Blood Pressure In Spontaneously Hypertensive Rats Research Articles

Cytochrome P450 1B1 contributes to increased blood pressure and cardiovascular and renal dysfunction in spontaneously hypertensive rats

We investigated the contribution of cytochrome P450 (CYP) 1B1 to hypertension and its pathogenesis by examining the effect of its selective inhibitor, 2,4,3',5'-tetramethoxystilbene (TMS), in spontaneously hypertensive rats (SHR). Blood pressure (BP) was measured bi-weekly. Starting at 8 weeks, TMS (600μg/kg, i.p.) or its vehicle was injected daily. At 14 weeks, samples were collected for measurement. TMS reversed increased BP in SHR (207 ± 7 vs. 129 ± 2mmHg) without altering BP in Wistar-Kyoto rats. Increased CYP1B1 activity in SHR was inhibited by TMS (RLU: aorta, 5.4 ± 0.7 vs. 3.7 ± 0.7; heart, 6.0 ± 0.8 vs. 3.4 ± 0.4; kidney, 411 ± 45 vs. 246 ± 10). Increased vascular reactivity, cardiovascular hypertrophy, endothelial and renal dysfunction, cardiac and renal fibrosis in SHR were minimized by TMS. Increased production of reactive oxygen species and NADPH oxidase activity in SHR, were diminished by TMS. In SHR, TMS reduced increased plasma levels of nitrite/nitrate (46.4 ± 5.0 vs. 28.1 ± 4.1μM), hydrogen-peroxide (36.0 ± 3.7 vs. 14.1 ± 3.8μM), and thiobarbituric acid reactive substances (6.9 ± 1.0 vs. 3.4 ± 1.5μM). Increased plasma levels of pro-inflammatory cytokines and catecholamines, and cardiac activity of extracellular signal-regulated kinase, p38 mitogen-activated protein kinase, c-Src tyrosine kinase, and protein kinase B in SHR were also inhibited by TMS. These data suggests that increased oxidative stress generated by CYP1B1 contributes to hypertension, increased cytokine production and sympathetic activity, and associated pathophysiological changes in SHR. CYP1B1 could be a novel target for developing drugs to treat hypertension and its pathogenesis.

Duration of Electrically Induced Atrial Fibrillation Is Augmented by High Voltage of Stimulus with Higher Blood Pressure in Hypertensive Rats

Objective. Many previous clinical studies have suggested that atrial fibrillation (AF) is closely associated with hypertension. However, the benefits of antihypertensive therapy on AF are still inconsistent, and it is necessary to explore the factors augmenting AF in hypertensive rats. The aim of the present study was to investigate the correlation between arterial pressure or voltage stimulus and to the duration of electrically induced AF in normotensive or hypertensive rats. Methods. AF was reproducibly induced by transesophageal atrial burst pacing in spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY). We did the burst pacing at high (20 V) or low (5 V) voltage. Results. Duration of AF did not correlate with systolic blood pressure (SBP) and stimulus voltage in WKY. However, only in SHR, duration of AF with high stimulus voltage significantly correlated with SBP and was significantly longer in high than in low voltage stimulus. Discussion and Conclusion. Duration of AF is augmented by high voltage stimulus with higher blood pressure in SHR.

Orexin, cardio-respiratory function, and hypertension

In this review we focus on the role of orexin in cardio-respiratory functions and its potential link to hypertension. (1) Orexin, cardiovascular function, and hypertension. In normal rats, central administration of orexin can induce significant increases in arterial blood pressure (ABP) and sympathetic nerve activity (SNA), which can be blocked by orexin receptor antagonists. In spontaneously hypertensive rats (SHRs), antagonizing orexin receptors can significantly lower blood pressure under anesthetized or conscious conditions. (2) Orexin, respiratory function, and central chemoreception. The prepro-orexin knockout mouse has a significantly attenuated ventilatory CO2 chemoreflex, and in normal rats, central application of orexin stimulates breathing while blocking orexin receptors decreases the ventilatory CO2 chemoreflex. Interestingly, SHRs have a significantly increased ventilatory CO2 chemoreflex relative to normotensive WKY rats and blocking both orexin receptors can normalize this exaggerated response. (3) Orexin, central chemoreception, and hypertension. SHRs have higher ABP and SNA along with an enhanced ventilatory CO2 chemoreflex. Treating SHRs by blocking both orexin receptors with oral administration of an antagonist, almorexant (Almxt), can normalize the CO2 chemoreflex and significantly lower ABP and SNA. We interpret these results to suggest that the orexin system participates in the pathogenesis and maintenance of high blood pressure in SHRs, and the central chemoreflex may be a causal link to the increased SNA and ABP in SHRs. Modulation of the orexin system could be a potential target in treating some forms of hypertension.

