2K1C Rats Research Articles

Effects of oxidative stress on endothelial modulation of contractions in aorta from renal hypertensive rats

BackgroundEndothelial dysfunction is linked to exaggerated production of superoxide anions. This study was conducted to examine the effects of oxidative stress on endothelial modulation of contractions in chronic two-kidney, one-clip (2K1C) renal hypertensive rats. MethodsThe 2K1C hypertension was induced by clipping the left renal artery; age-matched rats receiving sham treatment served as controls. Thoracic aortae were isolated and mounted in tissue baths for measurement of isometric tension. ResultsNorepinephrine-induced contraction was augmented by the removal of the endothelium, which was more pronounced in sham rats than in 2K1C rats. Nω-nitro-L-arginine methyl ester, an inhibitor of nitric oxide production, had a similar augmenting effect. Vitamin C inhibited the contraction in aortic rings with intact endothelium from 2K1C rats but not from sham rats. The contraction was also suppressed by treatment with diphenyleneiodonium or apocynin, inhibitors of nicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide phosphate (NADH/NADPH) oxidase, in the aortae with intact endothelium from 2K1C rats but not in those from sham rats. Superoxide anions generated by xanthine oxidase/hypoxanthine enhanced the contraction in the aortae with intact endothelium from sham rats, but had no effect in 2K1C rats. Enhanced contractile responses to norepinephrine by xanthine oxidase/hypoxanthine in sham rats were reversed by vitamin C. ConclusionThese results suggest that the effect on endothelial modulation of endothelium-derived nitric oxide is impaired in 2K1C hypertension. The impairment is, at least in part, related to increased production of superoxide anions by NADH/NADPH oxidase.

Occipital Artery Function during the Development of 2-Kidney, 1-Clip Hypertension in Rats.

This study compared the contractile responses elicited by angiotensin II (AII), arginine vasopressin (AVP), and 5-hydroxytryptamine (5-HT) in isolated occipital arteries (OAs) from sham-operated (SHAM) and 2-kidney, 1-clip (2K-1C) hypertensive rats. OAs were isolated and bisected into proximal segments (closer to the common carotid artery) and distal segments (closer to the nodose ganglion) and mounted separately on myographs. On day 9, 2K-1C rats had higher mean arterial blood pressures, heart rates, and plasma renin concentrations than SHAM rats. The contractile responses to AII were markedly diminished in both proximal and distal segments of OAs from 2K-1C rats as compared to those from SHAM rats. The responses elicited by AVP were substantially greater in distal than in proximal segments of OAs from SHAM rats and that AVP elicited similar responses in OA segments from 2K-1C rats. The responses elicited by 5-HT were similar in proximal and distal segments from SHAM and 2K-1C rats. These results demonstrate that continued exposure to circulating AII and AVP in 2K-1C rats reduces the contractile efficacy of AII but not AVP or 5-HT. The diminished responsiveness to AII may alter the physiological status of OAs in vivo.

Role of renal nerves in the treatment of renovascular hypertensive rats with L-arginine.

The purpose was to determine the role of renal nerves in mediating the effects of antihypertensive treatment with L-arginine in a renovascular hypertension model. The 2K1C (two-kidney one-clip model) hypertensive rats were submitted to bilateral surgical-pharmacological renal denervation. The animals were subdivided into six experimental groups: normotensive control rats (SHAM), 2K1C rats, 2K1C rats treated with L-arginine (2K1C + L-arg), denervated normotensive (DN) rats, denervated 2K1C (2K1C + DN) rats, and denervated 2K1C + L-arg (2K1C + DN + L-arg) rats. Arterial blood pressure, water intake, urine volume, and sodium excretion were measured. The 2K1C rats exhibited an increase in the mean arterial pressure (MAP) (from 106 ± 3 to 183 ± 5.8 mmHg, P < 0.01), whereas L-arg treatment induced a reduction in the MAP (143 ± 3.4 mmHg) without lowering it to the control level. Renal nerve denervation reduced the MAP to normotensive levels in 2K1C rats with or without chronic L-arg treatment. L-arg and denervation induced increases in water intake and urine volume, and L-arg caused a significant natriuretic effect. Our results suggest that renal sympathetic activity participates in the genesis and the maintenance of the hypertension and also demonstrate that treatment with L-arg alone is incapable of normalizing the MAP and that the effect of such treatment is not additive with the effect of kidney denervation.

