Sodium sensitivity of arterial blood pressure in L-NAME hypertensive but not eNOS knockout mice

The present studies determined the sensitivity of mean arterial pressure (MAP) to sodium intake in endothelial nitric oxide synthase (eNOS) knockout mice, wild-type mice (C56BL/6J), and wild-type mice intravenously administered the nonspecific NOS inhibitor N(G)-nitro-l-arginine methyl ester (L-NAME, 8.6 mg/kg/d). Arterial blood pressure was measured from chronically implanted femoral arterial catheters in conscious, freely moving mice. The MAP was unaltered from the low sodium ( approximately 200 microEq/d) intake level of 106 +/- 3 mm Hg in wild-type mice when sodium intake was increased to approximately 1000 microEq/d (n = 12). Chronic administration of L-NAME to wild-type mice led to a sodium-dependent increase in MAP from 111 +/- 7 mm Hg to 124 +/- 5 mm Hg when the mice were placed on an elevated sodium intake (n = 7). In contrast to the L-NAME-treated mice, MAP was unaltered in eNOS knockout mice (n = 8) when sodium intake was increased (128 +/- 3 mm Hg v 129 +/- 5 mm Hg). These experiments demonstrate that eNOS knockout and L-NAME-treated wild-type mice are hypertensive relative to wild-type controls when sodium intake is elevated, but only L-NAME-treated mice exhibited a sodium-sensitive increase in MAP. Therefore, nitric oxide produced by eNOS does not appear to be important in the physiologic adaptation to elevated sodium chloride intake.     

Structure of cerebral arterioles in mice deficient in expression of the gene for endothelial nitric oxide synthase

We examined effects of pharmacological inhibition of nitric oxide synthase (NOS) and genetic deficiency of the endothelial isoform of NOS (eNOS) on structure and mechanics of cerebral arterioles. We measured pressure, diameter, and cross-sectional area (CSA) of the vessel wall (histologically) in maximally dilated cerebral arterioles in mice that were untreated or treated for 3 months with the NOS inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME; 10 mg/kg per day in drinking water). Treatment with L-NAME increased systemic arterial mean pressure (SAP; 143+/-4 versus 121+/-4 mm Hg, P<0.05) and CSA (437+/-27 versus 310+/-34 microm2, P<0.05). These findings suggest that hypertension induced in mice by NOS inhibition is accompanied by hypertrophy of cerebral arterioles. To determine the role of the eNOS isoform in regulation of cerebral vascular growth, we examined mice with targeted disruption of one (heterozygous) or both (homozygous) genes encoding eNOS. Wild-type littermates served as controls. SAP and CSA were significantly increased in homozygous (SAP, 141+/-5 versus 122+/-3 mm Hg in wild-type mice, P<0.05; CSA, 410+/-18 versus 306+/-15 microm2 in wild-type mice, P<0.05), but not in heterozygous (SAP, 135+/-4 mm Hg; CSA, 316+/-32 microm2) eNOS-deficient mice. Carotid ligation normalized cerebral arteriolar pulse pressure did not prevent increases in CSA in homozygous eNOS-deficient mice. Thus, cerebral arterioles undergo hypertrophy in homozygous eNOS-deficient mice, even in the absence of increases in arteriolar pulse pressure. These findings suggest that eNOS plays a major role in regulation of cerebral vascular growth.     

PPAR(gamma) agonist rosiglitazone improves vascular function and lowers blood pressure in hypertensive transgenic mice

The peroxisome proliferator activated receptor (PPARgamma) agonist rosiglitazone has been reported to yield cardiovascular benefits in patients by a mechanism that is not completely understood. We tested whether oral rosiglitazone (25 mg/kg per day, 21 days) treatment improves blood pressure and vascular function in a transgenic mouse expressing both human renin and human angiotensinogen transgenes (R(+)A(+)). Rosiglitazone decreased systolic (138+/-5 versus 128+/-5 mm Hg) and mean blood pressure (145+/-5 versus 126+/-7 mm Hg) of R(+)A(+) mice as measured by tail-cuff and indwelling carotid catheters, respectively. Relaxation of carotid arteries to acetylcholine and authentic nitric oxide, but not papaverine, was impaired in R(+)A(+) mice when compared with littermate controls (RA(-)). There were no effects of rosiglitazone on RA(-) mice; however, relaxation to acetylcholine (49+/-10 versus 82+/-9% at 100 micromol/L) and nitric oxide (51+/-11 versus 72+/-6% at 10 micromol/L) was significantly improved in treated R(+)A(+) mice. Rosiglitazone treatment of R(+)A(+) mice did not alter the expression of genes, including endothelial nitric oxide synthase (eNOS), angiotensin 1 receptors, and preproendothelin-1, nor did it alter the levels of eNOS or soluble guanylyl cyclase protein. In separate studies, carotid arteries from R(+)A(+) and RA(-) mice relaxed in a concentration-dependent manner to rosiglitazone, suggesting possible PPARgamma-independent effects in the vasculature. This response was not inhibited with the nitric oxide synthase inhibitor N(omega)-nitro-l-arginine methyl ester (200 micromol/L) or the PPARgamma antagonist bisphenol A diglycidyl ether; 4,4'-isopropylidenediphenol diglycidyl ether (100 micromol/L). These data suggest that in addition to potential genomic regulation caused by PPARgamma activation, the direct effect of rosiglitazone in blood vessels may contribute to the improved blood pressure and vessel function.     

