Small Mesenteric Resistance Arteries Research Articles

Data for proteomic analysis of mouse middle cerebral arteries and small mesenteric resistance arteries

Data for proteomic analysis of mouse middle cerebral arteries and small mesenteric resistance arteries

Functional effects of visfatin in isolated rat mesenteric small resistance arteries

A new adipocytokine, visfatin is expressed in perivascular adipose tissue (PVAT) and exerts effects on vascular system in addition to its relationship with various pathological conditions. The present study aimed to investigate the functional effects of visfatin and the possible underlying mechanism(s) of the effects of visfatin in isolated rat mesenteric small resistance arteries. The study was conducted in small resistance arterial rings isolated from rat mesenteric vascular beds. While visfatin incubation did not produce significant alterations in contractile responses of mesenteric arterial rings to noradrenaline, relaxation responses to acetylcholine but not to sodium nitroprusside (SNP) were significantly reduced in endothelium-intact rings. The inhibitory effect of visfatin on responses to acetylcholine was not observed in endothelium-denuded preparations. Incubation of tissues with nicotinamide phosphoribosyl transferase (NAMPT) inhibitor FK866 or superoxide dismutase (SOD) reversed the inhibitory effects of visfatin on relaxation responses to acetylcholine. Co-incubation of visfatin with Nω-nitro-L-arginine methylester (L-NAME) did not produce a significant alteration in vascular responses to acetylcholine compared to L-NAME incubation alone. Mesenteric PVAT visfatin levels were significantly higher than and correlated positively with plasma visfatin levels. The results of our study indicated that visfatin-induced reductions in endothelium-dependent relaxations of rat isolated small resistance arteries are mediated by oxygen free radicals and a reduction in nitric oxide (NO) bioavailability. It was suggested that increment in systemic and/or local visfatin levels due to various pathologies including obesity and excessive weight gain may play a substantial role in initiation and/or propagation of vascular dysfunctions.

"A Step and a Ceiling": mechanical properties of Ca2+ spark vasoregulation in resistance arteries by pressure-induced oxidative activation of PKG.

We investigated the biomechanical relationship between intraluminal pressure within small mesenteric resistance arteries, oxidant activation of PKG, Ca2+ sparks, and BK channel vasoregulation. Mesenteric resistance arteries from wild type (WT) and genetically modified mice with PKG resistance to oxidative activation were studied using wire and pressure myography. Ca2+ sparks and Ca2+ transients within vascular smooth muscle cells of intact arteries were characterized using high‐speed confocal microscopy of intact arteries. Arteries were studied under conditions of varying intraluminal pressure and oxidation. Intraluminal pressure specifically, rather than the generic stretch of the artery, was necessary to activate the oxidative pathway. We demonstrated a graded step activation profile for the generation of Ca2+ sparks and also a functional “ceiling” for this pressure –‐sensitive oxidative pathway. During steady state pressure ‐ induced constriction, any additional Ca2+ sensitive‐K+ channel functional availability was independent of oxidant activated PKG. There was an increase in the amplitude, but not the Area under the Curve (AUC) of the caffeine‐induced Ca2+ transient in pressurized arteries from mice with oxidant‐resistant PKG compared with wild type. Overall, we surmise that intraluminal pressure within resistance arteries controls Ca2+ spark vasoregulation through a tightly controlled pathway with a graded onset switch. The pathway, underpinned by oxidant activation of PKG, cannot be further boosted by additional pressure or oxidation once active. We propose that these restrictive characteristics of pressure‐induced Ca2+ spark vasoregulation confer stability for the artery in order to provide a constant flow independent of additional pressure fluctuations or exogenous oxidants.

AMP‐Activated Protein Kinase (AMPK) Modulates Myoendothelial Junctions (MEJ) in Murine Microvessels