Participation of 5-HT and AT1Receptors within the Rostral Ventrolateral Medulla in the Maintenance of Hypertension in the Goldblatt 1 Kidney-1 Clip Model

The hypothesis that changes in neurotransmission within the rostral ventrolateral medulla (RVLM) are important to maintain the high blood pressure (BP) was tested in Goldblatt one kidney-one clip hypertension model (1K-1C). Male Wistar rats were anesthetized (urethane 1.2 g/kg, i.v.), and the effects of bilateral microinjections into the RVLM of the following drugs were measured in 1K-1C or control groups: glutamate (0.1 mol/L, 100 nL) and its antagonist kynurenic acid (0.02 mol/L, 100 nL), the angiotensin AT1 receptor antagonist candesartan (0.01 mol/L, 100 nL), and the nonselective 5-HT receptor antagonist methiothepin (0.06 mol/L, 100 nL). Experiments in 1K-1C rats were performed 6 weeks after surgery. In anesthetized rats glutamate response was larger in hypertensive than in normotensive rats (H: Δ67 ± 6.5; N: Δ43 ± 3.54 mmHg). In contrast, kynurenic acid microinjection into the RVLM did not cause any change in BP in either group. The blockade of either AT1 or 5-HT receptors within the RVLM decreased BP only in 1K-1C rats. A largest depressor response was caused by 5-HT receptor blockade. The data suggest that 5-HT and AT1 receptors act tonically to drive RVLM in 1K-1C rats, and these actions within RVLM contribute to the pathogenesis of this model of hypertension.

Distinct protein signature of hypertension-induced damage in the renal proteome of the two-kidney, one-clip rat model.

Hypertensive nephrosclerosis is one of the most frequent causes of chronic kidney failure. Proteome analysis potentially improves the pathophysiological understanding and diagnostic precision of this disorder. In the present exploratory study, we investigated experimental nephrosclerosis in the two-kidney, one-clip (2K1C) hypertensive rat model. The renal cortex proteome from juxtamedullary cortex and outer cortex of 2K1C male Wistar-Hannover rats (n = 4) was compared with the sham-operated controls (n = 6), using mass spectrometry-based quantitative proteomics. We combined a high abundant plasma protein depletion strategy with an extended liquid chromatographic gradient to improve peptide and protein identification. Immunohistology was used for independent confirmation of abundance. We identified 1724 proteins, of which 1434 were quantified with at least two unique peptides. Comparative proteomics revealed 608 proteins, including the platelet-derived growth factor receptor-β signalling pathway, with different abundances between the non-clipped kidney of hypertensive 2K1C rats and the corresponding kidney of the normotensive controls (P < 0.05, absolute fold change ≥1.5). Among the most significantly altered proteins in the whole cortex were periostin, transgelin, and creatine kinase B-type. Relative abundance of periostin alone allowed clear classification of 2K1C and controls. Enrichment of periostin in 2K1C rats was verified by immunohistology, showing positivity especially around the fibrotic vessels. The proteome is altered in hypertension-induced kidney damage. We propose periostin, especially in combination with transgelin and creatine kinase B-type, as possible proteomic classifier to distinguish hypertensive nephrosclerosis from the normal tissue. This classifier needs to be further validated with respect to early diagnosis of fibrosis, prognosis, and its potential as a novel molecular target for pharmacological interventions.

Morphine and nicotine addiction and withdrawal influence baroreflex sensitivity and blood pressure in two-kidney one clip hypertensive (2K1C) rats.

This study aimed to investigate the effects of addiction to morphine and nicotine as well as their withdrawal on both baroreflex sensitivity and blood pressure in hypertensive rats. In this experimental study 40 male rats were divided into two main groups as follows: in group I, hypertensive rats received saline for 8 weeks; in group II, hypertensive rats were treated with morphine and nicotine for 8 weeks. At the end of 8 weeks group II rats were divided into four sub-groups including, 3 sub-groups of those were put on drug withdrawal protocol. At the end of experiment, blood pressure, heart rate, plasma renin activity (PRA), serum NO concentration and baroreflex sensitivity (BRS) were measured. RESULTS demonstrated that BP and BRS were significantly lower in addicted to morphine and nicotine hypertensive rats compared to control (p < 0.05). Addiction withdrawal (in morphine and nicotine withdrawal rats) completely reversed BP and BRS to the pre-addiction levels (p < 0.05). Withdrawal in the only nicotine treated group lowered BP and BRS compared to group that had received morphine and nicotine together (p < 0.05). RESULTS of current study may propose simultaneous morphine and nicotine withdrawal can prevent cardiovascular complications raised due to withdrawal (Fig. 5, Ref. 58).