The central hypotensive effect induced by alpha 2-adrenergic receptor stimulation is dependent on endothelial nitric oxide synthase

OBJECTIVE: The aim of the present study was to determine whether the central antihypertensive effect of drugs that act via central alpha 2-adrenergic receptors is mediated by the nitric oxide-ergic system. METHODS: The hypotensive effects of dexmedetomidine, a 'pure' alpha2-adrenergic agonist, were compared in endothelial nitric oxide synthase knockout and wild-type control mice. RESULTS: When injected intravenously (5 mug/kg) in wild-type mice, dexmedetomidine elicited a depressor response (60 +/- 4 to 34 +/- 1 mmHg, P < 0.05), but had no hypotensive effect in endothelial nitric oxide synthase (eNOS) knockout mice (84 +/- 7 to 84 +/- 7 mmHg, P > 0.05). In the presence of N-omega-nitro-L-arginine, a nonselective nitric oxide synthase (NOS) blocker that does not cross the blood-brain barrier, the hypotensive effect of dexmedetomidine was not abolished (Delta MAP = 21 +/- 2 mmHg vs. Delta MAP = 26 +/- 3 mmHg, P > 0.05). CONCLUSIONS: It is concluded that the central cardiovascular effects of alpha 2-adrenergic agonists, such as dexmedetomidine, require an intact expression of eNOS within the brain. This study raises the interesting question of whether central eNOS itself might be considered as a target for new cardiovascular drugs regardless of any activation of alpha 2-adrenergic receptors.     

Endothelial NO/cGMP system contributes to natriuretic peptide-mediated coronary and peripheral vasodilation

We tested the hypothesis that the endothelial nitric oxide (NO)-soluble guanylyl cyclase system is involved in atrial natriuretic peptide (ANP) and C-type natriuretic peptide (CNP) mediated regulation of coronary and peripheral vascular resistance. Rat hearts were perfused via the aorta at constant flow and the effect of ANP and CNP on coronary perfusion pressure and release of cGMP was determined in the absence and presence of the nitric oxide synthase inhibitor NG-nitro-L-arginine (L-NNA; 0.2 mmol/L) and the specific inhibitor of soluble guanylyl cyclase ODQ (20 micromol/L), respectively (n = 6). ANP (10-300 nmol/L) reduced perfusion pressure from 133 +/- 2 to 53 +/- 2 mm Hg (-60%; control) in the presence of L-NNA from 132 +/- 1 to 71 +/- 1 mm Hg (-46%) and in the presence of ODQ from 133 +/- 1 to 85 +/- 2 (-36%) (n = 6; P < 0.05). Disruption of the coronary endothelium by perfusion of hearts with collagenase reduced the relaxant effect of ANP to a similar extent as L-NNA. Basal release of cGMP was increased up to sixfold by ANP and this increase was reduced by L-NNA and ODQ (n = 6; P < 0.05). The coronary relaxant effect of CNP (0.1-3 micromol/L) was similarly attenuated by L-NNA and ODQ (n = 6). In conscious mice, a low dose of L-NNA (30 nmol) consistently reduced the blood pressure lowering effect of ANP (30 nmol) by approximately 40% (n = 7), whereas the hypotensive effect of nitroprusside (0.15 micromol) was not affected (n = 5). We conclude that the coronary dilatory and hypotensive action of natriuretic peptides involves the endothelium and is partly mediated by soluble guanylyl cyclase. The data may explain previous observations in humans with congestive heart failure showing impaired vascular ANP responses.     