Myoendothelial junctions (MEJ) in microvessels are not only the site of direct cell communication via gap junctions but also represent a signaling microdomain critical for regulating vascular processes such as endothelium dependent hyperpolarization (EDH). The 5′‐adenosine monophosphate‐activated protein kinase (AMPK) has been shown to influence vascular tone. It has not yet been studied, however, whether AMPK could also affect MEJ. Thus, the aim of this project was to elucidate the impact of AMPK on MEJ dynamics and its potential mechanism of action.We studied isolated small mesenteric and skeletal muscle resistance arteries of mice using a pressure myograph system developed in our laboratory. Using confocal and two photon excitation fluorescence microscopy we identified the internal elastic lamina, which in these vessels exhibits small holes that allow direct contact between endothelial and smooth muscle cells. On average, we found that in about 35% of the holes, structures with a bright actin core as assessed by using F‐actin fluorescence in LifeAct mice or by staining with phalloidin which we could define as sites of MEJ. These structures showed expression of Cx37 and Cx43 suggesting that gap junctions (MEGJs) were also localized in these structures. Over observation times of up to 2 hours, we found that the number and localization of these MEJ varied, while they always positioned in the area of holes of IEL only. Thus, for the first time we could investigate MEJ dynamics. AMPK negatively regulates MEJ formation, since in AMPK α1, but not α2 knockout (KO) mice, the number of MEJ was significantly increased. In accordance, in wild type mice, incubation with the AMPK inhibitory compound C (CC) significantly augmented the number of MEJ. Furthermore, KO of AMPK α1, in intact mice as well as in cultured human and porcine smooth muscle cells, went along with an increase in PAI‐1 expression. Accordingly, incubation of arteries with exogenous PAI‐1 also increased the number of MEJ. The negative effect of AMPK on PAI‐1 could be explained by enhanced expression of the silencer of the PAI‐1 gene, small heterodimer partner (SHP). Since MEJ are also involved in EDH, we studied whether an enhanced formation of MEJ correlated with the magnitude of EDH induced vasodilation. Indeed, a higher amount of MEJs in α1 KO mice went along with a left shift of acetylcholine (ACh) ‐induced dilation dose effect curve.This work describes for the first time a role of the α1 subunit of AMPK as a potent modulator of MEJ dynamics which is probably mediated by controlling PAI‐1 expression. Our data also for the first time demonstrate a dynamic regulation of MEJ expression in intact blood vessels.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Differential effects of long-term slow-pressor and subpressor angiotensin II on contractile and relaxant reactivity of resistance versus conductance arteries.

Vascular reactivity was evaluated in three separate arteries isolated from rats after angiotensin II (Ang II) was infused chronically in two separate experiments, one using a 14‐day high, slow‐pressor dose known to produce hypertension and the other using a 7‐day low, subpressor but hypertensive‐sensitizing dose. There were three new findings. First, there was no evidence of altered vascular reactivity in resistance arteries that might otherwise explain the hypertension due to the high Ang II or the hypertensive‐sensitizing effect of the low Ang II dose. Second, the high Ang II dose exerted a novel differential effect on arterial contractile responsiveness to the sympathetic neurotransmitter, norepinephrine, depending on the level of sympathetic innervation. It clearly enhanced that responsiveness in the sparsely innervated aorta but not in small mesenteric resistance arteries or the proximal (conductance) portion of the caudal artery, both of which are densely innervated. This suggests that the increased expression of alpha adrenergic receptors after long‐term exposure to Ang II as previously reported for aortic smooth muscle, is prevented in densely innervated arteries, likely due to long‐term Ang II‐mediated increase in sympathetic neural traffic to those vessels. Third, the same high dose of Ang II impaired aortic relaxation in response to the nitric oxide (NO) donor nitroprusside without impairing aortic endothelium‐dependent relaxation. NO is the main relaxing substance released by aortic endothelium. Accordingly, it is possible that this dose of Ang II is also associated with enhanced release of and/or enhanced smooth muscle responsiveness to other endothelial relaxing substances in a compensatory capacity.

PP.12.08] EFFECTS OF MELATONIN ON THE PRODUCTION OF ADIPONECTIN AND THE EXPRESSION OF ADIPONECTIN RECEPTOR IN THE VISCERAL ADIPOSE TISSUE OF AGING MICE.