Antihypertensive Effect of Syzygium cumini in Spontaneously Hypertensive Rats.

This study evaluated the in vivo potential antihypertensive effect of hydroalcoholic extract of Syzygium cumini leaves (HESC) in normotensive Wistar rats and in spontaneously hypertensive rats (SHR), as well as its in vitro effect on the vascular reactivity of resistance arteries. The hypotensive effect caused by intravenous infusion of HESC (0.01–4.0 mg/kg) in anesthetized Wistar rats was dose-dependent and was partially inhibited by pretreatment with atropine sulfate. SHR received HESC (0.5 g/kg/day), orally, for 8 weeks and mean arterial pressure, heart rate, and vascular reactivity were evaluated. Daily oral administration of HESC resulted in a time-dependent blood pressure reduction in SHR, with a maximum reduction of 62%. In the endothelium-deprived superior mesenteric arteries rings the treatment with HESC reduced by 40% the maximum effect (E max⁡) of contraction induced by NE. The contractile response to calcium and NE of endothelium-deprived mesenteric rings isolated from untreated SHR was reduced in a concentration-dependent manner by HESC (0.1, 0.25, and 0.5 mg/mL). This study demonstrated that Syzygium cumini reduces the blood pressure and heart rate of SHR and that this antihypertensive effect is probably due to the inhibition of arterial tone and extracellular calcium influx.

Genetic Variants on Rat Chromosome 8 Exhibit Profound Effects on Hypertension Severity and Survival During Nitric Oxide Inhibition in Spontaneously Hypertensive Rats

Hypertension and mortality is aggravated by nitric oxide inhibition with N(G)-nitro-L-arginine methyl ester (L-NAME) in spontaneously hypertensive rats (SHRs) but not in Munich Wistar Frömter (MWF) rats. MWF rats carry major albuminuria quantitative trait loci on rat chromosome (RNO) 6 and RNO8; susceptibility of SHRs to L-NAME is enhanced by transfer of RNO6 from MWF rats into the SHR background. Here, we tested whether the sensitivity to L-NAME in SHRs is affected by transfer of RNO8 from MWF rats in consomic SHR-8(MWF) rats. In study 1, we analyzed survival in male SHR and SHR-8(MWF) rats in response to 18 weeks of treatment with either normal drinking water (vehicle-treated) or water containing 20mg/L L-NAME. In study 2, we analyzed blood pressure and renal damage in both strains in response to 6 weeks of treatment with L-NAME compared with vehicle-treated groups. In study 1, starting after 6 weeks of treatment with L-NAME, mortality reached 90% in SHRs in contrast with the group of L-NAME treated SHR-8(MWF) rats (P < 0.0001) in which all rats survived similar to vehicle-treated rats. In study 2, L-NAME resulted in a more pronounced increase in mean arterial blood pressures in SHRs compared with SHR-8(MWF) rats (216 ± 6 vs. 180 ± 11 mm Hg; P < 0.05). In contrast, tubulointerstitial kidney damage was even lower in SHRs compared with SHR-8(MWF) rats after L-NAME treatment (P < 0.05), whereas albuminuria was not different between strains. The blood pressure increase and impaired survival of SHRs in response to nitric oxide inhibition is profoundly influenced by genes on RNO8.