Chronic blockade of nitric oxide does not produce hypertension in baroreceptor denervated rabbits

Although the vascular action of endothelium-derived nitric oxide in modulating arterial pressure is well established, nitric oxide can also act as a neurotransmitter in the central nervous system. In addition, there is evidence for an interaction between nitric oxide and baroreceptor afferent processing; thus, nitric oxide may regulate blood pressure through central modulation of arterial baroreflexes. To test this possible interaction of nitric oxide and baroreflexes in the long-term regulation of blood pressure, we measured arterial pressure and heart rate responses to nitric oxide blockade by using L-NAME (50 mg/kg per day in drinking water) over 7 days in baroreceptor intact and sinoaortic denervated conscious rabbits. In the baroreceptor intact animals, blockade of nitric oxide leads to a significant increase in mean arterial pressure (from 75+/-2 to 84+/-3 mm Hg) and decrease in heart rate (from 233+/-8 to 195+/-8 bpm) that was sustained over the 7 days of nitric oxide blockade. In the sinoaortic denervated animals, blockade of nitric oxide initially led to a similar increase in arterial pressure (82+/-3 mm Hg on the second day), but in all sinoaortic denervated animals this increase was not sustained and recovered back to pre-L-NAME levels. This finding indicates that baroreflexes play an important role in the long-term control of blood pressure, and, second, that one mediator of this control is nitric oxide.     

Vascular function and blood pressure in GH transgenic mice.

Acromegaly is associated with cardiovascular disease. We studied vascular function and mean arterial blood pressure in transgenic mice overexpressing bovine GH. Mean arterial blood pressure was measured in conscious, unrestrained male and female bovine GH and littermate control mice during normal as well as high salt intake using telemetric devices. Structure in artificially perfused maximally dilated hindquarter vascular beds and vascular reactivity and endothelial function in small mesenteric vessels were studied in female bovine GH and control mice. Mean arterial blood pressure was increased in female bovine GH transgenic (126 +/- 3 mm Hg) and male bovine GH transgenic (129 +/- 4 mm Hg) compared with female (109 +/- 3 mm Hg, P < 0.05) and male (111 +/- 3 mm Hg, P < 0.05) controls respectively. Increased salt intake had no effect on mean arterial blood pressure. Perfusion studies showed a significant decrease in the average diameter of the female bovine GH transgenic hindquarter vascular bed (P < 0.05). The responses of isolated resistance arteries to nor-epinephrine, potassium-induced depolarization, acetylcholine, or sodium-nitroprusside did not significantly differ between bovine GH transgenic and control mice. We conclude that the phenotype of the bovine GH transgenic mice includes a salt-resistant form of hypertension. Furthermore, the increase in mean arterial blood pressure is accompanied by a significant structural narrowing of the resistance vasculature without changes in vascular reactivity or endothelial function. The results imply that hypertension in bovine GH transgenic mice is maintained mainly by a structurally based increase in peripheral vascular resistance.     

Cardiovascular and renal phenotype in mice with one or two renin genes

We compared the phenotype of two common mouse models, C-57BL/6J (C57), which carries only the Ren-1c gene, and 129/SvJ (Sv-129), with both Ren 1d and Ren-2. We hypothesized two renin gene Sv-129 would have increased blood pressure and the renin-angiotensin system would be more influential in regulating renal function compared with one renin gene mice. Sv-129 consistently had higher blood pressure than C-57, whether conscious (128 versus 108 mm Hg, P<0.001) or anesthetized (102 versus 88 mm Hg, P<0.001). Plasma renin concentration in both conscious and anesthetized C-57 mice was 3- to 4-fold higher than in Sv-129 (P<0.05), whereas renal cortical renin content was 2.5-fold higher (P<0.005). Renal blood flow and renal vascular resistance were the same in C-57 and Sv-129. Exogenous angiotensinogen produced identical pressor and renal vasoconstrictor responses in both strains. Blocking AT1 receptors with losartan reduced blood pressure by 19 mm Hg in both strains. Nitric oxide synthase inhibition by l-NAME increased blood pressure by 29 mm Hg in C-57 and 35 mm Hg in Sv-129 mice, but the decrease in renal blood flow was 30% less in C-57 (P<0.025). We conclude that Sv-129 mice with two renin genes have higher blood pressure but lower plasma and renal renin than C-57 mice with one renin gene. Renin substrate may limit angiotensin II production in the mouse. In Sv-129, the influence of nitric oxide on renal but not systemic resistance may be exaggerated. Renin from Ren-2 may act independently of normal renin control mechanisms.     