Objective: It has been previously demonstrated that inflammation in adipose tissue may be implicated in vascular dysfunction (Circulation 2009; 119(12):1661–1670). A senescence-accelerated prone mouse (SAMP8) is a model of age-related cognitive decline and vascular dysfunction. Several studies demonstrated that SAMP8 suffers from increased oxidative stress and that accelerated senescence was associated with increased oxygen radicals synthesis. We have also recently demostrated that the anticontractile effect of perivascular fat is impaired in a senescence-accelerated prone mouse, compared with control senescence-resistant mice (SAMR1). A long-term treatment with melatonin seems to decrease contractile responses to norepinephrine in mesenteric small arteries of SAMP8, thus restoring an anticontractile effect, probably through antioxidant mechanisms. However, it is not know whether melatonin is able to modulate the production of adiponectin and/or the expression of adiponectin receptors. Therefore, the aim of the study was to investigate the production of adiponectin and/or the expression of adiponectin receptors in the visceral adipose tissue of SAMP8 before and after chronic treatment with melatonin. Design and method: We investigated 7 SAMP8 and 7 SAMR1 treated for 10 months with melatonin (MEL, 10 mg/kg/day), an endogenous indoleamine with antioxidant and vasculoprotective properties, as well as 7 SAMP8 and 7 SAMR1 untreated controls (CTR) kept untreated for 10 months. Visceral fat (perirenal fat) was obtained by dissection, and the adiponectin content, as well as the expression of adiponectin receptor were evaluated by immunohistochemistry. Results: Results are shown in the figure (* p < 0.05 vs SAMR1 CTR; # p < 0.05 vs SAMR1 + MEL; + p < 0.05 vs SAMP8 + MEL). The adiponectin content, as well as the expression of adiponectin receptor were reduced in untreated SAMP8 compared with untreated SAMR1. Treament with MEL was able to increase the production of both proteins in SAMP8.Conclusions: Melatonin treatment is able to increase the production of adiponectin and the expression of adiponectin receptor in the visceral fat of aging mice. This observation might contribute to explain the improvement of the anticontractile action of perivascular fat observed in the mesenteric small resistance arteries of SAMP8 mice after chronic treatment with MEL.

Effect of long-term treatment with melatonin on vascular markers of oxidative stress/inflammation and on the anticontractile activity of perivascular fat in aging mice.

Some reports have suggested that inflammation in perivascular adipose tissue (PVAT) may be implicated in vascular dysfunction by causing the disappearance of an anticontractile effect. The aim of this study was to investigate the effects of chronic melatonin treatment on the functional responses of the small mesenteric arteries and on the expression of markers of inflammation/oxidative stress in the aortas of senescence-accelerated prone mice (SAMP8), a model of age-related vascular dysfunction. We investigated seven SAMP8 and seven control senescence-accelerated resistant mice (SAMR1) treated for 10 months with melatonin, as well as equal numbers of age-matched untreated SAMP8 and SAMR1. The mesenteric small resistance arteries were dissected and mounted on a wire myograph, and the concentration-response to norepinephrine was evaluated in vessels with intact PVAT and after the removal of the PVAT. The expression of markers of oxidative stress, inflammation and aging in the aortas was evaluated by immunostaining. In addition, the adiponectin content and the expression of adiponectin receptor 1 were evaluated in the visceral adipose tissue. In untreated SAMP8 mice, we observed an overexpression of oxidative stress and inflammatory markers in the vasculature compared with the controls. No anticontractile effect of the PVAT was observed in untreated SAMP8 mice. Long-term treatment of SAMP8 mice with melatonin increased the expression of some markers of vasoprotection, decreased oxidative stress and inflammation and restored the anticontractile effect of the PVAT. Decreased expression of adiponectin and adiponectin receptor 1 was also observed in visceral fat of untreated SAMP8, whereas a significant increase was observed after melatonin treatment.

Chronic black tea extract consumption improves endothelial function in ovariectomized rats.

Purpose Menopause escalates the risk of cardiovascular diseases in women. There is an unmet need for better treatment strategy for estrogen-deficiency-related cardiovascular complications. Here we investigated the impact of chronic black tea extract (BT) consumption on cardiovascular function and lipid metabolism using a rat model of estrogen deficiency.MethodsFemale Sprague–Dawley rats were ovariectomized (OVX) and treated with BT (15 mg/kg/day, 4 weeks; active ingredients: theaflavins) or estrogen (E2) treatment for 4 weeks. Serum was collected for measuring cholesterol, triacylglycerol and estradiol levels. Changes in vascular reactivity were examined. The protein levels of NADPH oxidases were assessed by Western blotting. Reactive oxygen species (ROS) level was measured using dihydroethidium fluorescence imaging. The concentrations of cGMP were measured using ELISA kit.ResultsAortic rings from control, BT-treated and E2-treated OVX rats exhibited a greater increase in Phe-induced contraction after inhibition of NO synthase compared with those from OVX rats. ACh-induced endothelium-dependent relaxations were augmented in aortae and renal arteries in BT/E2-treated OVX rats than in OVX rats. BT/E2 treatment improved flow-mediated dilatation in small mesenteric resistance arteries of OVX rats. BT/E2 treatment restored the eNOS phosphorylation level and reversed the up-regulation of NADPH oxidases and ROS overproduction in OVX rat aortae. ACh-stimulated cGMP production was significantly elevated in the aortae from BT- and E2-treated rats compared with those from OVX rats. BT/E2 treatment reduced circulating levels of total cholesterol.ConclusionsThe present study reveals the novel benefits of chronic BT consumption to reverse endothelial dysfunction and favorably modifying cholesterol profile in a rat model of estrogen deficiency and provides insights into developing BT as beneficial dietary supplements for postmenopausal women.