Treating hypertension by targeting orexin receptors: potential effects on the sleep-related blood pressure dipping profile

Treating hypertension by targeting orexin receptors: potential effects on the sleep-related blood pressure dipping profile

Renoprotective Effects of Melatonin in Young Spontaneously Hypertensive Rats with L-NAME

Nitric oxide (NO) deficiency occurs in humans and animals with hypertension and chronic kidney disease (CKD). An inhibitor of NO synthase, N(G)-nitro-l-arginine methyl ester (L-NAME) exacerbates kidney damage in the adult spontaneously hypertensive rat (SHR). We examined whether L-NAME exacerbated hypertensive nephrosclerosis in young SHRs and whether melatonin protects SHRs against kidney damage by restoration of the asymmetric dimethylarginine (ADMA)-NO pathway. Rats aged 4 weeks were randomly assigned into three groups (n=10 for each group): Group 1 (control), SHRs without treatment; Group 2 (L-NAME), SHRs received L-NAME (80mg/L) in drinking water; and Group 3 (L-NAME+melatonin), SHRs received L-NAME (80mg/L) and 0.01% melatonin in drinking water. All rats were sacrificed at 10 weeks of age. L-NAME exacerbates the elevation of blood pressure, renal dysfunction, and glomerular sclerosis in young SHRs. L-NAME induced an increase of ADMA and a decrease of arginine-to-ADMA ratio in the SHR kidney. Melatonin therapy prevented L-NAME-exacerbated hypertension and nephrosclerosis in young SHRs. In addition, melatonin restored L-NAME-induced reduction of dimethylarginine dimethylaminohydrolase (DDAH; ADMA-metabolizing enzymes) activity in the SHR kidney. Next, melatonin decreased renal ADMA concentrations, increased renal arginine-to-ADMA ratio, and restored NO production in L-NAME-treated young SHRs. Moreover, melatonin reduced the degree of oxidative damaged DNA product, 8-hydroxydeoxyguanosine immunostaining in L-NAME-treated SHR kidney. Our results indicated that L-NAME/SHR is a useful model for hypertensive nephrosclerosis in young rats. The blood pressure-lowering and renoprotective effects of melatonin is due to increases of DDAH activity, decreases of ADMA, and reduction of oxidative stress in L-NAME-treated SHR kidney. Specific therapy targeting the DDAH-ADMA pathway may be a promising approach to slowing chronic kidney disease progression in children.

An orally active angiotensin-(1–7) inclusion compound and exercise training produce similar cardiovascular effects in spontaneously hypertensive rats

Low angiotensin-(1–7) (Ang-(1–7)) concentration is observed in some cardiovascular diseases and exercise training seems to restore its concentration in the heart. Recently, a novel formulation of an orally active Ang-(1–7) included in hydroxy-propyl-beta-cyclodextrin (HPB-CD) was developed and chronically administered in experimental models of cardiovascular diseases. The present study examined whether chronic administration of HPB-CD/Ang-(1–7) produces beneficial cardiovascular effects in spontaneously hypertensive rats (SHR), as well as to compare the results obtained with those produced by exercise training. Male SHR (15-week old) were divided in control (tap water) or treated with HPB-CD/Ang-(1–7) (corresponding to 30μgkg−1day−1 of Ang-(1–7)) by gavage, concomitantly or not to exercise training (treadmill, 10 weeks). After chronic treatment, hemodynamic, morphometric and molecular analysis in the heart were performed. Chronic HPB-CD/Ang-(1–7) decreased arterial blood pressure (BP) and heart rate in SHR. The inclusion compound significantly improved left ventricular (LV) end-diastolic pressure, restored the maximum and minimum derivatives (dP/dT) and decreased cardiac hypertrophy index in SHR. Chronic treatment improved autonomic control by attenuating sympathetic modulation on heart and vessels and the SAP variability, as well as increasing parasympathetic modulation and HR variability. Overall results were similar to those obtained with exercise training. These results show that chronic treatment with the HPB-CD/Ang-(1–7) inclusion compound produced beneficial effects in SHR resembling the ones produced by exercise training. This observation reinforces the potential cardiovascular therapeutic effect of this novel peptide formulation.