Resveratrol Reverses the Impaired Vasodilation Observed in 2K-1C Hypertension through Endothelial Function Improvement

Background: The production of endothelial-derived factors induces either vasoconstriction or vasodilation; nitric oxide (NO) is the most distinguished relaxing factor. Endothelial dysfunction is associated with hypertension. The partial loss in the NO-promoted vasodilation is due to its decreased bioavailability and/or to an activity reduction of endothelium NO synthase (eNOS). Reactive oxygen species (ROS), present in oxidative stress, seize NO and diminish its bioavailability. Transresveratrol (RESV) has been proved to increase NO and eNOS levels. Thus, RESV could be capable of improving NO dependent vascular relaxation on aortic rings isolated from treated 2K-1C animals through ROS damage reduction. Aim: Evaluate the effects of RESV treatment on the relaxation of aortic rings isolated from treated 2K-1C rats while focusing on the effects of the treatment on systolic blood pressure. Methods: Male Wistar rats (180 g) were grouped: two 2K-1C and two Sham groups, one of each was treated with RESV (20 mg/kg, gavage) dissolved in Tween 80 and one of each was treated with water plus Tween 80 (control) for six weeks. The rats had their systolic blood pressure (SBP) measured before and after the treatments. Vascular reactivity studies were conducted in order to observe and compare acetylcholine (ACh)-induced relaxations in the presence and absence of the NOS inhibitor L-NAME (10-4 mol/L). Results: SBP for 2K-1C was significantly reduced in the treated group (179.13 ± 4.90 mmHg, n = 23) when compared to the untreated group (196.66 ± 6.06 mmHg, n = 15, p

Superoxide anions involved in sympathoexcitation and pressor effects of salusin‐β in paraventricular nucleus in hypertensive rats

Salusin-β in paraventricular nucleus (PVN) increases renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP), heart rate (HR) and arginine vasopressin (AVP) release in hypertensive rats but not in normal rats. The present study was designed to investigate the downstream molecular mechanism of salusin-β in the PVN in hypertension. Renovascular hypertension was induced by two-kidney, one-clip (2K1C) in male SD rats. Acute experiments were carried out 4 weeks after 2K1C or sham operation under anaesthesia. MrgA1 mRNA expression and salusin-β level in the PVN as well as plasma salusin-β level were increased in 2K1C rats. Bilateral PVN microinjection of salusin-β increased the RSNA, MAP and HR in 2K1C rats, which were abolished by the pre-treatment with polyethylene glycol-superoxide dismutase (PEG-SOD), the superoxide anion scavenger tempol, the NAD(P)H oxidase inhibitor apocynin or the protein kinase C (PKC) inhibitor chelerythrine chloride (CLC), but not affected by the AT1 receptor antagonist losartan, the Mas receptor antagonist A-779, the NOS inhibitor L-NAME or the GABAA and GABAB receptor antagonists gabazine+CGP-35348. Salusin-β-induced increases in superoxide anion level and NAD(P)H oxidase activity in the PVN were abolished by the PVN pre-treatment with CLC. Salusin-β increased AVP levels in rostral ventrolateral medulla and plasma, which were prevented by the pre-treatment with PEG-SOD, apocynin or CLC in 2K1C rats. Salusin-β augmented the enhanced activity of PKC in the PVN in 2K1C rats. Protein kinase C-NAD(P)H oxidase-superoxide anions pathway in the PVN is involved in salusin-β-induced sympathetic activation, pressor response and AVP release in renovascular hypertension.

Intermedin in Paraventricular Nucleus Attenuates Sympathetic Activity and Blood Pressure via Nitric Oxide in Hypertensive Rats

Intermedin (IMD) is a member of calcitonin/calcitonin gene-related peptide family, which shares the receptor system consisting of calcitonin receptor-like receptor (CRLR) and receptor activity-modifying proteins (RAMPs). This study investigated the effects of IMD in paraventricular nucleus (PVN) on renal sympathetic nerve activity and mean arterial pressure and its downstream mechanism in hypertension. Rats were subjected to 2-kidney 1-clip (2K1C) surgery to induce renovascular hypertension or sham operation. Acute experiments were performed 4 weeks later under anesthesia. IMD mRNA and protein were downregulated in 2K1C rats. Bilateral PVN microinjection of IMD caused greater decreases in renal sympathetic nerve activity and mean arterial pressure in 2K1C rats than in sham-operated rats, which were prevented by pretreatment with adrenomedullin receptor antagonist AM22-52 or nonselective nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine methyl ester, and attenuated by selective neuronal NO synthase inhibitor N(ω)-propyl-l-arginine hydrochloride or endothelial NO synthase inhibitor N(5)-(1-iminoethyl)-l-ornithine dihydrochloride. AM22-52 increased renal sympathetic nerve activity and mean arterial pressure in 2K1C rats but not in sham-operated rats, whereas calcitonin/calcitonin gene-related peptide receptor antagonist calcitonin/calcitonin gene-related peptide 8-37 had no significant effect. CRLR and RAMP3 mRNA, as well as CRLR, RAMP2, and RAMP3 protein expressions, in the PVN were increased in 2K1C rats. Microinjection of IMD into the PVN increased the NO metabolites (NOx) level in the PVN in 2K1C rats, which was prevented by AM22-52. Chronic PVN infusion of IMD reduced, but AM22-52 increased, blood pressure in conscious 2K1C rats. These results indicate that IMD in the PVN inhibits sympathetic activity and attenuates hypertension in 2K1C rats, which are mediated by adrenomedullin receptors (CRLR/RAMP2 or CRLR/RAMP3) and its downstream NO.

