Chronic Nitric Oxide Synthase Inhibition Research Articles

Effects of semaglutide, peptide YY3-36 and empagliflozin on metabolic dysfunctionsassociated fatty steatotic liver disease in diet-induced obese rats with chronic nitric oxide synthase-inhibition

Effects of semaglutide, peptide YY3-36 and empagliflozin on metabolic dysfunctionsassociated fatty steatotic liver disease in diet-induced obese rats with chronic nitric oxide synthase-inhibition

Effects of semaglutide, Peptide YY3-36 and empagliflozin on metabolic dysfunction associated fatty steatotic liver disease in diet-induced obese rats with chronic nitric oxide synthase-inhibition

Effects of semaglutide, Peptide YY3-36 and empagliflozin on metabolic dysfunction associated fatty steatotic liver disease in diet-induced obese rats with chronic nitric oxide synthase-inhibition

Suppression of nitric oxide synthase aggravates non-alcoholic steatohepatitis and atherosclerosis in SHRSP5/Dmcr rat via acceleration of abnormal lipid metabolism.

Non-alcoholic steatohepatitis (NASH) is a progressive subtype ofnon-alcoholic fatty liver disease (NAFLD) that is closely related to cardiovascular disease (CVD). Nitric oxide (NO) plays a critical role in the control of various biological processes. Dysfunction of the NO signaling pathway is associated with various diseases such as atherosclerosis, vascular inflammatory disease, and diabetes. Recently, it has been reported that NO is related to lipid and cholesterol metabolism. Chronic NO synthase (NOS) inhibition accelerates NAFLD by increasing hepatic lipid deposition. However, the detailed relationship between NO and abnormal lipid and cholesterol metabolism in NAFLD/NASH has not been completely explained. We aimed to determine the effects of NOS inhibition by N omega-nitro-L-arginine methyl ester hydrochloride (L-NAME), a NOS inhibitor, on NASH and CVD via lipid and cholesterol metabolism. Stroke-prone spontaneously hypertensive rats were fed a high-fat and high-cholesterol diet for 8weeks and administered L-NAME for the last 2weeks. Following blood and tissue sampling, biochemical analysis, histopathological staining, quantitative RT-PCR analysis, and western blotting were performed. L-NAME markedly increased hepatic triglyceride (TG) and cholesterol levels by promoting TG synthesis and cholesterol absorption from the diet. L-NAME increased the mRNA levels of inflammatory markers and fibrotic areas in the liver. Cholesterol secretion from the liver was promoted in rats administered L-NAME, which increased serum cholesterol. L-NAME significantly increased the level of oxidative stress marker and lipid deposition in the arteries. NOS inhibition simultaneously aggravates NASH and atherosclerosis via hepatic lipid and cholesterol metabolism.

Protective role of the mitochondrial fusion protein OPA1 in hypertension.

Hypertension is associated with excessive reactive oxygen species (ROS) production in vascular cells. Mitochondria undergo fusion and fission, a process playing a role in mitochondrial function. OPA1 is essential for mitochondrial fusion. Loss of OPA1 is associated with ROS production and cell dysfunction. We hypothesized that mitochondria fusion could reduce oxidative stress that defect in fusion would exacerbate hypertension. Using (a) Opa1 haploinsufficiency in isolated resistance arteries from Opa1+/- mice, (b) primary vascular cells from Opa1+/- mice, and (c) RNA interference experiments with siRNA against Opa1 in vascular cells, we investigated the role of mitochondria fusion in hypertension. In hypertension, Opa1 haploinsufficiency induced altered mitochondrial cristae structure both in vascular smooth muscle and endothelial cells but did not modify protein level of long and short forms of OPA1. In addition, we demonstrated an increase of mitochondrial ROS production, associated with a decrease of superoxide dismutase 1 protein expression. We also observed an increase of apoptosis in vascular cells and a decreased VSMCs proliferation. Blood pressure, vascular contractility, as well as endothelium-dependent and -independent relaxation were similar in Opa1+/- , WT, L-NAME-treated Opa1+/- and WT mice. Nevertheless, chronic NO-synthase inhibition with L-NAME induced a greater hypertension in Opa1+/- than in WT mice without compensatory arterial wall hypertrophy. This was associated with a stronger reduction in endothelium-dependent relaxation due to excessive ROS production. Our results highlight the protective role of mitochondria fusion in the vasculature during hypertension by limiting mitochondria ROS production.

