Impaired Nitric Oxide Synthesis Research Articles

GTP cyclohydrolase I gene transfer reverses tetrahydrobiopterin deficiency and increases nitric oxide synthesis in endothelial cells and isolated vessels from diabetic rats.

Nitric oxide (NO) synthesis in endothelial cells is impaired in diabetes. We previously showed that impaired NO synthesis in the spontaneously diabetic BB (BBd) rat is due to decreased levels of tetrahydrobiopterin (BH4), secondary to decreased expression of GTP cyclohydrolase I (GTPCH). The aim of this study was to utilize adenoviral GTPCH gene transfer to reverse BH4 deficiency and repair the ability of endothelial cells to produce NO. GTPCH gene transfer increased BH4 levels in BBd endothelial cells from 0.17 +/- 0.02 (mean +/-SE) to 73.37 +/- 14.42 pmol/million cells and NO production from 0.77 +/- 0.07 to 18.74 +/- 5.52 nmol/24 h/million cells. To demonstrate a functional effect of increasing BH4 concentrations in tissues, we transferred GTPCH into aortic rings from BBd and Zucker diabetic fatty (ZDF) rats, models of human type I and type II diabetes, respectively. GTPCH gene transfer led to a dose-dependent increase in acetylcholine-induced vasorelaxation, preventable by inhibiting NO synthase. Maximal relaxation of virus-treated rings (10(10) virus particles/ml) to acetylcholine was significantly higher than sham-treated rings (BBd 64% vs. 37%, P<0.005; ZDF 80% vs. 44%, P<0.05). This study demonstrates that GTPCH gene transfer can reverse BH4 deficiency in both type I and type II diabetes and provides an experimental basis for using gene therapy to treat cardiovascular complications in diabetic patients.

Partial gene deletion of endothelial nitric oxide synthase predisposes to exaggerated high-fat diet-induced insulin resistance and arterial hypertension.

Nitric oxide (NO) plays a major role in the regulation of cardiovascular and metabolic homeostasis, as evidenced by insulin resistance and arterial hypertension in endothelial NO synthase (eNOS) null mice. Extrapolation of these findings to humans is difficult, however, because eNOS gene deficiency has not been reported. eNOS gene polymorphism and impaired NO synthesis, however, have been reported in several cardiovascular disease states and could predispose to insulin resistance. High-fat diet induces insulin resistance and arterial hypertension in normal mice. To test whether partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension during metabolic stress, we examined effects of an 8-week high-fat diet on insulin sensitivity (euglycemic clamp) and arterial pressure in eNOS(+/-) mice. When fed a normal diet, these mice had normal insulin sensitivity and were normotensive. When fed a high-fat diet, however, eNOS(+/-) mice developed exaggerated arterial hypertension and had fasting hyperinsulinemia and a 35% lower insulin-stimulated glucose utilization than control mice. The partial deletion of the eNOS gene does not alter insulin sensitivity or blood pressure in mice. When challenged with nutritional stress, however, partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension, providing further evidence for the importance of this gene in linking metabolic and cardiovascular disease.

Type 2 diabetes is associated with impaired endothelium-dependent, flow-mediated dilation, but impaired glucose metabolism is not: The Hoorn Study