Mesenchymal Stem Cells (MSC) Prevented the Progression of Renovascular Hypertension, Improved Renal Function and Architecture

Renovascular hypertension induced by 2 Kidney-1 Clip (2K-1C) is a renin-angiotensin-system (RAS)-dependent model, leading to renal vascular rarefaction and renal failure. RAS inhibitors are not able to reduce arterial pressure (AP) and/or preserve the renal function, and thus, alternative therapies are needed. Three weeks after left renal artery occlusion, fluorescently tagged mesenchymal stem cells (MSC) (2×105 cells/animal) were injected weekly into the tail vein in 2K-1C hypertensive rats. Flow cytometry showed labeled MSC in the cortex and medulla of the clipped kidney. MSC prevented a further increase in the AP, significantly reduced proteinuria and decreased sympathetic hyperactivity in 2K-1C rats. Renal function parameters were unchanged, except for an increase in urinary volume observed in 2K-1C rats, which was not corrected by MSC. The treatment improved the morphology and decreased the fibrotic areas in the clipped kidney and also significantly reduced renal vascular rarefaction typical of 2K-1C model. Expression levels of IL-1β, TNF-α angiotensinogen, ACE, and Ang II receptor AT1 were elevated, whereas AT2 levels were decreased in the medulla of the clipped kidney. MSC normalized these expression levels. In conclusion, MSC therapy in the 2K-1C model (i) prevented the progressive increase of AP, (ii) improved renal morphology and microvascular rarefaction, (iii) reduced fibrosis, proteinuria and inflammatory cytokines, (iv) suppressed the intrarenal RAS, iv) decreased sympathetic hyperactivity in anesthetized animals and v) MSC were detected at the CNS suggesting that the cells crossed the blood-brain barrier. This therapy may be a promising strategy to treat renovascular hypertension and its renal consequences in the near future.

Effects of telmisartan and linagliptin when used in combination on blood pressure and oxidative stress in rats with 2-kidney-1-clip hypertension

To investigate the effects of linagliptin alone and in combination with the angiotensin II receptor blocker (ARB), telmisartan on blood pressure (BP), kidney function, heart morphology and oxidative stress in rats with renovascular hypertension. Fifty-seven male Wistar rats underwent unilateral surgical stenosis of the renal artery [2-kidney-1-clip (2k1c) method]. Animals were randomly divided into four treatment groups (n = 14-18 per group) receiving: telmisartan (10 mg/kg per day in drinking water), linagliptin (89 ppm in chow), combination (linagliptin 89 ppm + telmisartan 10 mg/kg per day) or placebo. An additional group of 12 rats underwent sham surgery. BP was measured one week after surgery. Hypertensive animals entered a 16-week dosing period. BP was measured 2, 4, 8, 12 and 16 weeks after the initiation of treatment. Blood and urine were tested for assessment of kidney function and oxidative stress 6, 10, 14 and 18 weeks after surgery. Blood and urine sampling and organ harvesting were finally performed. Renal stenosis caused an increase in mean ± SD systolic BP as compared with the sham group (157.7 ± 29.3 vs. 106.2 ± 20.5 mmHg, respectively; P < 0.001). Telmisartan alone and in combination with linagliptin, normalized SBP (111.1 ± 24.3 mmHg and 100.4 ± 13.9 mmHg, respectively; P < 0.001 vs. placebo). Telmisartan alone and in combination with linagliptin significantly prevented cardiac hypertrophy, measured by heart weight and myocyte diameter. Renal function measured by cystatin C was not affected by 2k1c surgery. Telmisartan significantly increased plasma concentration of cystatin C. 2k1c surgery initiated fibrosis in both kidneys. Telmisartan promoted further fibrotic changes in the clipped kidney, as measured by protein expression of Col1a1 and histology for interstitial fibrosis and glomerulosclerosis. In non-clipped kidneys, telmisartan demonstrated antifibrotic properties, reducing Col1a1 protein expression. Plasma levels of oxidized low-density lipoprotein were higher in the placebo-treated 2k1c rats as compared to sham-operated animals. The increase was abolished by linagliptin alone (P = 0.03 vs. placebo) and in combination with telmisartan (P = 0.02 vs. placebo). Combination therapy also significantly reduced plasma concentration of carbonyl proteins (P = 0.04 vs. placebo). Inhibition of type 4 dipeptidyl peptidase with linagliptin did not counter BP-lowering effects of ARB in 2k1c rats. Linagliptin reduced lipid and protein oxidation in 2k1c rats, and this effect was BP-independent.