Unique gene program of rat small resistance mesenteric arteries as revealed by deep RNA sequencing

Deep sequencing of RNA samples from rat small mesenteric arteries (MA) and aorta (AO) identified common and unique features of their gene programs. ∼5% of mRNAs were quantitatively differentially expressed in MA versus AO. Unique transcriptional control in MA smooth muscle is suggested by the selective or enriched expression of transcription factors Nkx2-3, HAND2, and Tcf21 (Capsulin). Enrichment in AO of PPAR transcription factors and their target genes of mitochondrial function, lipid metabolism, and oxidative phosphorylation is consistent with slow (oxidative) tonic smooth muscle. In contrast MA was enriched in contractile and calcium channel mRNAs suggestive of components of fast (glycolytic) phasic smooth muscle. Myosin phosphatase regulatory subunit paralogs Mypt1 and p85 were expressed at similar levels, while smooth muscle MLCK was the only such kinase expressed, suggesting functional redundancy of the former but not the latter in accordance with mouse knockout studies. With regard to vaso-regulatory signals, purinergic receptors P2rx1 and P2rx5 were reciprocally expressed in MA versus AO, while the olfactory receptor Olr59 was enriched in MA. Alox15, which generates the EDHF HPETE, was enriched in MA while eNOS was equally expressed, consistent with the greater role of EDHF in the smaller arteries. mRNAs that were not expressed at a level consistent with impugned function include skeletal myogenic factors, IKK2, nonmuscle myosin, and Gnb3. This screening analysis of gene expression in the small mesenteric resistance arteries suggests testable hypotheses regarding unique aspects of small artery function in the regional control of blood flow.

7D.10

Objective: It has been previously demonstrated that inflammation in adipose tissue may be implicated in vascular dysfunction (Circulation 2009; 119(12):1661–1670). A senescence-accelerated prone mouse (SAMP8) is a model of age-related cognitive decline and vascular dysfunction. Several studies demonstrated that SAMP8 suffers from increased oxidative stress and that accelerated senescence was associated with decreased eNOS and nNOS and increased oxygen radicals synthesis. Aim of the study was to investigate functional responses of small mesenteric arteries in a senescence-accelerated prone mouse (SAMP8) before and after chronic treatment with melatonin.Design and method: We investigated 7 SAMP8 and 7 SAMR1 normal controls. Mesenteric small resistance arteries were dissected and mounted on a wire myograph, according to Mulvany-Halpern technique (internal diameter about 200 μm). A concentration-response to norepinephrine (NE, from 10–9 to 10-5 Mol/l) was evaluated in vessels with intact prerivascular fat tissue (WF) and in vessels in which perivascular fat tissue was removed (NoF). Investigations were repeated in 7 SAMP8 and 7 SAMR1 after 54 weeks of chronic treatment with melatonin, an endogenous hormone with antioxidant and vasculoprotective properties. Results: In SAMR1 control mice anticontractile effect of perivascular fat was present (WF vs. NoF: ANOVA p = 0.04), while in aging SAMP8 mice the effect was less pronounced (WF vs. NoF: ANOVA p = NS) (see figure). Long-term treatment with melatonin had no effect in SAMR1 either in WF or NoF vessels, while it decreased the contractile response to norepinephrine in noF vessels of SAMP8 (ANOVA p < 0.001); the effect of melatonin treatment in WF vessels was not statistically significant. Conclusions: The anticontractile effect of perivascular fat is impaired in a senescence-accelerated prone mouse, compared with controls. A long-term treatment with melatonin seems to decrease contractile responses to norepinephrine in NOF mesenteric small arteries of SAMP8, thus restoring an anticontractile effect, probably through antioxidant mechanisms.

Conditional knockout of smooth muscle sodium calcium exchanger type-1 lowers blood pressure and attenuates Angiotensin II-salt hypertension.