Renalase mRNA levels in the brain, heart, and kidneys of spontaneously hypertensive rats with moderate and high hypertension

BackgroundRenalase is a recently discovered secretory protein involved in regulation of arterial blood pressure in humans and animals. Results of animal experiments from independent laboratories indicate that administration of human recombinant renalase decreases blood pressure and some genetically predisposed hypertensive rats have lowered renalase levels.Material/MethodsThe levels of renalase mRNA expression in brain hemispheres, heart, and kidneys of spontaneously hypertensive rats (SHR) with moderate (140–180 mm Hg) or high (>180 mm Hg) hypertension and of control Wistar-Kyoto (WKY) rats were analyzed using real-time PCR.ResultsSpontaneously hypertensive rats with high hypertension (>180 mm Hg) had a lower renalase mRNA level in brain hemispheres, and higher heart and kidney renalase mRNA levels compared with control WKY rats. In SHR with a moderate increase in arterial blood pressure (140–180 mm Hg), the tissue renalase mRNA changed in the same direction but did not reach the level of statistical significance as compared with control rats.ConclusionsThe results indicate that the development of hypertension in SHR is accompanied by altered expression of the renalase gene in the examined organs as compared with control WKY rats. The brain and peripheral tissues renalase mRNA levels demonstrate opposite trends, which are obviously crucial for impaired regulation of blood pressure in SHR.

Antioxidant effect of doxycycline decreases MMP activity and blood pressure in SHR

Increased matrix metalloproteinase (MMP) levels are involved in vascular remodeling of hypertension. In this study, we hypothesized that doxycycline (a MMP inhibitor) could exert antioxidant effects, reverse establish vascular remodeling, and lower blood pressure in spontaneously hypertensive rats (SHR). SHR and Wistar-Kyoto rats received either doxycycline at 30mg/kg/day by gavage or vehicle. Systolic blood pressure (SBP) was assessed weekly by tail cuff. After 5weeks of treatment, morphologic changes in the aortic wall were studied in hematoxylin/eosin sections. MMP activity and expression were determined by in situ zymography using DQ gelatin and immunofluorescence for MMP-2. Dihydroethidium was used to evaluate aortic reactive oxygen species (ROS) production by fluorescence microscopy. Doxycycline reduced SBP by 25mmHg. However, the antihypertensive effects were not associated with significant reversal of hypertension-induced vascular hypertrophy. SHR showed increased aortic MMP-2 levels which co-localized with higher aortic MMP activity and ROS levels, and all those biochemical alterations associated with hypertension were blunted by treatment with doxycycline. These results show that MMP inhibition with doxycycline in SHR with established hypertension resulted in antioxidant effects, lower gelatinolytic activity, and antihypertensive effects which were not associated with reversal of hypertension-induced vascular remodeling.

Two different approaches to restore renal nitric oxide and prevent hypertension in young spontaneously hypertensive rats: l-citrulline and nitrate

Nitric oxide (NO) deficiency mediates oxidative stress in the kidney and is involved in the development of hypertension. NO synthesis occurs via 2 pathways: nitric oxide synthase (NOS) dependent and NOS-independent. We tested whether the development of hypertension is prevented by restoration of NO by dietary l-citrulline or nitrate supplementation in young spontaneously hypertensive rats (SHRs). Male SHRs and normotensive Wistar Kyoto control rats (WKYs)s age 4weeks were assigned to 4groups: untreated SHRs and WKYs, and SHRs and WKYs that received 0.25% l-citrulline for 8weeks. In our second series of studies, we replaced l-citrulline with 1mmol/kg/d sodium nitrate. All rats were sacrificed at age 12weeks. We found an increase in the blood pressure of SHRs was prevented by dietary supplementation of l-citrulline or nitrate. Both treatments restored NO bioavailability and reduced oxidative stress in SHR kidneys. l-Citrulline therapy reduced levels of l-arginine and asymmetric dimethylarginine (ADMA)-an endogenous inhibitor of NOS-and increased the l-arginine-to-ADMA ratio in SHR kidneys. Nitrate treatment reduced plasma levels of l-arginine and ADMA concurrently in SHRs. Our findings suggest that both NOS-dependent and -independent approaches in the prehypertensive stage toward augmentation of NO can prevent the development of hypertension in young SHRs.

