethidium fluorescence and ferric-xylenol orange, respectively. Contraction to KCl and relaxation to acetylcholine remained unchanged irrespective of senescence and diet. Although similar contractions to phenylephrine were observed in SAMR1 and SAMP8, accelerated senescence was associated with decreased eNOS and nNOS and increased O 2 (·-) synthesis. Senescence-related alterations were compensated, at least partly, by the contribution of NO derived from iNOS and the enhanced endogenous antioxidant capacity of superoxide dismutase 1 to maintain vasoconstriction. Administration of a WD induced qualitatively different alterations in phenylephrine contractions of mesenteric arteries from SAMR1 and SAMP8. SAMR1 showed increased contractions partly as a result of decreased NO availability generated by decreased eNOS and nNOS and enhanced O 2 (·-) formation. In contrast, WD feeding in SAMP8 resulted in reduced contractions due to, at least in part, the increased functional participation of iNOS-derived NO. In conclusion, senescence-dependent intrinsic alterations during early stages of vascular senescence may promote vascular adaptation and predispose to further changes in response to high-fat intake, which may lead to the progression of aging-related cardiovascular disease, whereas young subjects lack the capacity for this adaptation.