Neuronal nitric oxide synthase-deficient mice have impaired renin release but normal blood pressure

BACKGROUND: Nitric oxide deficiency is involved in the development of hypertension, but the mechanisms are currently unclear. This study was conducted to further elucidate the role of neuronal nitric oxide synthase (nNOS) in blood pressure regulation and renin release in relation to different sodium loads. METHODS: Blood pressure and heart rate were measured telemetrically and assessed during periods of physical activity and inactivity. Urinary solute excretion was measured by metabolism cages and plasma renin concentration (PRC) was determined by radioimmunoassay; all in nNOS knockout (nNOS(-/-)) and wild-type (nNOS(+/+)) mice after 10 days of low (0.01% NaCl) and high (4% NaCl) sodium diets. RESULTS: The resting heart rate was reduced in nNOS(-/-) mice, but the two genotypes had similar blood pressure during the low (nNOS(+/+) 104 +/- 2 mm Hg; nNOS(-/-) 103 +/- 2 mm Hg) and high (nNOS(+/+) 107 +/- 3 mm Hg; nNOS(-/-) 108 +/- 2 mm Hg) sodium diets. During the high sodium diet, PRC did not differ between the genotypes (nNOS(+/+) 743 +/- 115 10(-5) Goldblatt units; nNOS(-/-) 822 +/- 63 10(-5) Goldblatt units), but during the low sodium diet, nNOS(-/-) mice failed to increase PRC (nNOS(+/+) 2164 +/- 220 10(-5) Goldblatt units; nNOS(-/-) 907 +/- 101 10(-5) Goldblatt units) and renal renin mRNA. On the low sodium diet, nNOS(-/-) mice also showed increased urine flow rate and osmolar excretion, observations not made during a high sodium diet. CONCLUSIONS: Our results show that nNOS is necessary for stimulation of renin in response to sodium restriction. Furthermore, nNOS(-/-) mice are normotensive, and their blood pressure responds normally to an increased dietary sodium intake, indicating that nNOS deficiency does not cause salt-sensitive hypertension.     

Intraportal administration of glyceryl trinitrate or nitroprusside exerts more systemic than intrahepatic effects in anaesthetised cirrhotic rats.

BACKGROUND/AIMS: Increased intrahepatic vascular tone can be pharmacologically manipulated in isolated cirrhotic livers. Intrahepatic endothelial dysfunction may lead to a decreased production of the potent endogenous vasodilator nitric oxide in cirrhotic livers. The aims of the study were to determine whether portal pressure can be lowered in vivo by injecting nitric oxide donors glyceryl trinitrate or nitroprusside directly in the portal vein and whether this is related to a decrease in intrahepatic resistance. METHODS: In anaesthetised CCl4 cirrhotic rats, intraportal doses of glyceryl trinitrate 0.5, 1 or 5 microg/kg/ min or nitroprusside 1, 5 or 10 microg/kg/min did not decrease portal pressure but only arterial pressure. Systemic and splanchnic haemodynamics were measured before and during 15 min intraportal infusion of glyceryl trinitrate 10 microg/kg/min or nitroprusside 20 microg/kg/min. RESULTS: Glyceryl trinitrate decreased portal pressure from 14.0+/-1.1 to 11.8+/-1.4 mm Hg, splanchnic perfusion pressure from 102+/-10 to 74+/-5 mm Hg and portal sinusoidal flow from 2.11+/-0.38 to 1.70+/-0.35 ml/min/g liver (all p<0.05). Nitroprusside did not decrease portal pressure significantly but led to a reduction of the splanchnic perfusion pressure (104+/-9 to 66+/-7 mm Hg) and the portal sinusoidal flow (2.39+/-0.50 to 1.77+/-0.31 ml/min/g liver; all p<0.05). Portal sinusoidal resistance was not altered by either drug. CONCLUSIONS: Intraportal infusion of nitric oxide donors decreased arterial pressure more than portal pressure. Portal sinusoidal resistance remained unaffected, but the liver parenchyma became less perfused with high doses. The systemic effects of nitric oxide donating drugs prevailed.     

Endothelial nitric oxide gene knockout mice: cardiac phenotypes and the effect of angiotensin-converting enzyme inhibitor on myocardial ischemia/reperfusion injury.