Combined antihypertensive effect of luteolin and buddleoside enriched extracts in spontaneously hypertensive rats

Ethnopharmacological relevanceFlos Chrysanthemi is used in a variety of diseases in traditional Chinese medicine including hypertension, and the total flavonoids (rich in luteolin (LUT) and buddleoside (BUD)) of Flos Chrysanthemi is known to modulate vascular functions and reduce the blood pressure. However, the active flavonoids and their synergistic effects on anti-hypertension are still unclear. To investigate the combined anti-hypertension effects of LUT and BUD enriched extracts on spontaneously hypertensive rats (SHR), as well as the anti-hypertensive mechanism of LUT&BUD mixture. Materials and methodsCODA Mouse & Rat Tail-Cuff Blood Pressure System was used to measure the systolic blood pressure (SBP) and diastolic blood pressure (DBP) of SHR after treated with extracts contains with LUT and/or BUD. The expressions of Ang II, PRA, ALD, ET, PGI2 and TXB2 were investigated by ELASA. Serum NO concentration was measured by the method of Nitric acid reductase. ResultsA single administration of LUT, BUD, or LUT:BUD=1:1 significantly reduced SBP by about 3.35mmHg, 4.39mmHg and 15.42mmHg, respectively. Chronic administration of LBM (at 60mg/kg; p.o. for 30 days) reduced both SBP and DBP by 4.04% and 5.24% of the vehicle group, respectively. Oral administration of LBM at 60mg/kg inhibited the serum levels of ANG, ALD and ET, but increased serum NO concentration. ConclusionThis study shows the synergistic anti-hypertension effects of LUT and BUD in SHR. The effects of LBM on blood pressure are associated with RAAS and endothelial system. Thus, our experiments suggest that the combination of luteolin and buddleoside from Flos Chrysanthemi are potentially useful for the therapeutic treatments for hypertension.

Characterization of Ile‐His‐Arg‐Phe, a novel rice‐derived vasorelaxing peptide with hypotensive and anorexigenic activities

Recently, we found that dipeptide Arg-Phe (RF) had cholecystokinin (CCK)-dependent vasorelaxing activity. The RF sequence is often observed in the primary structure of natural food proteins. In the current study, we investigated enzymatic conditions for the release of RF-related peptides from rice glutelin, a major storage protein, using gastrointestinal proteases. RF-related peptides were then characterized. It was found that RF and Ile-His-Arg-Phe (IHRF) were released in the chymotrypsin digest of the partial structure of rice glutelin. We then focused on previously unidentified IHRF, corresponding to rice glutelin(155-158). IHRF had vasorelaxing activity in the mesenteric artery of spontaneous hypertensive rats (SHRs). Orally administered IHRF lowered systolic blood pressure in SHRs. The antihypertensive activity of IHRF was more potent and long-lasting than that of RF. IHRF-induced vasorelaxing activity was not blocked by inhibitors of nitric oxide synthase and cyclooxygenase, but by an antagonist for CCK₁ receptor. IHRF also had CCK-like suppressive activities in food intake and gastrointestinal transit. IHRF increased intracellular Ca²⁺ flux and CCK release in the enteroendocrine cell STC-1. IHRF, a novel CCK-dependent vasorelaxing peptide, decreases both blood pressure and food intake in rodents.

Protective effect of a pomace olive oil concentrated in triterpenic acids in alterations related to hypertension in rats: Mechanisms involved

Despite the amount of information and research on the effects of virgin olive oil and its components in cardiovascular disease, little attention has been paid to the effects of pomace olive oil, an olive oil subproduct traditionally used in Spain. The aim of the present study was to evaluate the potential effects of a pomace olive oil concentrated in triterpenic acids (POCTA) on blood pressure, cardiac hemodynamics, and functional and molecular vascular alterations associated with hypertension in spontaneously hypertensive rats (SHR). The study showed that POCTA attenuated the increase of blood pressure in SHR. This effect was associated with an improvement in endothelium-dependent relaxation, enhancement of vascular expression of endothelial nitric oxide synthase, and reduction of tumor necrosis factor alpha, transforming growth factor beta, and collagen I. Furthermore, POCTA improved cardiac hemodynamics (left ventricular systolic pressure and left ventricular end-diastolic pressure) and decreased relative kidney and lung weights. POCTA exerts antihypertensive effects together with vascular and hypertension target organ protection in SHR. Since interest in pomace olive oil has been low, the results of this study contribute to increasing awareness of its biological and nutritional values.