Propranolol does not increase the contractile response to epinephrine in endothelium-intact aorta from renal hypertensive rat

Renal hypertension (2K-1C) is characterized by the increased sympathetic activity and oxodative stress induced by angiotensin II. Therefore, the epinephrine plasma levels can potentiate the contractile response in 2K-1C rat aorta. This study aimed to evaluate the effect of inhibition of β-adrenoceptor (β-AR) activation on the contractile response induced by epinephrine in intact endothelium (E+) and in denuded aorta (E−) of 2K-1C rats compared to normotensive sham-operated rats (2K). All the procedures were performed in accordance with the standards and policies of Animal Care and Use Committee of the University of São Paulo. Male rats (180–200 g) were anesthetized with tribromoethanol and after a midline laparotomy a silver clip was placed around the left renal artery. Normotensive two-kidney rats (2K) were only submitted to laparotomy. The systolic blood pressure (SBP) was measured by an indirect tail-cuff method 6 weeks after surgery. Rats were considered to be hypertensive when systolic arterial pressure was higher than 160 mm Hg. Rats were killed and the aortic rings were isolated. Concentration–effect curves were constructed for epinephrine in E+ and E− aortas in the absence (Control) or after incubation with 10 μM propranolol (P+) for 20 min. The maximal effect (ME, in grams of tension) and the potency (pD2) were evaluated. Propranolol did not change the contractile response induced by epinephrine in aortas E− from 2K rats (Control, ME: 2.9 ± 0.2 g; pD2: 7.74 ± 0.09; n = 9) and P+ (ME: 3.7 ± 0.3 g; pD2: 7.48 ± 0.19; n = 4) and 2K-1C (Control, ME: 2.1 ± 0.1 g; pD2: 7.92 ± 0.10; n = 7) and P+ (ME: 2.2 ± 0.2 g; pD2: 8.44 ± 0.78; n = 5). In E+ aorta, propranolol increased the ME to epinephrine in 2K rat aortas from 2.6 ± 01 g; n = 11 to 3.1 ± 0.2 g; n = 6) (P <0.01) and it reduced the pD2 values in 2K-1C rat aortas from 7.37 ± 0.16; n = 9 to 6.70 ± 0.22; n = 4) (P <0.05). Our results indicate that the contractile response induced by epinephrine involves the activation of β-AR in the endothelial cells of 2K rat aortas that leads to a lower contractile responses of 2K rat aorta but not in 2K-1C hypertensive rat aorta. Supported by FAPESP and CNPq .

Hydrogen peroxide modulates phenylephrine-induced contractile response in renal hypertensive rat aorta

Endothelium-derived factors play an important role in vascular tone control. This study aimed to evaluate how endothelium and reactive oxygen species (ROS) contribute to phenylephrine (PE)-induced contraction in renovascular hypertensive (2K–1C) and normotensive (2K) rats aortas. The effects of the superoxide scavenger Tiron (0.1mM and 1mM) or catalase (30U/ml, 90U/ml, 150U/ml and 300U/ml) on the PE-induced contraction were evaluated in both intact endothelium (E+) and denuded (E−) aortas. Endothelium removal increased the PE-induced contractions. The maximum contractile response decreased only in 2K–1C rat E+ aorta, and catalase (30U/ml, 90U/ml, 150U/ml) partially reversed this effect. Endothelium increased the basal hydrogen peroxide (H2O2) production in 2K and 2K–1C rats aortas. PE-stimulated H2O2 production was higher in 2K–1C (E+/E−) than in 2K (E+/E−). Inhibition of the enzymes cyclooxygenase, NADPH-oxidase, xanthine-oxidase, and superoxide dismutase reduced the PE-stimulated H2O2 production in 2K–1C rat aorta. The decreased contraction to PE in 2K–1C rat aorta is partially due to endothelial H2O2 production; however, in denuded aorta, it contributes to maintaining the contractile response. Superoxide plays an important role on the PE-induced contraction in 2K rat denuded aorta, whereas in 2K–1C rat aorta, it is H2O2 that plays an important role in this effect.