We tested the hypothesis that nitric oxide (NO) released by endothelial NO synthase (eNOS) is not only important in blood pressure regulation but also involved in cardiac function and remodeling and in the cardioprotective effect of angiotensin-converting enzyme inhibitors (ACEi). With the use of a 2D Doppler echocardiography system equipped with a 15-MHz linear transducer, we evaluated left ventricular (LV) morphology and function in conscious eNOS knockout mice (eNOS(-/-); n=15) and their wild-type littermates (eNOS(+/+); n=16). We also studied whether in eNOS(-/-) mice (1) myocardial ischemia/reperfusion injury is more severe and (2) the cardioprotective effect of ACEi is diminished or absent. In comparison with the wild type, eNOS(-/-) mice had significantly increased systolic blood pressure (128+/-3 versus 108+/-5 mm Hg; P<0.001) and decreased heart rate (531+/-22 versus 629+/-18 bpm; P<0.001) associated with increased LV posterior wall thickness (0.80+/-0.04 versus 0.64+/-0.02 mm; P<0.001) and LV mass (18.3+/-0.9 versus 13.1+/-0.5 mg/10 g body weight; P<0.01). Despite hypertension and LV hypertrophy, LV chamber dimension, shortening fraction and ejection fraction (indicators of LV contractility), and cardiac output did not differ between the 2 strains, which indicates that LV function in eNOS(-/-) mice is well compensated. We also found that in eNOS(+/+) mice, ACEi decreased the ratio of myocardial infarct size to area at risk from 62.7+/-3.9% to 36.3+/-1.6% (P<0. 001), whereas in eNOS(-/-) mice this effect of ACEi was almost abolished: the ratio of myocardial infarct size to area at risk was 67.2+/-2.9% in the vehicle-treated group and 62.7+/-3.9% in mice treated with ACEi. Moreover, infarct size in vehicle-treated eNOS(-/-) mice was not significantly different from eNOS(+/+) mice given the same treatment. We concluded that (1) endothelium-derived NO plays an important role in the regulation of blood pressure homeostasis; (2) NO released under basal conditions has no significant impact on cardiac function; and (3) ACEi protect the heart against ischemia/reperfusion injury in mice and that this effect is mediated in part by endothelium-derived NO.     

Reduced production of nitric oxide during haemodialysis.

OBJECTIVE: To examine the influence of systemic nitric oxide (NO) synthesis on blood pressure in patients with chronic renal failure undergoing haemodialysis, since nitric oxides are susceptible to renal excretion or are dialysed, a different indicator that is unaffected by renal function, such as the level of exhaled NO was evaluated. We examined the levels of the endogenous NO before and after a haemodialysis session. DESIGN AND METHODS: We evaluated the serum concentrations of nitrite/nitrate and the rate of nitric oxide release into exhaled air in 10 patients with hypertension who were receiving maintenance haemodialysis. RESULTS: The serum concentrations of nitrite/nitrate before haemodialysis were significantly higher than those in 10 normal controls (183 +/- 151 microM vs 42 +/- 17 microM, P < 0.05). These levels decreased significantly by the end of haemodialysis (42 +/- 26 microM). Because the amount of nitric oxide in the deepest expirate correlated well with the duration of exhalation, we were able to derive the rate of release of NO. The rate of NO release was 0.034 +/- 0.012 nmol/sec before haemodialysis, similar to that in normal controls (0.031 +/- 0.013nmol/sec). The rate was significantly reduced after dialysis (0.023 +/- 0.010 nmol/sec) (P < 0.05). The mean pre-dialysis mean blood pressure (109 +/- 11 mm Hg) and the post-dialysis blood pressure (106 +/- 9 mm Hg) were the same. CONCLUSIONS: These data indicate that NO production does not appear to have a critical role in control of arterial blood pressure across haemodialysis in patients with chronic renal failure.     

Control of blood pressure variability in caveolin-1-deficient mice: role of nitric oxide identified in vivo through spectral analysis

AIMS: In endothelial cells, caveolin-1 (cav-1) is known to negatively modulate the activation of endothelial nitric oxide synthase, a key regulator of blood pressure (BP). However, the impact of genetic alteration of cav-1 on vascular nitric oxide (NO) production and BP homeostasis in vivo is unknown. METHODS AND RESULTS: We used spectral analysis of systolic blood pressure (SBP) variability in mice chronically equipped with telemetry implants to identify frequency ranges (0.05-0.4 Hz; very low frequency, VLF) specifically responding to NO, independently of changes in absolute BP or systemic neurohormone levels. VLF variability was inversely correlated to aortic vasodilator-stimulated Ser(239) phosphoprotein (VASP) phosphorylation, reflecting NO bioactivity. We show that mice deficient in cav-1 have decreased VLF variability paralleled with enhanced systemic and vascular production of NO at unchanged mean SBP levels. Conversely, VLF variability was increased upon acute injection of mice, with a peptide containing the caveolin-scaffolding domain (CSD; residues 82-101) fused to an internalization sequence of antennapedia that decreased vascular and circulating NO in vivo. CONCLUSION: These data highlight the functional importance of cav-1 for the production of bioactive NO in conduit arteries and its control of central BP variability. Given the impact of the latter on target organ damage, this raises the interest for genetic, pharmacological, or molecular interventions that modulate cav-1 expression in diseases with NO-dependent endothelial dysfunction.     