The functions of smooth muscle sodium calcium exchanger (NCX) in the vasculature are controversial and poorly understood. To determine the possible roles of NCX in the vascular phenotype and function, we developed a novel mouse model (SM‐NCX1 KO) in which the smooth muscle‐specific NCX type‐1 (NCX1) was conditionally knocked out using tamoxifen‐inducible Cre‐loxP recombination technique. SM‐NCX1 KO mice exhibit significantly lower blood pressure and attenuated angiotensin II (Ang II)‐salt‐induced hypertension (measured by radio telemetry and intra‐arterial catheterization). Isolated, pressurized mesenteric small resistance arteries from SM‐NCX1 KO mice, compared to control arteries, were characterized by the following: (1) ~90% reduced NCX1 protein expression; (2) impaired functional responses to (i) acute NCX inhibition by SEA0400 or SN‐6, (ii) NCX activation by low [Na+]o, and (iii) Na+ pump inhibition by ouabain; (3) attenuated myogenic reactivity; and (4) attenuated vasoconstrictor response to phenylephrine but not Ang II. These results provided direct evidence that arterial NCX1 normally mediates net Ca2+ influx that helps maintain basal vascular tone in small resistance arteries and blood pressure under physiological conditions. Importantly, NCX1 contributes to blood pressure elevation in Ang II‐salt hypertension, possibly by regulating α‐adrenergic receptor activation.

Anticontractile activity of perivascular fat in obese mice and the effect of long-term treatment with melatonin.

It has been demonstrated previously that inflammation in perivascular adipose tissue (PVAT) may be implicated in vascular dysfunction. The aim of this study was to investigate the functional responses of small mesenteric arteries in a hyperphagic animal model of obesity after chronic treatment with melatonin, an endogenous hormone with antioxidant and vasculoprotective properties. Ten obese mice (ob/ob) and 10 control lean mice (CLM) were treated with melatonin 100 mg/kg per day in the drinking water for 8 weeks. Mesenteric small resistance arteries were dissected and mounted on a wire myograph and a concentration-response to norepinephrine was evaluated in vessels with intact PVAT and after PVAT was removed and in the presence of iberiotoxin, a selective blocker of BKCA channels as well as under conditions of induced hypoxia in vitro. The presence of PVAT reduced the contractile response to norepinephrine in both ob/ob and CLM; however, the effect was significantly reduced in ob/ob. The anticontractile effect of PVAT completely disappeared with iberiotoxin preincubation. After melatonin treatment, inflammation was significantly ameliorated, and the contractile response in ob/ob and CLM was significantly reduced when PVAT was removed. Anticontractile effect of PVAT that is lost in obesity can be rescued using melatonin. A reduced expression of adiponectin and adiponectin receptor was observed in perivascular fat of ob/ob, whereas significant increase was observed in ob/ob treated with melatonin. Melatonin seems to exert a protective effect on arteries from both ob/ob and CLM, counteracting the adverse effect of hypoxia and iberiotoxin.

Chronic intermittent hypoxia alters blood pressure and mesenteric small resistance arterial responsiveness in mice (710.9)

Blood pressure (BP) is elevated in rats and mice exposed to chronic intermittent hypoxia (CIH). It is unclear if altered vascular function, such as myogenic reactivity (MR), contributes to BP elevation in CIH. We aimed to characterize the vascular alterations in small resistance arteries from CIH‐exposed mice. C57BL/6J mice were exposed to either CIH (1 min alterations of 5% and 21% O2) or normoxia (sham control, constant 21% O2), 8 hrs/day for 5 weeks. Compared to sham controls, CIH‐exposed mice had significantly higher BP, measured by radio telemetry. Mesenteric small resistance arteries were isolated and pressurized for diameter and wall thickness measurement. Myogenic constriction in response to intraluminal pressure increase (i.e., MR) was significantly attenuated, especially at pressures 蠅 70 mm Hg. At 70 mm Hg, vasodilations to acetylcholine and sodium nitroprusside, but not to external Ca2+ removal, were significantly impaired in CIH, whereas vasoconstrictions to phenylephrine (PE), endothelin‐1, U46619, and high [K+]o were similar between sham and CIH arteries. When intraluminal pressure was increased to 110 mm Hg, however, PE‐induced vasoconstriction was significantly reduced, and nitric oxide‐dependent vasodilation was further attenuated in CIH arteries. Arterial remodeling was not obvious since wall thickness, wall to lumen ratio, and passive diameters in Ca2+‐free solution were similar between two groups. Thus, CIH 1) elevates BP; 2) reduces MR; 3) reduces arterial responsiveness to vasodilators, and, at higher intraluminal pressure, to vasoconstrictors; 4) has no effect on arterial remodeling in mouse mesenteric small resistance arteries. It is likely that the altered arterial responsiveness is involved in the regulation of BP in CIH.Grant Funding Source: NIH/NHLBI R01 HL107654