Abstract 524: Vinegar Enhanced the Anti-hypertensive Effect of Saccharina Japonica Intake in Rats With 2-kidney, 1-clip Renovascular Hypertension

Objective: Saccharina japonica (SJ) is one of brown algae and is cultivated or grows wild in Japan and neighbor countries. SJ is rich in alginate, the intake of which was reported to decrease blood pressure (BP) in spontaneously hypertensive rats. It is also rich in fucoidan, which was reported to improve the renal blood flow in rats with acute renal failure. We recently observed that intake of SJ decreased BP in 2-kidney, 1-clip renovascular hypertensive (2K1C) rats, probably through nitric oxide, and did not change BP in normotensive rats. In Japan, there are many traditional processed foods made of SJ with vinegar, which is originally used for softening SJ and raising its preserving property. Vinegar is also suggested to suppress angiotensin-converting enzyme activity. In this study, we investigated the interactive effects of SJ and vinegar on BP in 2K1C rats. Methods: Male Sprague-Dawley rats (6 wks) were treated with sham operation (SHAM) or clipping the left renal artery (2K1C). After surgery, SHAM or 2K1C rats started receiving a control diet (CTL) or a diet containing 5.0% (w/w) freeze-dried SJ powder (SJ) for 6 weeks, and a tap water (Veh) or a water containing 5.0% (v/v) of 4.3% acidity rice vinegar (VG). The systolic BP (SBP) was measured by a tail-cuff method every week. At the end of the protocol, mean arterial BP (MAP) was measured in each rat under anesthesia. Then, thoracic aorta was removed and the section of aorta was stained with hematoxylin and eosin. Results: Analysis of variance revealed significant effects in time, animal model (SHAM vs 2K1C), diet (CTL vs SJ) and model x diet ( P &lt; 0.001, each), but not in water (Veh vs VG) or model x water. At the end of the protocol, in CTL-Veh, 2K1C was higher in SBP than SHAM (179 ± 33 vs 130 ± 20 mmHg, P &lt; 0.001). In 2K1C-Veh, the SJ diet decreased SBP (156 ± 6 mmHg) compared with CTL ( P &lt; 0.05). Vinegar did not significantly change SBP (184 ± 19 mmHg) in 2K1C-CTL, but decreased SBP (142 ± 11 mmHg) in 2K1C-SJ ( P &lt; 0.05) as low as in SHAM-SJ-VG (139 ± 3 mmHg, P = NS). The observations in MAP were similar to in the SBP. Moreover, compared to SHAM, 2K1C-CTL-Veh showed the aortic media thickening, which was reduced in 2K1C-SJ-Veh/VG. Conclusion: A certain daily intake of vinegar did not lower BP by itself, but enhanced the BP reduction by a SJ diet, in 2K1C rats.

Abstract 622: Role of Endothelial Nitric Oxide Synthase in a Capsaicin Diet-Induced Alleviation of Blood Pressure-Elevation in 2-Kidney, 1-Clip Hypertensive Rats

Objective: Capsaicin directly acts on the sensory nerve ending and releases neuropeptides, which activate a vasodilator, nitric oxide (NO) and decrease blood pressure (BP). We demonstrated that BP elevation was alleviated by consuming a low concentration of capsaicin diet in 2-kidney, 1-clip (2K1C) hypertensive rats. We also observed that when 2K1C rats took the NO synthase (NOS) inhibitor, NG-nitro-L-arginine methyl ester, during the protocol, the effect of alleviating hypertension by dietary capsaicin was diminished. Thus, we hypothesized that NO is involved in the effect of capsaicin in 2K1C rats. In this study, to test the hypothesis, we observed endothelial NOS (eNOS) expression in 2K1C rats fed a diet containing capsaicin. Methods: Six-week old male Sprague-Dawley rats were treated with sham operation (SHAM) or clipping the left renal artery (2K1C). One week after the surgery, each group of rats were further divided into 2 groups randomly, which received either a control diet (CTL) or a diet containing 0.006% capsaicin (CAP) for 6 weeks. The systolic BP (SBP) was measured by a tail-cuff method once per week throughout the protocol. At the end of the protocol, MAP was measured in each rat under anesthesia. After euthanasia, the thoracic aortas were collected for extracting mRNA. Then, eNOS mRNA expression in aorta was evaluated in each group of rats by reverse transcriptase PCR. Results and Discussion: The present data of BP confirmed that capsaicin diet alleviates BP elevation in 2K1C rats. At six weeks after the surgery, SBP in 2K1C-CTL rats (183 ± 6 mmHg) was significantly higher than SHAM-CTL (124 ± 6 mmHg, p &lt; 0.05), but SBP in 2K1C-CAP (140 ± 4 mmHg) was significantly lower than that in 2K1C-CTL. SHAM-CAP (129 ± 8 mmHg) did not show a significant difference in SBP with SHAM-CTL. MAP at the end of the protocol showed the same tendency as SBP. Moreover, eNOS mRNA expression was enhanced in 2K1C-CTL (0.39 ± 0.06) compared to SHAM-CTL (0.31 ± 0.04, p &lt; 0.05). Although it was not enhanced in SHAM-CAP (0.30 ± 0.01) compared to SHAM-CTL, it was tended to be enhanced in 2K1C-CAP (0.43 ± 0.02) compared to 2K1C-CTL. These data suggested that dietary capsaicin decreased BP with elevation of eNOS mRNA expression in 2K1C rats, but did not change eNOS mRNA nor BP in SHAM normotensive rats.