Abstract 439: Role of Hindbrain Endogenous Melanocortin Receptor Activity in Contributing to Hypertension in SHR

We previously demonstrated that activation of the central nervous system (CNS) melanocortin system plays a key role in the modulation of sympathetic nerve activity (SNA) and blood pressure (BP) regulation. In the present study, we examined whether chronic inhibition of melanocortin-4 receptors (MC4R) in the hindbrain attenuates the elevated blood pressure in spontaneously hypertensive rats (SHR). The long-term cardiovascular and metabolic effects of MC4R antagonism were assessed in SHR and normotensive Wistar-Kyoto (WKY) rats. Male WKY rats (n=6) and SHR (n=7) were implanted with telemetry probes to measure BP and heart rate (HR) 24-hrs/day, and an intracerebroventricular (ICV) cannula was placed into the fourth ventricle (4 th V). After 10 days of recovery and 5 days of control measurements, the MC4R antagonist (SHU-9119, 1 nmol/h, 4 th V) was infused for 10 days followed by a 5-day recovery period. Chronic hindbrain MC4R antagonism significantly increased food intake and body weight in WKY (17±1 to 35±2 g/day and 280±8 to 353±8 g) and SHR (19±2 to 35±2 g/day and 323±7 to 371±11 g). No changes were observed in blood glucose levels (99±4 to 87±4 and 89±5 to 89±4 mg/dl, respectively for WKY and SHR). Chronic SHU-9119 infusion reduced MAP and HR similarly in WKY (-5±1 mmHg and -47±3 bpm) and SHR (-8±3 mmHg and -44±3 bpm). These results suggest that although hindbrain MC4R activity is important for appetite and HR regulation, it does not play a major role in mediating the elevated blood pressure in SHR. (NHLBI-PO1 HL51971/AHA SDG5680016)

Abstract 454: Hypertension Control by Decrease of Sympathetic Hyperactivity in SHR Rats: Comparison Between Renal Denervation and Dopamine ß-hydroxylase Inhibition

Activation of the sympathetic nervous system has been described in the development and progression of systemic hypertension. Recently, catheter-based renal nerve ablation for treatment of drug-resistant hypertension has been developed. An alternative strategy for modulation of sympathetic nerve function is to reduce the biosynthesis of norepinephrine (NE) via inhibition of dopamine β-hydroxylase (DβH), the enzyme that catalysis the conversion of dopamine (DA) to NE in sympathetic nerves. Renal denervation was assessed in the rat by transient (90 sec) ischemia of renal tissues. Bilateral renal artery occlusion (RAO) was assessed in spontaneously hypertensive rats (SHR) to evaluate the effect of renal denervation on NE levels and blood pressure throughout a 28-day period. Oral treatment with the selective peripheral DβH inhibitor etamicastat (30 mg/Kg body weight) was performed in a second cohort of SHR and NE levels and blood pressure was assessed. Renal denervation resulted in marked depletion of NE levels, selective for the renal sympathetic innervations, 5 days after RAO in renal cortex (16.5±7.7% of control values, P&lt;0.0001). Decreased NE levels persisted up to 28 days after RAO (40.8±19.4% of control values, P&lt;0.0001). No differences in left ventricle NE levels were observed. Renal denervation significantly decreased systolic blood pressure (SBP) 5 days (165±6 vs. 187±2 mmHg, P&lt;0.05) and 14 days post-surgery (194±3 vs. 204±2 mmHg, P&lt;0.05), but had no effect on heart rate. Systolic blood pressure recovered to basal values 28 days post-surgery. Likewise, etamicastat treatment resulted in a depletion of NE levels in both renal cortex (70.9±9.1% of control values, P&lt;0.05) and left ventricle (55.8±3.9% of control values, P&lt;0.01). Etamicastat treatment decreased SBP (228±7 vs. 174±9 mmHg, P&lt;0.001) without effects on heart rate. In conclusion, these results suggest that renal denervation and DβH inhibition similarly decreases sympathetic hyperactivity and high blood pressure in the SHR. Downregulation of sympathetic drive with DβH inhibitors may be useful in the treatment of hypertension.