Abstract 533: Vascular Smooth Muscle Na/Ca Exchanger-1 Modulates Myogenic Reactivity and Blood Pressure in Chronic Intermittent Hypoxia

Chronic intermittent hypoxia (CIH) results in systemic hypertension and altered vascular reactivity in human and rodents. We hypothesize that vascular smooth muscle (VSM) Na/Ca exchanger type-1 (NCX1) is a potential molecular mechanism mediating CIH-induced changes in vascular reactivity and blood pressure (BP). Wild type (WT) and transgenic (TG) mice with VSM-specific NCX1 overexpression were exposed to either CIH (1 min 5% nadir O 2 , followed by 1 min 21% O 2 ; 8 hrs/day for 5 weeks), or normoxia (constant 21% O 2 ). CIH resulted in greater BP elevation in TG (102 ± 3 mm Hg vs. 92 ± 6 mm Hg in normoxia, P &lt; 0.05, n = 6 and 3, measured by intra-carotid artery catheterization under 1.5% isoflurane) than in WT (88 ± 4 mm Hg vs. 83 ± 2 mm Hg in normoxia, NS, n = 3 and 5) mice. Mesenteric small resistance arteries were then isolated and pressurized for diameter and wall thickness measurement. Compared to normoxia controls, CIH-exposed WT arteries exhibited increased resting myogenic tone (21 ± 2% vs. 16 ± 2% of PD in normoxia, P &lt; 0.05, n = 5 each group). Moreover, myogenic reactivity in WT arteries of CIH-exposed mice shifted downward at lower range of intraluminal pressure (20-80 mm Hg), and upward at higher pressure (100-140 mm Hg). The effects of CIH were significantly greater in TG arteries compared to those in WT arteries, suggesting that VSM NCX1 augmented the myogenic response to CIH. Vasoconstrictions induced by phenylephrine (0.003-100 μM), greater in TG arteries, or high (60 mM) external K + solution, were similar between control and CIH groups in the same genotype, suggesting that arterial contractility to agonist stimulation was not affected by CIH. Arterial wall thickness or passive diameters under various intraluminal pressures in Ca 2+ -free solution were not different between control and CIH groups in either genotype, suggesting a lack of arterial stiffness or remodeling. In summary, 1) CIH results in greater BP elevation in TG mice; 2) CIH leads to biphasic changes in myogenic reactivity in small resistance arteries; 3) Overexpression of VSM NCX1 exaggerates BP elevation in CIH, likely by modulating myogenic reactivity, suggesting that VSM NCX1 is a potential mechanism mediating CIH-induced hypertension.

Effect of Captopril and Telmisartan on Anti-contractile Property of Perivascular Adipose Tissue Lost after Hypoxia in Rat Mesenteric Small Resistance Arteries

Final body weight (gr) 211 3,85 0,005 246,4 11,7 < 0,0001vsCTRL 263,3 7,3 c-fPWV cm/sec 378,9 7,2 <,0001 560,8 12,1 < 0,0001 363,3 5,9 MeanBP mmHg 108,8 4,8 0,002 129,8 2,6 0,003 109,3 2,7 Superoxide nmol/min/mm 97,4 8,3 0,01 142,3 14,6 0,04 105,0 9,8 SystolicBP mmHg 133,1 6,1 0,07 153,3 12,9 0,11 135,9 4,1 DiastolicBP mmHg 96,7 4,6 0,09 109,1 6,9 0,1 97,2 2,6 2,5 0,08 < 0,0001< 0,0001vsCTRL 5,2 0,2

Role of store-operated calcium entry in imperatorin-induced vasodilatation of rat small mesenteric artery

Store-operated Ca 2+ entry (SOCE) has recently been proposed to contribute to Ca 2+ influx in vascular smooth muscle cells (VSMCs). Imperatorin is known for its potent vasodilatory effects as a dietary furanocoumarin. The study was designed to examine the hypothesis that SOCE have a functional role in imperatorin-induced vasodilation. Small mesenteric resistance arteries and mesenteric VSMCs were obtained from rats. Isometric tensions of isolated artery rings were measured by a sensitive myograph system. Laser scanning confocal microscopy was used to determine the intracellular Ca 2+ concentration of fluo-3-loaded VSMCs. Imperatorin (1–100 μM) relaxed artery rings precontracted by phenylephrine in a concentration-dependent manner. In cultured mesenteric VSMCs, passive store depletion by thapsigargin and active store depletion by phenylephrine both induced Ca 2+ influx due to SOCE. Imperatorin didn't inhibit SOCE-mediated increases in cytosolic Ca 2+ levels evoked by the emptying of the stores. In isolated artery rings, imperatorin didn't inhibit SOCE-induced contractions due to store depletion. Our results exclude SOCE mechanism of vasodilatation by imperatorin. But imperatorin is partly similar with nifedipine in vasorelaxation effect.