Cardiac remodelling in a swine model of chronic thromboembolic pulmonary hypertension: comparison of right vs. left ventricle.

Key points Right ventricle (RV) function is the most important determinant of survival and quality of life in patients with chronic thromboembolic pulmonary hypertension (CTEPH).The changes in right and left ventricle gene expression that contribute to ventricular remodelling are incompletely investigated.RV remodelling in our CTEPH swine model is associated with increased expression of the genes involved in inflammation (TGFβ), oxidative stress (ROCK2, NOX1 and NOX4), and apoptosis (BCL2 and caspase‐3).Alterations in ROCK2 expression correlated inversely with RV contractile reserve during exercise.Since ROCK2 has been shown to be involved in hypertrophy, oxidative stress, fibrosis and endothelial dysfunction, ROCK2 inhibition may present a viable therapeutic target in CTEPH. Right ventricle (RV) function is the most important determinant of survival and quality of life in patients with chronic thromboembolic pulmonary hypertension (CTEPH). The present study investigated whether the increased cardiac afterload is associated with (i) cardiac remodelling and hypertrophic signalling; (ii) changes in angiogenic factors and capillary density; and (iii) inflammatory changes associated with oxidative stress and interstitial fibrosis. CTEPH was induced in eight chronically instrumented swine by chronic nitric oxide synthase inhibition and up to five weekly pulmonary embolizations. Nine healthy swine served as a control. After 9 weeks, RV function was assessed by single beat analysis of RV–pulmonary artery (PA) coupling at rest and during exercise, as well as by cardiac magnetic resonance imaging. Subsequently, the heart was excised and RV and left ventricle (LV) tissues were processed for molecular and histological analyses. Swine with CTEPH exhibited significant RV hypertrophy in response to the elevated PA pressure. RV–PA coupling was significantly reduced, correlated inversely with pulmonary vascular resistance and did not increase during exercise in CTEPH swine. Expression of genes associated with hypertrophy (BNP), inflammation (TGFβ), oxidative stress (ROCK2, NOX1 and NOX4), apoptosis (BCL2 and caspase‐3) and angiogenesis (VEGFA) were increased in the RV of CTEPH swine and correlated inversely with RV–PA coupling during exercise. In the LV, only significant changes in ROCK2 gene‐expression occurred. In conclusion, RV remodelling in our CTEPH swine model is associated with increased expression of genes involved in inflammation and oxidative stress, suggesting that these processes contribute to RV remodelling and dysfunction in CTEPH and hence represent potential therapeutic targets.

The Probiotic Lactobacillus fermentum Prevents Dysbiosis and Vascular Oxidative Stress in Rats with Hypertension Induced by Chronic Nitric Oxide Blockade

Whether the probiotic Lactobacillus fermentum CECT5716 (LC40) ameliorates hypertension in rats with chronic nitric oxide (NO) synthase inhibition is tested. Rats are randomly divided into four different groups and treated for 4weeks: a) vehicle (control), b) vehicle plus NG -nitro-l-arginine methyl ester (l-NAME; 50mg100mL-1 in drinking water), c) LC40 (109 colony-forming unitsd-1 by gavage), and d) LC40 plus l-NAME. l-NAME induces gut dysbiosis, characterized mainly by an increased Fimicutes/Bacteroidetes (F/B) ratio and reduced Bifidobacterium content, increased Th17 cells and reduced Treg in mesenteric lymph nodes (MLN), increased aortic Th17 infiltration and reactive oxygen species, reduced aortic endothelium-dependent relaxant response to acetylcholine, and hypertension. LC40 prevents gut dysbiosis, alters the Th17/Treg balance in MLN, vascular oxidative stress, and inflammation, slightly improves endothelial dysfuncion but do not inhibit the development of l-NAME-induced hypertension. Chronic LC40 treatment, in this model of chronic inhibition of NO synthesis, reduces early events involved in atherosclerosis development, such as vascular oxidative stress and pro-inflammatory status, as a result of prevention of gut dysbiosis and immune changes in MLN, but not hypertension, confirming the critical role of NO in the antihypertensive effects of LC40 in genetic hypertension.