Abstract 253: Inhibition of Necrosis Attenuates Blood Pressure and Renal Inflammation in Male SHR with no Effect in Females

Inflammation plays an important role in the pathogenesis of hypertension, and we recently published that male spontaneously hypertensive rats (SHR) have more pro-hypertensive, pro-inflammatory Th17 cells in their kidneys than females. However, the mechanisms initiating the inflammatory response remain unclear. Damage-associated molecular patterns (DAMPs) are released from dying cells and induce immune system activation. High mobility group box 1 (HMGB1) is a DAMP actively secreted by apoptotic cells in an oxidized form, or passively released by necrotic cells in a reduced form. Reduced HMGB1 has prolonged activity and is more pro-inflammatory. We hypothesized that sex differences in cell death contribute to sex differences in the immune cell profile and hypertension. Renal necrosis and apoptosis were measured by flow cytometry in 12 wk old male and female SHR. Male SHR have greater necrotic cell death (% total renal cells: 5.3±0.8 vs.1.0±0.4; p=0.004; N=6), while female SHR have more apoptosis (% total renal cells: 1.6±0.4 vs.4.0±0.4; p=0.004; N=4-5). Male SHR also had greater reduced HMGB1 in renal cortex compared to females (HMGB1 oxidized/reduced ratio: 1.0±0.1 vs. 1.7±0.1; p=0.002; N=6). We next measured blood pressure by telemetry in male and female SHR treated with vehicle or necrosis inhibitor NecroX-5 (1 mg/kg per day; 2 wks). The renal T cell and macrophage levels were measured by flow cytometry. NecroX-5 treatment in male SHR prevented age-related increases in blood pressure compared to vehicle-treated rats (ΔmmHg/week: 0.8±1.1 vs. 4.2±0.3; p=0.05; N=3). Necrox5 also significantly decreased renal necrosis (% total renal cells: 5.3±0.8 vs. 0.8±0.1; p=0.005; N=3-6), macrophages (% total renal cells: 2.8±0.6 vs. 0.8± 0.1; p=0.04; N=3-4), Th17 cells (% T cells: 4.5±0.3 vs. 2.7±0.3; p=0.009; N=3-4) and the pro-inflammatory cytokine IL-17 (% total renal cells: 10.0±0.4 vs. 5.3±0.7; p=0.002; N=3-4) in male SHR. NecroX-5 had no effect on blood pressure or pro-inflammatory markers in females. We conclude that necrosis is an important contributor in progression of hypertension in male SHR. Sex differences in cell death may contribute to sex differences in blood pressure and the immune profile.

Effect of melatonin supplementation and cross-fostering on renal glutathione system and development of hypertension in spontaneously hypertensive rats

Antenatal and postnatal environments are hypothesised to influence the development of hypertension. This study investigates the synergistic effect of cross-fostering and melatonin supplementation on the development of hypertension and renal glutathione system in spontaneously hypertensive rats (SHR). In one experiment, 1-day-old male SHR pups were fostered to either SHR (shr-SHR) or Wistar-Kyoto rats, (shr-WKY). In a concurrent experiment, SHR dams were given melatonin in drinking water (10 mg/kg body weight) from day 1 of pregnancy. Immediately following delivery, 1-day-old male pups were fostered either to SHR (Mel-shr-SHR) or WKY (Mel-shr-WKY) dams receiving melatonin supplementation until weaning on day 21. Upon weaning, melatonin supplementation was continued to these pups until the age of 16 weeks. Systolic blood pressures (SBP) were recorded at the age of 4, 6, 8, 12 and 16 weeks. Renal antioxidant activities were measured. Mean SBP of shr-WKY, Mel-shr-SHR and Mel-shr-WKY was significantly lower than that in shr-SHR until the age of 8 weeks. At 12 and 16 weeks of age, mean SBP of Mel-shr-WKY was lower than those in non-treated shr-SHR and shr-WKY pups but was not significantly different from that in Mel-shr-SHR. Renal glutathione peroxidase (GPx) and glutathione S-transferase (GST) activities were significantly higher in Mel-shr-SHR and Mel-shr-WKY at 16 weeks of age. It appears that combination of cross-fostering and melatonin supplementation exerts no synergistic effect on delaying the rise in blood pressure in SHR. The elevated GPx and GST activities are likely to be due to the effect of melatonin supplementation.