Abstract 645: Atorvastatin and Sildenafil Reduce Oxidative Stress, MMP-2 And MMP-9 Levels, TGF-beta Expression and Prevent Vascular Remodeling in 2K1C-hypertension

Hypertension-induced vascular remodeling is associated with oxidative stress, and matrix metalloproteinase (MMP) and TGF (transforming growth factor)-beta up-regulation. Atorvastatin (ATORVA) and sildenafil (SILD) exert pleiotropic effects that may result in cardiovascular protection. We evaluated the effects of ATORVA and SILD alone or in association on oxidative stress, MMPs and TGF-beta up-regulation, and vascular hypertrophy induced by 2 kidney-1 clip (2K1C) hypertension. Sham and 2K1C rats were orally treated with vehicle, ATORVA (50 mg/kg), SILD (45 mg/kg) or both for 8 weeks. ATORVA, SILD, or both drugs exerted antihypertensive effects (systolic blood pressure: 148±7, 156±5, and 138±4 mmHg, respectively, vs. 207±4 mmHg in 2K1C untreated rats; P&lt;0.05). All treatments prevented the increases in the aortic cross-sectional area and media/lumen ratio in 2K1C rats (P&lt;0.05). Hypertension increased NADPH oxidase activity when compared with control groups and the treatments reduced oxidative stress. Aortas from 2K1C rats showed higher MMP-2 levels when compared with sham (0.399±0.06 vs. 0.173±0.05 A.U.) and all treatments attenuated 2K1C hypertension-induced increases in MMP-2 levels (ATORVA: 0.191±0.04; SILD: 0.136±0.05; ATORVA+SILD: 0.173±0.03; P&lt;0.05). However, these drugs had no in vitro effects on human recombinant MMP-2 activity (P&gt;0.05). Moreover, 2K1C rats showed higher MMP-9 (12.64±0.86 vs. 8.70±0.92) and TGF-beta expression (14.25±0.97 vs. 9.07±1.67) compared with sham groups. All treatments prevented 2K1C hypertension-induced increases in these pro-fibrotic factors. Treatment with ATORVA or SILD, or both, exerted antihypertensive effects and prevented 2K1C hypertension-induced vascular remodeling through antioxidant and anti-fibrotic mechanisms. Our results suggest that ATORVA and SILD may prevent the vascular alterations of hypertension.

Abstract 55: PPAR-δ Binding to Heme Oxygenase 1 Promoter Prevents Angiotensin II Induced Vascular and Adipocyte Dysfunction in a Model of Renovascular Hypertension

Introduction: Renin-Angiotensin System (RAS) induces oxidative stress and contributes to various pathological conditions including insulin resistance and the metabolic syndrome. Recent studies have shown the role of PPARδ-agonist in attenuation of Angiotensin II induced oxidative stress.The aim of this study was to explore the effectiveness of a PPARδ-agonist in the prevention of Ang II-induced vascular and adipocyte dysfunction and the possible interaction between PPARδ and HO-1 system in a model of enhanced oxidative stress, the Goldblatt 2K1C model. Methods: We first established a direct stimulatory effect of the PPARδ-agonist (GW 501516) on the HO-1 gene by demonstrating increased luciferase activity in COS-7 cells transfected with a luciferase-HO-1 promoter construct. SD rats were divided in 4 groups: sham operated animals, 2K1C rats and 2K1C rats treated with GW 501516, in the absence or presence of the HO activity inhibitor, stannous mesoporphyrin (SnMP). Results: 2K1C animals had increased blood pressure, visceral adiposity, adipocyte hypertrophy, increased inflamatory cytokines, increased circulatory and adipose tisssue levels of renin and Ang II along with increased adipose tissue gp91 phox expression (p&lt;0.05) when compared to sham operated animals. Treatment with GW 501516 increased adipose tissue HO-1 and adiponectin levels (p&lt;0.01) along with enhancement of Wnt10b and β-catenin expression. HO-1 induction was accompanied by the decreased expression of Wnt 5b, Mest and C/EBPα levels and an increased number of small adipocytes (p&lt;0.05). These effects of GW501516 were reversed in 2K1C animals exposed to SnMP (p&lt;0.05). Conclusion: This novel study corroborates the existence of a PPARδ-HO-1 axis whose direct stimulation via the administration of an exogenous PPARδ-agonist induces HO-1 expression, abates RAS associated adipocyte and vascular dysfunction and induces adipocyte remodeling with smaller adipocytes while decreasing inflammation and increasing adiponectin secretion.