Nitric Oxide Synthase Inhibition Induces Renal Medullary Hypoxia in Conscious Rats.

BackgroundRenal hypoxia, implicated as crucial factor in onset and progression of chronic kidney disease, may be attributed to reduced nitric oxide because nitric oxide dilates vasculature and inhibits mitochondrial oxygen consumption. We hypothesized that chronic nitric oxide synthase inhibition would induce renal hypoxia.Methods and ResultsOxygen‐sensitive electrodes, attached to telemeters, were implanted in either renal cortex (n=6) or medulla (n=7) in rats. After recovery and stabilization, baseline oxygenation (pO 2) was recorded for 1 week. To inhibit nitric oxide synthase, N‐ω‐nitro‐l‐arginine (L‐NNA; 40 mg/kg/day) was administered via drinking water for 2 weeks. A separate group (n=8), instrumented with blood pressure telemeters, followed the same protocol. L‐NNA rapidly induced hypertension (165±6 versus 108±3 mm Hg; P<0.001) and proteinuria (79±12 versus 17±2 mg/day; P<0.001). Cortical pO 2, after initially dipping, returned to baseline and then increased. Medullary pO 2 decreased progressively (up to −19±6% versus baseline; P<0.05). After 14 days of L‐NNA, amplitude of diurnal medullary pO 2 was decreased (3.7 [2.2–5.3] versus 7.9 [7.5–8.4]; P<0.01), whereas amplitudes of blood pressure and cortical pO 2 were unaltered. Terminal glomerular filtration rate (1374±74 versus 2098±122 μL/min), renal blood flow (5014±336 versus 9966±905 μL/min), and sodium reabsorption efficiency (13.0±0.8 versus 22.8±1.7 μmol/μmol) decreased (all P<0.001).ConclusionsFor the first time, we show temporal development of renal cortical and medullary oxygenation during chronic nitric oxide synthase inhibition in unrestrained conscious rats. Whereas cortical pO 2 shows transient changes, medullary pO 2 decreased progressively. Chronic L‐NNA leads to decreased renal perfusion and sodium reabsorption efficiency, resulting in progressive medullary hypoxia, suggesting that juxtamedullary nephrons are potentially vulnerable to prolonged nitric oxide depletion.

The Effect of Chronic NO Synthase Inhibition on the Vasoactive and Structural Properties of Thoracic Aorta, NO Synthase Activity, and Oxidative Stress Biomarkers in Young SHR.

Although the role of nitric oxide (NO) in essential hypertension is still unclear, the effects of long-term NO deficiency have not yet been investigated during the critical juvenile period in spontaneously hypertensive rats (SHR). We aimed to analyze the effects of chronic NO synthase (NOS) inhibition on systolic blood pressure (sBP), vasoactivity, morphological changes and superoxide level in the thoracic aorta (TA), NOS activity in different tissues, and general biomarkers of oxidative stress in plasma of young SHR. Four-week-old SHR were treated with NG-nitro-L-arginine methyl ester (L-NAME, 50 mg/kg/day, p.o.) for 4-5 weeks. L-NAME treatment induced a transient sBP increase only, and surprisingly, slightly inhibited endothelium-dependent relaxation of TA. Hereby, the inhibition of NOS activity varied from tissue to tissue, ranging from the lowest in the TA and the kidney to the highest in the brain stem. In spite of an increased sensitivity of adrenergic receptors, the maximal adrenergic contraction of TA was unchanged, which was associated with changes in elastin arrangement and an increase in wall thickness. The production of reactive oxygen species in the TA was increased; however, the level of selected biomarkers of oxidative stress did not change. Our findings proved that the TA of young SHR responded to chronic NO deficiency by the development of adaptive mechanisms on the functional (preserved NO-derived vasorelaxation, unincreased contraction) and molecular (preserved NOS activity) level.