Correction of vascular hypercontractility in spontaneously hypertensive rats using shRNAs-induced delta protein kinase C gene silencing

Potassium conductance in vascular smooth muscle (VSM) is known to be altered in arterial hypertension. High level of protein kinase C (PKC) activity is a common feature for hypertension of different genesis. The main goal of this study was to investigate the efficacy of the RNA interference (RNAi) technique targeting PKC delta-isoform gene as a possible pharmacological tool to restore vasodilator potential in spontaneously hypertensive rats (SHR). Experimental design of the study comprised RNAi and patch-clamp techniques, RT-PCR analysis and standard acetylcholine test. Total outward currents and acetylcholine-induced endothelium-dependent relaxant responses were blunted in SHR. BKCa alpha subunit mRNA expression in SHR was unchanged whereas KV and KATP mRNA expression appeared significantly increased. PKC inhibitor, chelerythrine (100nM), restored potassium channels activity in SHR. PKC-delta-isoform protein expression and PKC-delta-isoform mRNA expression are 2.5–4 fold increased in VSM from SHR. PKC gene silencing with the short hairpin RNAs (shRNAs)-plasmid delivery system administered intravenously led to an increment in maximal amplitude of acetylcholine-relaxation, restored outward K+ currents and PKC-delta-isoform mRNA and protein expression. Arterial blood pressure in SHR was normalized following shRNAs administration. We conclude that BKCa channels are likely to be the most PKC-dependent member of K+ channels family responsible for vascular hypercontractility in SHR while Kv and KATP channels may constitute a reserve mechanism for the maintenance of vasodilator potential under BKCa channelopathy. It is likely that RNAi technique is a good therapeutic approach to inactivate PKC gene and to normalize vascular functions and high arterial blood pressure in SHR.

Administration of Telmisartan Reduced Systolic Blood Pressure and Oxidative Stress Probably Through the Activation of PI3K/Akt/eNOS Pathway and NO Release in Spontaneously Hypertensive Rats

We investigated the effects of telmisartan, the blocker of angiotensin II receptor 1, on the regulation of systolic blood pressure (SBP) and oxidative stress through endothelial nitric oxide (NO) release in spontaneously hypertensive rats (SHRs). SHRs randomly received placebo, oral feeding of telmisartan (5 mg/kg or 10 mg/kg) every day and Wistar-Kyoto rats (WKYs) served as normotensive control. The SBP of rat was measured before and weekly thereafter. After a total of 8-week treatment, rats were killed for experimental measurements. Parameters that subject to measurements in isolated aorta endothelial cells include: NO concentration, protein expression levels of angiotensin II receptor 1, nitrotyrosine, 8-isoprostane, SOD, PI3K, Akt, AMPK and eNOS. In addition, L-NMMA, a general inhibitor of nitric oxide synthase, was also applied to test the inhibition of NO concentration. We found that SBPs were significantly lower in telmisartan therapy group than in placebo treated hypertensive rats and WKYs (p<0.05). The NO concentration was significantly higher in telmisartan-treated group with increased activity of the PI3K/Akt pathway and activated eNOS signaling. Blockade of Akt activity reversed such effects. Activation of AMPK also contributed to the phosphorylation of eNOS. L-NMMA treatment reduced less NO concentration in SHR rats than the telmisartan co-treated groups. Oxidative stress in SHRs was also attenuated by telmisartan administration, shown by reduced formation of nitrotyrosine, 8-isoprostane, and recovered SOD protein level. Telmisartan enhanced NO release by activating the PI3K/Akt system, AMPK phosphorylation and eNOS expression, which attenuated the blood pressure and oxidative stress in SHRs.