CAPTOPRIL BUT NOT TELMISARTAN, RESTORES THE ANTI-CONTRACTILE EFFECTS OF PERIVASCULAR FAT TISSUE LOST AFTER HYPOXIA IN RAT MESENTERIC SMALL RESISTANCE ARTERIES: PP.11.422

It has been previously demonstrated that inflammation in adipose tissue may be implicated in vascular dysfunction (Circulation 2009; 119(12):1661–1670). In particular, adipocytes secrete adiponectin, a physiological modulator of local vascular tone through an increased nitric oxide bioavailability. This capacity is lost in obesity by the development of adipocyte hypertrophy, leading to hypoxia, inflammation, and oxidative stress. Aim of the study was to investigate the feasibility a pharmacological modulation of the concerned effect in an animal model (Wistar-Kyoto normotensive rats: WKY). Materials and Methods: We investigated 25 WKY of 12 weeks of age. Mesenteric small resistance arteries were dissected and mounted on a wire myograph, according to Mulvany-Halpern technique (internal diameter about 200 μm). A concentration-response to norepinephrine (NE, from 10–9 to 10–5 Mol/l) was evaluated in the following conditions: 1) in vessels with intact prerivascular fat tissue (WF); 2) in vessels in which perivascular fat tissue was removed (NoF); 3) in WF vessels kept in hypoxic condition (no oxygen supply) (WF + Hyp); 4) in NoF vessels kept in hypoxic condition (NoF + Hyp); 5) in WF + Hyp vessels incubated with telmisartan 10–2 Mol/l for 3 hour (WF + Hyp + Telm); 6) in WF + Hyp vessels incubated with captopril 10–2 Mol/l for 3 hour (WF + Hyp + Capt). Results are summarized in the figure (active media stress: KPa, mean of two vessels for each rat).A significantly greater reactivity to NE was observed in NoF vessels compared with WF vessels (ANOVA p = 0.003 between curves), confirming previous observations. This increased reactivity is similar to that observed in WF + Hyp (ANOVA p = NS between, NoF and WF + Hyp), therefore hypoxia seems to abolish the anti-vasoconstrictor effects of perivascular fat. Captopril was able to completely prevent the effect of hypoxia (ANOVA p < 0.05 between WF + Hyp and WF + Hyp + Capt), while telmisartan had a very modest effect. In conclusion, the ACE inihibitor captopril, but not the angiotensin-receptor blocker telmisartan, seems to be able to restore the anti-contractile effects of perivascular fat tissue lost after hypoxia, possibly through an increased bradykinin activity.

EFFECT OF ANTIHYPERTENSIVE TREATMENT WITH FOSINOPRIL ON MECHANICAL PROPERTIES OF SMALL MESENTERIC ARTERIES OF SPONTANEOUSLY HYPERTENSIVE RATS: PP.11.433

A pronounced fibrosis, as documented by an increased collagen:elastin ratio may be observed in the tunica media of mesenteric small resistance arteries of spontaneously hypertensive rats (SHR), compared with Wistar-Kyoto controls (Sharifi EM et al, J Hypertens 1998; 16:457–466). However, the consequences of such fibrotic changes in the microvasculature in terms of elastic properties are controversial, and either no change or a paradoxical increase in distensibility/decreased elastic modulus was observed in SHR (Mulvany MJ, Hypertension 1992 20: 7–9; Laurant P et al, J Vasc Res 1997, 34: 117–125). The aim of this study was to evaluate the effects of the ACE inhibitor fosinopril given either at hypotensive, high dose (FOS hd) or at a non-hypotensive, low dose (FOS ld), on mechanical properties (relationship between incremental elastic modulus/wall stress) of mesenteric resistance arteries in SHR, compared with WKY controls. Fosinopril was administered in the drinking water from the 6th to the 12th week of age. Rats were sacrificed at 12 weeks of age and heart weight/body weight ratio was measured. Mesenteric arterioles were dissected and mounted on a micromyograph (Mulvany's technique), and the media to lumen ratio (M/L) was measured. Results are summarized in the Table (** = p < 0.01 vs. untreated SHR; #p < 0.001 vs. untreated WKY).During the 6 weeks of treatment, systolic blood pressure in SHR treated with high dose fosinopril was significantly lower in comparison with untreated SHR, while no difference was observed with low dose fosinopril. On the contrary, in SHR treated with both high dose and low dose fosinopril, a statistically significant reduction of M/L was observed. However, no difference in the relation between incremental elastic modulus and wall stress of mesenteric resistance arteries between untreated SHR and WKY, or between treated and untreated SHR was observed. In conclusion, our data suggest that changes in the structure of mesenteric small resistance arteries in SHR are not associated with parallel changes in the mechanical properties/vascular distensibility.