Effect of co-administration of morphine and nicotine on cardiovascular function in two-kidney one clip hypertensive (2K1C) rats

Cardiovascular morbidity and mortality are potentiated with smoking and hypertension. The aim of this study was to investigate the effects of morphine and nicotine co-administration on cardiovascular function in two-kidney one-clip hypertensive (2K1C) rats. Thirty-two male rats were divided into four groups as follow: Vehicle, morphine, nicotine and nicotine + morphine. All drugs were administered for 8 weeks. Baroreflex sensitivity (BRS), heart rate and blood pressure were measured using a Power Lab data acquisition. Plasma rennin activity (PRA) and serum concentration of nitric oxide (NO) were measured using Elisa method. To induce hypertension, the renal artery of left kidney was clipped for 8 weeks. A significant decrease in systolic blood pressure (SBP), diastolic blood pressure (DBP) and mean arterial pressure (MAP) was observed in nicotine + morphine group compared to vehicle and nicotine groups (p<0.05). Serum concentration of NO was lower in nicotine + morphine group compared to morphine group and significantly higher than nicotine group. The BRS was lower in the nicotine + morphine group compared to other groups. The PRA level was higher in nicotine + morphine compared to morphine group but it was higher than nicotine group. This study demonstrated that prolonged co-consumption of morphine and nicotinedecreased PRA and blood pressure and increased the serum concentration of NO in hypertensive rats. Co-administration of morphine and nicotine decreased BRS in 2k1c hypertensive rats probably via central nervous system.

AT1 Receptors Prevent Salt-Induced Vascular Dysfunction in Isolated Middle Cerebral Arteries of 2 Kidney-1 Clip Hypertensive Rats

Elevated blood pressure, elevated angiotensin II (ANG II), and ANG II suppression with high salt (HS) diet all contribute to vascular dysfunction. This study investigated the interplay of HS diet and vascular function in a high renin model of hypertension. Male Sprague-Dawley rats were subjected to 2 kidney-1 clip (2K1C) Goldblatt hypertension for 4 weeks and compared with sham-operated controls. Middle cerebral arteries (MCA) of 2K1C rats and sham-operated controls fed normal salt (NS; 0.4% NaCl) diet dilated in response to acetylcholine (ACh) and reduced partial pressure of oxygen (PO2). Switching to HS (4% NaCl) diet for 3 days to reduce plasma renin activity (PRA) eliminated vasodilation to ACh and reduced PO2 in sham-operated controls, with no effect on vasodilation in 2K1C rats. AT1 receptor blockade (losartan, 20 mg/kg/day; 1 week) eliminated vasodilator responses to ACh and reduced PO2 in 2K1C rats fed NS or HS diet. ANG II infusion (5 ng/kg/min, intravenous) for 3 days to prevent salt-induced reductions in plasma ANG II restored vascular relaxation in MCA of sham-operated controls fed HS diet. Copper/zinc superoxide dismutase expression and total superoxide dismutase activity were significantly higher in arteries of 2K1C rats fed HS diet vs. sham-operated controls. These results suggest that the sustained effects of elevated ANG II levels in 2K1C hypertension maintain endothelium-dependent vasodilatation via AT1 receptor-mediated preservation of antioxidant defense mechanisms despite significant elevations in blood pressure and salt-induced suppression of PRA.

Sodium nitroprusside activates potassium channels in the vena cava in normotensive but not in hypertensive rats

Despite the importance of the venous system in the regulation of blood pressure, there are few studies that evaluate venous function in health and disease, and the effects of drugs on venous function. Blood pressure depends directly on the peripheral resistance and cardiac output. Unlike the peripheral resistance, in which the contractile activity of the arteries is the key factor, cardiac output depends primarily on the venomotor tone. An increase in cardiac blood pressure can be caused by an increase in blood volume, structural changes in the walls of the veins, leading to a reduced compliance thereof, or an increase in the contractile activity of venous smooth muscle. This study examined the effect of sodium nitroprusside (SNP), a classical nitric oxide donor, on vascular relaxation in the vena cava from normotensive (2K) and renal hypertensive (2K-1C) rats. We studied the effect of this compound in vena cava rings from normotensive and renal hypertensive rats. We showed for the first time that the vascular relaxation induced by SNP is impaired in vena cavas from hypertensive rats because of an impaired functional activity of potassium channels. Another relevant finding of this study is that the sarcoplasmic reticulum Ca(2+) ATPase is not involved in the venorelaxation induced by SNP.