Pressure-induced cardiac overload induces upregulation of endothelial and myocardial progenitor cells

AIM: The regulation of angiogenesis in the hypertrophied overloaded heart is incompletely understood. Bone-marrow-derived progenitor cells have been shown to contribute to endothelial homeostasis, repair, and new blood vessel formation. We therefore studied the effects of pressure overload on angiogenesis and progenitor cells. METHODS AND RESULTS: Pressure overload induced by transaortic constriction (TAC, C57/Bl6 mice, 360 microm for 35 days) increased left ventricular (LV) systolic pressure, the ratio of heart weight to tibia length, cardiomyocyte diameters, and cardiac apoptosis and fibrosis compared to sham-operated mice. In the TAC group, the number of cycling Ki67 pos cells increased from none to 0.1 +/- 0.02% in cardiomyocytes and from 0.17 +/- 0.02% to 0.65 +/- 0.1% in non-cardiomyocytes, P < 0.001. stem cell antigen 1(pos)/vascular endothelial growth factor receptor 2 pos endothelial progenitor cells (EPC) increased to 210 +/- 25% in the blood and to 196 +/- 21% in the bone marrow (P < 0.01). TAC upregulated cultured spleen-derived DiLDL pos/lectin pos EPC to 221 +/- 37%, P < 0.001. Cardiac hypertrophy and upregulation of EPC secondary to cardiac pressure overload were associated with increased extra-cardiac neoangiogenesis (54 +/- 12% increase, P < 0.05). In endothelial nitric oxide synthase double knockout mice, the upregulation of EPC by TAC was abolished. Maladaptive myocardial remodelling in TAC mice was characterized by a reduction of CD31 pos cells. In mice transplanted with green fluorescent protein pos bone marrow, TAC markedly increased myocardial bone marrow-derived CD31 pos cells from 2.37 +/- 0.4% to 7.76 +/- 1.5% and MEF2 pos cells from 1.8 +/- 0.4/mm2 to 20.5 +/- 5.3/mm2, P < 0.05. CONCLUSION: Pressure-induced myocardial hypertrophy leads to upregulation of systemic EPCs, increased extra-cardiac angiogenesis, and upregulation of intra-myocardial bone marrow-derived endothelial and myocyte precursor cells. The data show that afterload-dependent regulation of bone marrow-derived progenitor cells contributes to angiogenesis in myocardial hypertrophy.     

L-arginine attenuates hypertension in pregnant rats with reduced uterine perfusion pressure

A chronic reduction in uterine perfusion pressure in the pregnant rat is associated with significant elevations in mean arterial pressure, proteinuria, and reductions in kidney function as is chronic nitric oxide blockade, suggesting that nitric oxide deficiency may contribute to the clinical manifestations of preeclampsia. The purpose of this study was to determine whether supplementation with L-arginine, the precursor for nitric oxide, attenuates the hypertension produced in response to a chronic reduction in uterine perfusion pressure in the pregnant rat. Reduced uterine perfusion was initiated at day 14 of gestation with arterial pressure determined at day 19 of gestation in conscious, chronically instrumented rats. Arterial pressure was significantly elevated in pregnant rats with chronic reductions in uterine perfusion as compared with pregnant control rats (132+/-2 versus 109+/-2 mm Hg, P<0.01, respectively). Treatment with L-arginine (2%) in the drinking water was initiated at day 10 of gestation. l-arginine supplementation resulted in a significant decrease in arterial pressure in both pregnant rats with reduced uterine perfusion pressure (113+/-2 mm Hg treated, P<0.01 versus untreated pregnant with reduced uterine perfusion pressure) and pregnant control (97+/-3 mm Hg treated, P<0.01 versus untreated pregnant) rats. However, supplementation with L-arginine decreased blood pressure by 19 mm Hg in pregnant with reduced uterine perfusion pressure (untreated versus treated) as compared with 12 mm Hg in pregnant (untreated versus treated) rats. Thus, these results suggest that l-arginine supplementation may be beneficial in attenuating the hypertension in preeclampsia.     