The novel organic mononitrate NDHP attenuates hypertension and endothelial dysfunction in hypertensive rats

RationaleDevelopment and progression of cardiovascular diseases, including hypertension, are often associated with impaired nitric oxide synthase (NOS) function and nitric oxide (NO) deficiency. Current treatment strategies to restore NO bioavailability with organic nitrates are hampered by undesirable side effects and development of tolerance. In this study, we evaluated NO release capability and cardiovascular effects of the newly synthesized organic nitrate 1, 3-bis (hexyloxy) propan-2-yl nitrate (NDHP). MethodsA combination of in vitro and in vivo approaches was utilized to assess acute effects of NDHP on NO release, vascular reactivity and blood pressure. The therapeutic value of chronic NDHP treatment was assessed in an experimental model of angiotensin II-induced hypertension in combination with NOS inhibition. ResultsNDHP mediates NO formation in both cell-free system and small resistance arteries, a process which is catalyzed by xanthine oxidoreductase. NDHP-induced vasorelaxation is endothelium independent and mediated by NO release and modulation of potassium channels. Reduction of blood pressure following acute intravenous infusion of NDHP was more pronounced in hypertensive rats (two-kidney-one-clip model) than in normotensive sham-operated rats. Toxicological tests did not reveal any harmful effects following treatment with high doses of NDHP. Finally, chronic treatment with NDHP significantly attenuated the development of hypertension and endothelial dysfunction in rats with chronic NOS inhibition and angiotensin II infusion. ConclusionAcute treatment with the novel organic nitrate NDHP increases NO formation, which is associated with vasorelaxation and a significant reduction of blood pressure in hypertensive animals. Chronic NDHP treatment attenuates the progression of hypertension and endothelial dysfunction, suggesting a potential for therapeutic applications in cardiovascular disease.

Carbon monoxide does not contribute to vascular tonus improvement in exercise-trained rats with chronic nitric oxide synthase inhibition

Carbon monoxide (CO), an end product of heme oxygenase (HO) that is involved in the regulation of vascular tonus, may show a compensatory effect in nitric oxide (NO) deficiency. This study aimed to assess the effect of the HO/CO system on the vascular tone in exercise-trained rats with hypertension induced by chronic NO synthase (NOS) inhibition. Hypertension was induced by N-nitro-l-arginine methyl ester (25 mg/kg/day in drinking water), and exercise training comprised swimming 1 h/day, 5 days/week, for 6 weeks. Systolic blood pressure (BP) was measured weekly using a tail-cuff method. The effects of hypertension and/or exercise-training on the constriction and relaxation responses of the thoracic aorta and resistance arteries of the mesenteric and gastrocnemius vascular beds were evaluated. NOS inhibition produced a gradually developed hypertension, and the magnitude of the increase in BP was significantly attenuated by exercise training. Although phenylephrine (Phe)-induced contraction responses of aorta incubated with an HO-1 inhibitor were reduced in hypertensive animals, there was no difference in the hypertensive-exercise group. However, thoracic aortas in the hypertensive-exercise group exhibited significantly more relaxation in response to a CO donor. There was no change in Phe-induced contraction with or without HO inhibition CO donor relaxation responses in both resistance arteries. These results suggest that the HO/CO system does not contribute to diminishing BP by exercise training in a NOS inhibition-induced hypertension model.

Effects of chronic nitric oxide synthase inhibition on V’O2max and exercise capacity in mice