Background: Type 2 diabetes (DM2) and impaired glucose metabolism (IGM) are associated with an increased cardiovascular disease risk. Impaired endothelial synthesis of nitric oxide (NO) is an important feature of atherothrombosis and can be estimated from endothelium-dependent flow-mediated dilation (FMD). It is controversial whether or not FMD is impaired in DM2 and IGM. We investigated this issue in a population-based setting. Methods and results: In the study population ( n=650; 246 with normal glucose metabolism (NGM), 135 with IGM and 269 with DM2; mean age: 67.6 years), FMD and endothelium-independent nitroglycerine-mediated dilation (NMD) were ultrasonically estimated from the brachial artery and expressed as the absolute change in diameter in mm. The increase in diameter (mean ± standard deviation) in NGM, IGM and DM2 was 0.19±0.15, 0.19±0.18 and 0.13±0.17 for FMD and 0.45±0.21, 0.43±0.24 and 0.45±0.25 for NMD. After adjustment for age, sex, baseline diameter and percentage increase in peak systolic velocity, DM2, as compared to NGM, remained associated with impaired FMD (regression coefficient β (95%CI)) as compared to NGM, −0.06 mm (−0.09 to −0.03). IGM was not associated with impaired FMD ( β, 0.01 mm (−0.02 to 0.04)). Additional adjustment for conventional cardiovascular risk factors did not alter these associations. Hyperglycemia or hyperinsulinemia explained 2% of the association between DM2 and FMD. NMD was not associated with glucose tolerance. Conclusions: This study shows that DM2 is independently associated with impaired FMD. Hyperglycemia and hyperinsulinemia contribute minimally to this association. Impaired FMD may therefore, in part, explain the increased cardiovascular disease risk in DM2, whereas the normal FMD in IGM suggests that other forms of endothelial dysfunction are important in explaining the increased cardiovascular disease risk in IGM.

The therapeutic effect of natriuretic peptides in heart failure; differential regulation of endothelial and inducible nitric oxide synthases.

The abnormal regulation of nitric oxide synthase activity represents an underlying feature of heart failure. Increased peripheral vascular resistance, and decreased renal function may be in part related to impaired endothelium-dependent nitric oxide (NO) synthesis. Paradoxically, the chronic production of NO by inducible nitric oxide synthase (iNOS) in heart failure exerts deleterious effects on ventricular contractility, and circulatory function. Consequently, pharmacologically improving endothelium-dependent NO synthesis and the concomitant inhibition of iNOS activity would be therapeutically advantageous. Interestingly, natriuretic peptides have been shown to differentially regulate endothelial NOS (eNOS) and iNOS activity. Moreover, in both patients and animal models of heart failure, pharmacologically increasing plasma natriuretic peptide levels ameliorated vascular tone, renal function, and ventricular contractility. Based on these observations, the following review will explore whether the therapeutic benefit of the natriuretic peptide system in heart failure may occur in part via the amelioration of endothelium-dependent NO synthesis, and the concomitant inhibition of cytokine-mediated iNOS expression.

Cardiorenal risk as a new frontier of nephrology: research needs and areas for intervention.

Understanding and modifying the causes of the high cardiovascular morbidity and mortality associated with renal disease is the greatest challenge faced by renal physicians. About one person in 20 has a serum creatinine level above normal (> or =1.5 mg/dl in males and > or =1.4 mg/dl in females), signifying mild kidney disease. People with hypertension, hyperlipidaemia, and/or diabetes (approximately 350000 people per million in the general population) have the highest risk of renal failure. Anaemia, extracellular volume expansion, increased angiotensin II and aldosterone levels, high calcium-phosphate product, inflammation, hyperhomocysteinaemia, and impaired nitric oxide synthesis all amplify the risk of cardiovascular disease in patients with renal failure. These factors may adversely affect the cardiovascular system by influencing the generation of reactive oxygen species, thus contributing to high oxidative stress. Further research into optimal follow-up of patients with mild renal insufficiency is needed. Identification of 'problematic' and/or treatment-resistant patients should be a primary goal. Greater understanding of the genetic and environmental precursors of diseases associated with renal insufficiency would also be beneficial, particularly for younger patients. Observational studies aimed at linking these risk factors to well-defined and measured renal and cardiovascular outcomes should increase knowledge of renal disease progression and cardiovascular risk in these populations.

Angiotensin II type 1 receptor blockade restores nitric oxide-dependent renal vascular responses in renovascular hypertension.