Premature death and age-related cardiac dysfunction in male eNOS-knockout mice

The aims of our study were to determine mortality, and age- and genotype-related cardiac phenotype in endothelial nitric oxide synthase (NOS) knockout (-/-) and wild-type (+/+) mice. Male and female (-/-) and male and female (+/+) conscious mice were studied at different ages by echocardiography and tail-cuff blood pressure (BP) measurement. Only 50% male (-/-) mice lived longer than 21 months whereas 89% (+/+) mice were still alive after 24 months (P < 0.005). There was little mortality in female mice of either genotype. Both (-/-) and (+/+) male mice have normal cardiac dimensions and function at 5.5 months. However, (-/-) mice developed cardiac dilation and dysfunction at 21 months as evidenced by a significant increase (P < 0.05) in left ventricular (LV) end-diastolic diameter from 2.69 +/- 0.13 to 3.13 +/- 0.09 mm, LV end-systolic diameter from 1.28 +/- 0.11 to 1.86 +/- 0.12 mm, LV end-diastolic cavity volume from 21 +/- 2.8 to 31 +/- 2.5 microl and LV mass from 19 +/- 2.5 to 27 +/- 1.9 mg/10 g and a significant decrease (P < 0.05) in ejection fraction (from 65 +/- 3.3% to 41 +/- 4.6%), shortening fraction (from 53 +/- 2.2% to 41 +/- 3.4%), LV posterior wall thickening (from 27 +/- 2% to 12 +/- 4%) and septum thickening (from 27 +/- 2% to 12 +/- 4%) compared with those at 5.5 months. There was a clear increase in cardiac weight and cardiac dilation by hematoxylin and eosin in male (-/-) mice at 21 months. BP in male (-/-) mice fell with the cardiac dysfunction, whereas female (-/-) mice were hypertensive even at 21 months. The level of mRNA for neuronal NOS and inducible NOS was greater in all females compared to males. These results indicate that male (-/-) mice have a significantly shorter lifespan than (+/+) or female mice, and male (-/-) mice develop cardiac dysfunction with age.     

Assessment of renal functional phenotype in mice lacking gp91PHOX subunit of NAD(P)H oxidase

To determine the role of endogenous superoxide (O2-) in the kidney, we assessed renal hemodynamics and excretory function in gp91(PHOX) (a NAD(P)H oxidase subunit) gene knockout (KO) mice and compared these findings with those of wild-type (WT) strain C57BL/6 mice. Renal blood flow (RBF) and glomerular filtration rate (GFR) were determined by PAH and inulin clearances respectively in anesthetized mice (n=8 in each group). There were higher baseline RBF (4.3+/-0.4 versus 2.5+/-0.2 mL/min per gram; P<0.002) and lower renal vascular resistance (RVR) (16+/-1.4 versus 29+/-2.3 mm Hg/mL/min per gram; P<0.0001) in KO compared with WT without a significant difference in mean arterial pressure (MAP) (67+/-2 versus 71+/-2 mm Hg) and GFR (0.66+/-0.09 versus 0.73+/-0.05 mL/min per gram) between the strains. Intravenous infusion of angiotensin II (Ang II) (2 ng/min per gram of body weight) for 30 minutes caused a lesser degree of decreases in RBF (-8% versus -33%) and of increases in RVR (+73% versus +173%) in KO compared with WT. GFR was increased (43%) in KO but not in WT during Ang II infusion. Urinary excretion of nitrate/nitrite was higher in conscious KO (n=5) than in WT (n=5), indicating an increase in nitric oxide bioavailability that could be the cause of high RBF and low RVR in KO. These data indicate that gp91(PHOX), a subunit of NAD(P)H oxidase, plays a regulatory role in the maintenance of renal vascular tone. These results also suggest that the mechanism of Ang II-mediated renal vascular action involves concomitant generation of O2-.     

Endothelial function in normotensive and high-normal hypertensive subjects

To evaluate the impact of a mild increment in blood pressure level on endothelial function, we evaluated 61 healthy volunteers (24 women, 37 men, and aged 35-50 years). All subjects underwent a blood chemistry panel to exclude any metabolic abnormalities and were submitted to a Doppler ultrasound of the brachial artery to assess endothelial function. We assessed the endothelial response to reactive hyperaemia and exogenous nitric oxide administration considering an increase in systolic blood pressure (SBP) at each 10-mm Hg interval. Our study population was divided as follows: SBP <115 mm Hg (SG1, n=13), SBP > or =115 mm Hg and <125 mm Hg (SG2, n=20), SBP > or = 125 mm Hg and <135 mm Hg (SG3, n=13) and SBP > or = 135 mm Hg and < 140 mm Hg (SG4, n=15). We found a significant difference in flow-mediated dilation among SG2, SG3 and SG4, 16.2+/-5.6, 13.4+/-5.2 and 11.5+/-3.6%, P<0.05, respectively). After nitrate administration, we observed a nonsignificant decrease in brachial artery dilation among groups, P=0.217. Our data showed in a healthy normotensive population, without any risk factor for atherosclerotic disease that small increases in SBP but not in diastolic blood pressure may impair endothelial function even in subjects considered as high-normal, meaning that this population deserves more attention than usually ascribed to intervene and prevent complications, as endothelial dysfunction may represent an early change in those who develop hypertension later in life.     