Acute inhibition of NOS by L-NAME (Nω-nitro-L-arginine methyl ester) is known to decrease maximal oxygen consumption (V'O2max) and impair maximal exercise capacity, whereas the effects of chronic L-NAME treatment on V'O2max and exercise performance have not been studied so far. In this study, we analysed the effect of L-NAME treatment, (LN2 and LN12, respectively) on V'O2max and exercise capacity (in maximal incremental running and prolonged sub-maximal incremental running tests), systemic NO bioavailability (plasma nitrite (NO2-) and nitrate (NO3-)) and prostacyclin (PGI2) production in C57BL6/J mice. Mice treated with L-NAME for 2weeks (LN2) displayed higher V'O2max and better running capacity than age-matched control mice. In LN2 mice, NO bioavailability was preserved, as evidenced by maintained NO2- plasma concentration. PGI2 production was activated (increased 6-keto-PGF1α plasma concentration) and the number of circulating erythrocytes (RBC) and haemoglobin concentration were increased. In mice treated with L-NAME for 12weeks (LN12), NO bioavailability was decreased (lower NO2- plasma concentration), and 6-keto-PGF1α plasma concentration and RBC number were not elevated compared to age-matched control mice. However, LN12 mice still performed better during the maximal incremental running test despite having lower V'O2max. Interestingly, the LN12 mice showed poorer running capacity during the prolonged sub-maximal incremental running test. To conclude, short-term (2weeks) but not long-term (12weeks) treatment with L-NAME activated robust compensatory mechanisms involving preservation of NO2- plasma concentration, overproduction of PGI2 and increased number of RBCs, which might explain the fully preserved exercise capacity despite the inhibition of NOS.

Effects of a novel ACE inhibitor, 3-(3-thienyl)-l-alanyl-ornithyl-proline, on endothelial vasodilation and hepatotoxicity in l-NAME-induced hypertensive rats.

Nitric oxide (NO) is a widespread biological mediator involved in many physiological and pathological processes, eg, in the regulation of vascular tone and hypertension. Chronic inhibition of NO synthase by NG-nitro-l-arginine methyl ester (l-NAME) hydrochloride results in the development of hypertension accompanied by an increase in vascular responsiveness to adrenergic stimuli. Recently, we developed a novel sulfur-containing angiotensin-converting enzyme inhibitor: 3-(3-thienyl)-l-alanyl-ornithyl-proline (TOP). Our previous studies indicated a superior nature of the molecule as an antihypertensive agent in spontaneously hypertensive rats (showing the involvement of renin–angiotensin–aldosterone system) in comparison to captopril. The aim of the present study was to investigate the effect of TOP on NO pathway in l-NAME-induced hypertensive rats, and captopril was included as the standard treatment group. Treatment with both TOP (20 mg/kg) and captopril (40 mg/kg) prevented the development of hypertension in l-NAME model, but TOP showed better restoration of NO and normal levels of angiotensin-converting enzyme. In addition, in vitro vasorelaxation assay showed an improvement in endothelium-dependent vasodilation in both the cases. Further, the biochemical (malondialdehyde, alanine aminotransferase, and aspartate aminotransferase) and the histopathological effects of TOP on rat liver tissues revealed a protective nature of TOP in comparison to captopril in the l-NAME model. In conclusion, TOP at 50% lesser dose than captopril was found to be better in the l-NAME model.

Acute and chronic role of nitric oxide, renin-angiotensin system and sympathetic nervous system in the modulation of calcium sensitization in Wistar rats.

Principal vasoactive systems - renin-angiotensin system (RAS), sympathetic nervous system (SNS), nitric oxide (NO) and prostanoids - exert their vascular effects through the changes in calcium levels and/or calcium sensitization. To estimate a possible modulation of calcium sensitization by the above vasoactive systems, we studied the influence of acute and chronic blockade of particular vasoactive systems on blood pressure (BP) changes elicited in conscious normotensive rats by acute dose-dependent administration of Rho-kinase inhibitor fasudil. Adult male chronically cannulated Wistar rats were used throughout this study. The acute inhibition of NO synthase (NOS) by L-NAME enhanced BP response to fasudil, the effect being considerably augmented in rats deprived of endogenous SNS. The acute inhibition of prostanoid synthesis by indomethacin modified BP response to fasudil less than the acute NOS inhibition. The chronic NOS inhibition caused moderate BP elevation and a more pronounced augmentation of fasudil-induced BP changes compared to the effect of acute NOS inhibition. This indicates both short-term and long-term NO-dependent attenuation of calcium sensitization. Long-term inhibition of RAS by captopril caused a significant attenuation of BP changes elicited by fasudil. In contrast, a long-term attenuation of SNS by chronic guanethidine treatment (in youth or adulthood) had no effect on BP response to fasudil, suggesting the absence of SNS does not affect calcium sensitization in vascular smooth muscle of normotensive rats. In conclusion, renin-angiotensin system contributes to the long-term increase of calcium sensitization and its effect is counterbalanced by nitric oxide which decreases calcium sensitization in Wistar rats.