It was previously reported that enhancement of renal vascular responses to angiotensin II in hypertensive rats is related to decreased release of nitric oxide. Thus, it was suggested that impairment of nitric oxide synthesis during development of hypertension is related to a decreased nitric oxide synthase mRNA expression by an angiotensin II-dependent mechanism. The current study evaluated whether the blockade of angiotensin II type 1 receptor during the development of hypertension restored nitric oxide synthase mRNA expression, nitric oxide synthesis, and nitric oxide-dependent modulation of angiotensin II vasoconstrictor effects. It was shown that losartan treatment prevented increased vascular responses to angiotensin II in hypertensive rats and that this effect was associated with restoration of nitric oxide synthase mRNA expression and nitric oxide synthase activity. Furthermore, angiotensin II-dependent nitric oxide release in hypertensive rats was potentiated by losartan treatment. Angiotensin II (1 microg) released renal nitrites by 485 +/- 178, 470 +/- 150, 185 +/- 45, and 515 +/- 100 nmol/ml/30 s in the kidneys from normotensive, losartan-treated normotensive rats, hypertensive, and losartan-treated hypertensive rats, respectively. The data suggest that during development of hypertension, angiotensin II downregulates nitric oxide synthase mRNA expression, blunting nitric oxide vasodilatory tone and increasing vascular sensitivity to vasoconstrictor agents in the renal circulation.

Positive Association of the Endothelial Nitric Oxide Synthase Gene Polymorphisms With High-Altitude Pulmonary Edema

A defect of nitric oxide (NO) synthesis in the lung of high-altitude pulmonary edema (HAPE) has been suggested to contribute to its exaggerated pulmonary hypertension. Several polymorphisms have been identified in the gene encoding endothelial nitric oxide synthase (eNOS), which is a key enzyme responsible for NO synthesis, some of which were reported to be associated with vascular disorders. We studied 41 HAPE-susceptible subjects (HAPE-s) and 51 healthy climbers (control group) in a Japanese population. We examined 2 polymorphisms of the eNOS gene, including the Glu298Asp variant and 27-base pair (bp) variable numbers of tandem repeats using polymerase chain reaction followed by restriction fragment length polymorphism. The Asp allelic frequency of the Glu298Asp variant was 25.6% in the HAPE-s and 9.8% in the controls, which was significantly different between the two groups (P=0.0044). The eNOS4a allelic frequency of 27-bp variable numbers of tandem repeats was 23.2% in the HAPE-s, significantly higher than that of 6.9% in the controls (P=0.0016). In HAPE-s group, 11 of 41 (26.8%) subjects possessed simultaneously both of the two significant alleles, but among the controls, none did, which showed a high statistical difference between the two groups (P=0.000059). Both polymorphisms of the eNOS gene were significantly associated with HAPE. A genetic background may underlie the impaired NO synthesis in the pulmonary circulation of HAPE-s. These polymorphisms could be genetic markers for predicting the susceptibility to HAPE.

The Glu298Asp polymorphism in endothelial nitric oxide synthase gene is associated with coronary in-stent restenosis

Background: Reduced or impaired synthesis of nitric oxide promotes the proliferation of vascular smooth muscle cells, and thus may induce the neointimal formation leading to coronary in-stent restenosis. Recent reports have suggested that the Glu298Asp polymorphism in exon 7 of the endothelial nitric oxide synthase gene is associated with coronary spasm and acute myocardial infarction. In this study, we have examined the implication of this polymorphism with regard to coronary restenosis after Palmaz–Schatz stent deployment. Methods: Eighty-nine lesions in 85 consecutive patients were treated with Palmaz–Schatz stents, and were prospectively followed up for 6 months. The lesions were classified into a restenosis group (% diameter stenosis=50%) and a non-restenosis group. Assessment was made using an automated quantitative angiographic system. We performed polymerase chain reaction-restriction fragment length polymorphism analysis to detect the missense Glu298Asp variant in exon 7 of the endothelial nitric oxide synthase gene. Results: Coronary risk factors and angiographic findings of stenotic lesions did not differ between the groups. Univariate analyses showed that the missense Glu298Asp variant was the only statistically significant predictor of restenosis (odds ratio, 4.27; P=0.025). In addition, multiple logistic regression analysis revealed the missense Glu298Asp variant as the only independent predictor for in-stent restenosis (odds ratio, 3.90; P=0.036). Conclusions: The missense Glu298Asp variant may be an independent risk factor for in-stent restenosis.