Evidence for normal nitric oxide-mediated vasodilator tone in conscious rats with cirrhosis.

Because it has been hypothesized that the hyperkinetic circulation in portal hypertension is the result of increased synthesis of nitric oxide, we compared the hemodynamic effects of nitric oxide synthesis--specific agonist (L-arginine) and antagonist between normal and cirrhotic conscious rats. The dose-response curves showed that L-arginine significantly decreased arterial pressure and increased heart rate. These changes started at the 200 mg/kg dose and were similar in both groups of rats. In both groups of rats NG-monomethyl-L-arginine (25 mg/kg) significantly decreased cardiac output by 35%. In cirrhotic rats, NG-monomethyl-L-arginine decreased portal pressure from 15.3 +/- 0.9 mm Hg to 13.6 +/- 0.7 mm Hg and portal tributary blood flow from 7.8 +/- 0.7 ml.min-1.100 gm-1 to 5.9 +/- 0.7 ml.min-1.100 gm-1; it significantly increased portal territory vascular resistance from 950 +/- 108 dyn.sec.cm-5.100 gm-1 x 10(3) to 1,579 +/- 258 dyn.sec.cm-5.100 gm-1 x 10(3). In normal rats, portal tributary blood flow decreased similarly, by 27%, and portal territory vascular resistance increased by 55%. In neither group was hepatic arterial blood flow altered. Before and after NG-monomethyl-L-arginine administration, arterial cyclic GMP concentrations were not significantly different between normal and cirrhotic rats. In conclusion, this study shows evidence of a normal role for nitric oxide-mediated vasodilatation in rats with cirrhosis and that inhibition of nitric oxide synthesis reduces portal hypertension. These results did not support the hypothesis that nitric oxide synthesis is increased in cirrhosis.     

Dimethylarginine dimethylaminohydrolase promotes endothelial repair after vascular injury.

OBJECTIVES: We sought to determine if a reduction in asymmetric dimethylarginine (ADMA) enhances endothelial regeneration. BACKGROUND: Asymmetric dimethylarginine is an endogenous inhibitor of nitric oxide synthase (NOS). Increased plasma levels of ADMA are associated with endothelial vasodilator dysfunction in patients with vascular disease or risk factors. Asymmetric dimethylarginine is eliminated largely by the action of dimethylarginine dimethylaminohydrolase (DDAH), which exists in 2 isoforms. Dimethylarginine dimethylaminohydrolase-1 transgenic (TG) mice manifest increased DDAH activity, reduced plasma and tissue ADMA levels, increased nitric oxide synthesis, and reduced systemic vascular resistance. METHODS: The left femoral arteries of DDAH1 TG mice and wild-type (WT) mice were injured by a straight spring wire, and regeneration of the endothelial cell (EC) monolayer was assessed. Endothelial sprouting was assayed with growth factor-reduced Matrigel. RESULTS: Regeneration of the EC monolayer was more complete 1 week after injury in TG mice (WT vs. TG: 40.0 +/- 6.5% vs. 61.2 +/- 6.4%, p < 0.05). The number of CD45 positive cells at the injured sites was reduced by 62% in DDAH TG mice (p < 0.05). Four weeks after injury, the neointima area and intima/media ratio were attenuated in DDAH TG mice (WT vs. TG: 0.049 +/- 0.050 mm2 vs. 0.031 +/- 0.060 mm2, 3.1 +/- 0.5 vs. 1.7 +/- 0.2, respectively, p < 0.05). Endothelial cell sprouting from vascular segments increased in TG mice (WT vs. TG: 24.3 +/- 3.9 vs. 39.0 +/- 2.2, p < 0.05). CONCLUSIONS: We find for the first time an important role for DDAH in EC regeneration and in neointima formation. Strategies to enhance DDAH expression or activity might be useful in restoring the endothelial monolayer and in treating vascular disease.