Systemic Aldosterone, But Not Angiotensin II, Plays a Pivotal Role in the Pathogenesis of Renal Injury in Chronic Nitric Oxide-Deficient Male Rats

Chronic inhibition of nitric oxide synthase by N(ω)-nitro-L-arginine methyl ester (L-NAME) causes progressive renal injury and systemic hypertension. Angiotensin II (Ang II) has been conventionally regarded as one of the primary causes of renal injury. We reported previously that such renal injury was almost completely suppressed by both an Ang II type I receptor blocker and an aldosterone antagonist. The aldosterone antagonist also inhibited the systemic Ang II elevation. Therefore, it remains to be elucidated whether Ang II or aldosterone directly affects the development of such renal injury. In the present study, we investigated the role of aldosterone in the pathogenesis of renal injury induced by L-NAME-mediated chronic nitric oxide synthase inhibition in male Wistar rats (aged 10 wk). Serial analyses demonstrated that the renal injury and inflammation in L-NAME-treated rats was associated with elevation of both Ang II and aldosterone. To investigate the direct effect of aldosterone on the renal injury, we conducted adrenalectomy (ADX) and aldosterone supplementation in L-NAME-treated rats. In ADX rats, aldosterone was undetectable, and renal injury and inflammation were almost completely prevented by ADX, although systemic and local Ang II and blood pressure were still elevated. Aldosterone supplementation reversed the beneficial effect of ADX. The present study indicates that aldosterone rather than Ang II plays a central and direct role in the pathogenesis of renal injury by L-NAME through inflammation, independent of its systemic hemodynamic effects.

Acute and Chronic Inhibition of NOS Causes a Switch in Vasodilator Mechanism from Nitric Oxide to Hydrogen Peroxide in the Human Microcirculation

This study tests the hypothesis that inhibition of NO-synthase (NOS) increases mitochondria-derived (mt) reactive oxygen species (ROS), resulting in changes in the mediator of flow-induced dilation (FID). In healthy individuals nitric oxide (NO) is the primary vasodilator responsible for FID, while in microvessels (MV) of patients with coronary artery disease (CAD) mtROS are responsible. NO inhibits mtROS and reduces smooth muscle proliferation to contribute to prevention of atherosclerosis. Hydrogen peroxide (H2O2)promotes cell proliferation and is pro-atherosclerotic. MV (~200µm) obtained from surgical discarded adipose were dissected and cannulated on glass pipettes in a heated organ chamber, perfused with Krebs and aerated with 74% N2, 21% O2, 5% CO2. After equilibration, MV were constricted with endothelin-1 and responses to graded increases in flow were recorded. Papaverine (100µM) was used to assess endothelial independent relaxation. Neither 1h nor 16h incubation with L-NAME (100µM before and during experiment) affected the FID compared to Vehicle (V) (% Max dilation: V 76.1 ±5.1; L-NAME: 1h 84.3 ±3; 16h: 87.8±2.7 P< 0.05 ANOVA). In V treated MV FID was L-NAME inhibitable (10.1±8.3) but not effected by PEG-cat while with L-NAME pretreatment PEG-cat inhibited dilation (L-NAME: 1h +Peg-Cat 18.4±12.9 16h + Peg-Cat 12.6 ±1.6). Our results suggest that inhibition of NOS in MV from healthy individuals causes a primary shift in the mediator of dilation from NO to H2O2 , similar to subjects with CAD. Our data are consistent with the concept that that threshold levels of NO are necessary to suppress mtROS in human MV. Loss of NO bioavailability causes a compensatory increase in mtROS, which mediates FID.