Effects of Melatonin and Pycnogenol on Small Artery Structure and Function in Spontaneously Hypertensive Rats

It was suggested that oxidative stress has a key role in the development of endothelial dysfunction, as well as microvascular structural alterations. Therefore, we have investigated 2 substances with antioxidant properties: melatonin and Pycnogenol. We treated 7 spontaneously hypertensive rats (SHRs) with melatonin and 7 with Pycnogenol for 6 weeks. We compared results obtained with those observed in 7 SHRs and 7 Wistar-Kyoto normotensive control rats kept untreated. Mesenteric small resistance arteries were dissected and mounted on a wire myograph, and a concentration-response curve to acetylcholine was performed. Aortic contents of metalloproteinase 2, Bax, inducible NO synthase, and cyclooxygenase 2 were evaluated, together with the aortic content of total collagen and collagen subtypes and apoptosis rate. A small reduction in systolic blood pressure was observed. A significant improvement in mesenteric small resistance artery structure and endothelial function was observed in rats treated with Pycnogenol and melatonin. Total aortic collagen content was significantly greater in untreated SHRs compared with Wistar-Kyoto control rats, whereas a full normalization was observed in treated rats. Apoptosis rate was increased in the aortas of untreated SHRs compared with Wistar-Kyoto control rats; an even more pronounced increase was observed in treated rats. Bax and metalloproteinase 2 expressions changed accordingly. Cyclooxygenase 2 and inducible NO synthase were more expressed in the aortas of untreated SHRs compared with Wistar-Kyoto control rats; this pattern was normalized by both treatments. In conclusion, our data suggest that treatment with Pycnogenol and melatonin may protect the vasculature, partly independent of blood pressure reduction, probably through their antioxidant effects.

Role of Heme Oxygenase in Modulating Endothelial Function in Mesenteric Small Resistance Arteries of Spontaneously Hypertensive Rats

It has been proposed that endothelial dysfunction is due to the excessive degradation of nitric oxide (NO) by oxidative stress. The enzyme heme-oxygenase (HO) seems to exert a protective effect on oxidative stress in the vasculature, both in animal models and in humans. The objective of this study is to evaluate the effects of inhibition or activation of HO on endothelial function in mesenteric small resistance arteries of spontaneously hypertensive rats (SHR). Six SHR were treated with cobalt protoporphyrin IX 50 mg/Kg (CoPP), an activator of HO; six SHR with stannous mesoporphyrin 30 mg/Kg (SnMP), an inhibitor of HO, and six SHR with saline. As controls, six Wistar-Kyoto rats (WKY) were treated with CoPP, six WKY with SnMP, and six WKY with saline. Drugs were injected in the peritoneum once a week for 2 weeks. Systolic blood pressure (SBP) was measured (tail cuff method) before and after treatment. Mesenteric small resistance arteries were mounted on a micromyograph. Endothelial function was evaluated as a cumulative concentration-response curve to acetylcholine (ACH), before and after preincubation with N(G)-methyl-L-arginine (L-NMMA, inhibitor of NO synthase), and to bradykinin (BK). In SHR treatment with CoPP, improved ACH-and BK-induced vasodilatation (ANOVA p < 0.001) and this improvement was abolished by L-NMMA (ANOVA p < 0.001). SnMP was devoid of effects on endothelial function. In WKY, both activation and inhibition of HO did not substantially affect endothelium-mediated vasodilatation. The stimulation of HO seems to induce an improvement of endothelial dysfunction in SHR by possibly reducing oxidative stress and increasing NO availability.