Polymorphism in intron 23 of the endothelial nitric oxide synthase gene (NOS3) is not associated with hypertension.

Nitric oxide (NO) is synthesised in the vascular endothelium by nitric oxide synthase (NOS3) and is an important factor in the regulation of blood pressure. Impaired synthesis of NO due to mutations in the NOS3 gene is associated with hypertension. To date several allelic variants of the NOS3 gene have been identified and their possible linkage with hypertension investigated. We studied the distribution of genotypes and frequency of alleles of the G11T polymorphism in intron 23 of the NOS3 gene in patients with hypertension and in a control group of healthy individuals. The polymorphism was determined by PCR-RFLP analysis. The distribution of genotypes in the patients with hypertension and in the healthy individuals did not differ significantly from the values predicted from Hardy-Weinberg equilibrium for the general population. No major differences in the distribution of the G11T polymorphism in the patients and healthy individuals were found (P > 0.05).

Implications physiopathologiques et cliniques des r??cepteurs AT1/AT2 de l??angiotensine II dans les pathologies m??taboliques

The coexistence of hypercholesterolaemia and diabetes dramatically and synergistically increases the risk of microvascular and macrovascular complications in patients. A single unifying mechanism of increased production of reactive oxygen species (ROS) by angiotensin II (Ang II) may serve as a causal link between hyperglycaemia and hypercholesterolaemia and many of the major pathways responsible for atherogenic and diabetic disorders. Several lines of evidence suggest a crucial role for Ang II-mediated oxidative stress in the pathogenesis of hyperglycaemia- and hypercholesterolemia-associated endothelial dysfunction. Endothelial dysfunction in these scenarios may be due to impaired nitric oxide (NO) synthesis and/or inactivation of endothelium-derived NO by ROS. That Ang II plays an important role in the development of atherosclerosis and glomerulosclerosis is supported by numerous studies indicating that angiotensin receptor blockers (ARBs) retard the progression of these diseases in both experimental animal models and humans. Evidence indicates that Ang II contributes to atherogenesis at both transcriptional and translational levels by upregulating adhesion molecule mRNA and protein synthesis. The recent demonstration of Ang II AT(2) receptors in the adult kidney and their potential to oppose the vasoconstrictive, antinatriuretic, and profibrotic properties of AT(1) receptors suggests that the balance of intrarenal AT(1) and AT(2) receptors may be important in determining the cellular responses to Ang II in diabetic nephropathy. Results of these studies suggest that hypercholesterolaemia and hyperglycaemia can induce a pro-inflammatory response within coronary arteries and the kidney glomerulus. This response involves production of well described macrophage chemotactic and adhesion molecules, which results in macrophage recruitment and the development of acute and chronic injury. Glomerular macrophage recruitment in experimental diabetes occurs via Ang II-stimulated monocyte chemoattractant protein (MCP)-1 expression, suggesting that the renin-angiotensin system is an important regulator of local MCP-1 expression, and strongly implicating macrophage recruitment and activation in the pathogenesis of early diabetic glomerular injury. Diabetes-associated vascular complications may also involve an activation of the nuclear factor (NF)-kappaB by hyperglycaemia. NF-kappaB activation is related to AT(1) receptor-mediated pathways, and is believed to be dependent on activation of the Rho proteins belonging to the superfamily of low molecular weight guanosine triphosphatases (GTPases) that regulate intracellular signalling. Preincubation of vascular smooth muscle cells with insulin doubled NF-kappaB transactivation stimulated by Ang II and hyperglycaemia, suggesting a potential mechanism for crosstalk between the renin-angiotensin system and hyperglycaemia. Taken together, these data suggest that activation of the renin-angiotensin system is a mechanism for the initiation and progression of inflammatory cell infiltration found in early changes common to both hypercholesterolaemia and hyperglycaemia. While the base of information regarding ARBs in high-risk patients with diabetes and hypercholesterolemia is lacking, preclinical and pilot trial data suggest that the ARBs are reno- and vasculoprotective in these patients. Therapeutic blockade of Ang II AT(1) receptors in diabetic and hypercholesterolaemic humans by ARBs, with concomitant elevation in plasma and tissue Ang II levels, may provide vascular and renal protection not only by reducing AT(1) receptor-mediated pro-oxidative effects, but also by unopposed AT(2) receptor stimulation.