The optimization of a chronic nitric oxide synthase (NOS) inhibition model of pre-eclampsia by evaluating physiological changes

ObjectivesIn order to address the gap in our understanding of the pathogenesis and pathophysiology of PE, we optimized the NOS inhibition animal model by comparing changes in different parameters at various time frames during pregnancy, in both early and late-onset PE. Study Design120 nulliparous Sprague-Dawley rats were divided into 5 groups (n=24). A pregnant control, two groups that represented early and late-onset PE and two groups that were treated with sildenafil citrate (SC) to show reversal of the pre-eclamptic-like symptoms. ResultsOur results showed that treatment with l-NAME caused significant changes in physiological parameters for both early and late-onset PE groups. There was a significant increase in systolic blood pressure (SBP) levels in the early-onset PE group (128.5±5.71mmHg) and late-onset PE group (128.3±6.15mmHg) on day 19 compared to the SBPs on day 0, (p<0.01). Urine excretion volumes in the early-onset PE group (13.62±3.18mL) and in the late-onset PE (13.28±2.60mL), compared to the pregnant control group (11.96±1.9mL) were also increased (p<0.05). There was also an increase in total urinary protein in the early-onset PE group (0.62±0.08g/L and the late-onset PE group (0.45±0.05g/L), when compared to the pregnant control group (0.38±0.07) (p<0.05). We also found a decrease in fetal numbers in the PE group in comparison to the pregnant control and SC treated groups. The remission of these signs was seen after delivery of the fetuses. We also demonstrated that treatment of this syndrome with SC prevented the development of these signs. ConclusionsThe NOS inhibition model can be used for the study of the pathogenesis and pathophysiology of PE, since the pathogenic changes mimic those of early and late-PE.

Differential regulation of adipose tissue and vascular inflammatory gene expression by chronic systemic inhibition of NOS in lean and obese rats.

We tested the hypothesis that a decrease in bioavailability of nitric oxide (NO) would result in increased adipose tissue (AT) inflammation. In particular, we utilized the obese Otsuka Long Evans Tokushima Fatty rat model (n = 20) and lean Long Evans Tokushima Otsuka counterparts (n = 20) to determine the extent to which chronic inhibition of NO synthase (NOS) with Nω‐nitro‐l‐arginine methyl ester (L‐NAME) treatment (for 4 weeks) upregulates expression of inflammatory genes and markers of immune cell infiltration in retroperitoneal white AT, subscapular brown AT, periaortic AT as well as in its contiguous aorta free of perivascular AT. As expected, relative to lean rats (% body fat = 13.5 ± 0.7), obese rats (% body fat = 27.2 ± 0.8) were hyperlipidemic (total cholesterol 77.0 ± 2.1 vs. 101.0 ± 3.3 mg/dL), hyperleptinemic (5.3 ± 0.9 vs. 191.9 ± 59.9 pg/mL), and insulin‐resistant (higher HOMA IR index [3.9 ± 0.8 vs. 25.2 ± 4.1]). Obese rats also exhibited increased expression of proinflammatory genes in perivascular, visceral, and brown ATs. L‐NAME treatment produced a small but statistically significant decrease in percent body fat (24.6 ± 0.9 vs. 27.2 ± 0.8%) and HOMA IR index (16.9 ± 2.3 vs. 25.2 ± 4.1) in obese rats. Further, contrary to our hypothesis, we found that expression of inflammatory genes in all AT depots examined were generally unaltered with L‐NAME treatment in both lean and obese rats. This was in contrast with the observation that L‐NAME produced a significant upregulation of inflammatory and proatherogenic genes in the aorta. Collectively, these findings suggest that chronic NOS inhibition alters transcriptional regulation of proinflammatory genes to a greater extent in the aortic wall compared to its adjacent perivascular AT, or visceral white and subscapular brown AT depots.

Chapter 12 - Chronic nitric oxide synthase inhibition does not impair upper airway muscle adaptation to chronic intermittent hypoxia in the rat

Nitric oxide (NO) is an important modulator of striated muscle function. Nitric oxide synthase (NOS) expression and activity is altered by hypoxia and NO is implicated in respiratory muscle remodeling following chronic sustained hypoxia. We sought to determine if NO is implicated in upper airway dilator muscle adaptation to chronic intermittent hypoxia (CIH). Thirty-two adult male Wistar rats (284±13, mean±SD) were exposed to alternating bouts of hypoxia (90 s; 5% O2 at the nadir) and normoxia (210 s; 21% O2) for 12 cycles per hour, 8h/day for 3 weeks. Sham animals were exposed to normoxia in parallel. Half of the animals in both groups received the nNOS inhibitor-L-NNA (2mM) in the drinking water throughout the study (N=8 for all groups). Sternohyoid (pharyngeal dilator) muscle contractile and endurance properties were determined ex vivo. Sternohyoid muscle myosin heavy chain (MHC) isoform composition and cross-sectional area was determined by fluorescence microscopy. Chronic nNOS blockade did not alter sternohyoid muscle peak force or force-frequency relationship in sham or CIH-treated animals. In contrast, chronic nNOS blockade significantly decreased sternohyoid muscle endurance with equivalent effects in sham and CIH-treated rats. Our results suggest that NO is an important modulator of sternohyoid muscle endurance. However, our data provide no evidence to suggest that NO is implicated in upper airway muscle adaptation to CIH.

Reverse myocardial effects of intermedin in pressure-overloaded hearts: role of endothelial nitric oxide synthase activity.

Intermedin (IMD) is a cardiac peptide synthesized in a prepro form, which undergoes a series of proteolytic cleavages and amidations to yield the active forms of 47 (IMD(1-47)) and 40 amino acids (IMD(8-47)). There are several lines of evidence of increased IMD expression in rat models of cardiac pathologies, including congestive heart failure and ischaemia; however, its myocardial effects upon cardiac disease remain unexplored. With this in mind, we investigated the direct effects of increasing concentrations of IMD(1-47) (10(-10) to10(-6) m) on contraction and relaxation of left ventricular (LV) papillary muscles from two rat models of chronic pressure overload, one induced by transverse aortic constriction (TAC), the other by nitric oxide (NO) deficiency due to chronic NO synthase inhibition (NG-nitro-l-arginine, l-NAME), and respective controls (Sham and Ctrl). In TAC and l-NAME rats, exogenous administration of IMD(1-47) elicited concentration-dependent positive inotropic and lusitropic effects. By contrast, in Sham and Ctrl rats, IMD(1-47) induced a negative inotropic response without a significant effect on relaxation. Both TAC and l-NAME rats presented LV hypertrophy, elevated LV systolic pressures, preserved systolic function and elevated peroxynitrite levels. In the normal myocardium (Ctrl and Sham), IMD(1-47) induced a 3-fold increase of endothelial nitric oxide synthase (eNOS) phosphorylation at Ser(1177), indicating enhanced eNOS activity. In TAC and l-NAME rats, eNOS phosphorylation was increased at baseline, and its response to IMD(1-47) was blunted. In addition, the distinct myocardial response to IMD(1-47) was accompanied by distinct subcellular mechanisms. While in Sham rats the addition of IMD(1-47) induced the phosphorylation of cardiac troponin I due to NO/cGMP activation, in TAC rats IMD(1-47) induced phospholamban phosphorylation possibly associated with cAMP/protein kinase A activation. Therefore, we demonstrated for the first time a reversed myocardial response to IMD(1-47) neurohumoral stimulation due to impairment of eNOS activation in TAC and l-NAME rats. These results not only reveal the distinct myocardial effects and subcellular mechanisms for IMD(1-47) in normal and hypertrophic hearts, but also highlight the potential pathophysiological relevance of cardiac endothelial dysfunction in neurohumoral myocardial action.

Protective Effect of Aerobic Exercise against Some of Proinflammatory Cytokines-Induced Chronic Nitric Oxide Synthase Inhibition in Renal Tissue Rats

Protective Effect of Aerobic Exercise against Some of Proinflammatory Cytokines-Induced Chronic Nitric Oxide Synthase Inhibition in Renal Tissue Rats