Anaemia in end-stage renal disease: pathophysiological considerations

Anaemia is a frequent complication of many diseases but the mechanisms that link reduced blood oxygen content to the long-term consequences of anaemia are incompletely understood. The maintenance of oxygen supply to the tissues during anaemia involves complex cardiovascular adaptations, including an increase in cardiac output, reduced peripheral resistance and increased oxygen extraction from haemoglobin (Hb). In addition, hypoxia-inducible factors are associated with the transcriptional activation of genes involved in adaptive mechanisms that increase oxygen delivery and provide alternative metabolic pathways. The complex pathophysiology of chronic kidney disease alters the adaptations to anaemia in uraemic patients. The increased cardiac output induced by anaemia is associated with left ventricular hypertrophy and cardiac disease in renal patients. Alterations in endothelial cell function, common in renal disease, may diminish endothelium-induced vasodilatation, increase the risk of atherosclerosis and impair angiogenesis. Many potential reasons for erythropoietin-induced hypertension in uraemic patients have been postulated, including increased blood viscosity as haematocrit rises, a reversal of hypoxic vasodilatation, increased blood volume that is not compensated by haemodialysis, ultrafiltration and impaired nitric oxide synthesis, preventing vascular relaxation in response to increased blood viscosity. In view of this impaired vascular reactivity, rapid increases in haematocrit should be avoided during epoetin treatment. As the interaction between anaemia and uraemia is very complex, it is not possible to derive the optimal Hb concentration for individual patients by using simple physiological or pathophysiological models and there is a need for good randomized controlled clinical trials to address this issue.

REDUCED BAROREFLEX SENSITIVITY AND BLUNTED ENDOGENOUS NITRIC OXIDE SYNTHESIS PRECEDE THE DEVELOPMENT OF HYPERTENSION IN TGR(mREN2)27 RATS

Transgenic TGR(mREN2)27 (TGR) rats are an animal model of fulminant hypertension characterized by an inverse circadian blood pressure profile. The present study addressed the contribution of nitric oxide (NO) synthesis and baroreflex function to hypertension and the inverse blood pressure pattern. NO synthesis was measured at four different times of day indirectly by excretion of NO metabolites (NOx: NO−2 and NO−3) in the urine of 5- and 11-week-old TGR and Sprague-Dawley (SPRD) controls. Blood pressure, heart rate, and motor activity were recorded in age-matched rats of both strains using an implantable telemetry system. Beat-to-beat recording of blood pressure and pulse interval was performed hourly in 6-week-old animals over 24h. From these data, baroreflex sensitivity (BRS) was calculated by linear regression of spontaneous fluctuations of blood pressure and corresponding changes of pulse interval. Baroreflex sensitivity was lower in prehypertensive TGR rats than in SPRD rats, and the reduction was restricted to the daily resting period. In both strains, NOx excretion showed circadian rhythmicity, with peak values during the activity period at night. Interestingly, excretion of NOx was reduced during the resting period in 5-week-old TGR rats prior to the development of hypertension. Impairment of NO synthesis and baroreflex function precede the development of hypertension in TGR rats. The reduction of both parameters was restricted to the resting period and, therefore, could be involved in the development of the inverse circadian blood pressure profile of TGR rats. (Chronobiology International, 18(2), 215–226, 2001)

Macrophage Preconditioning with Synthetic Malaria Pigment Reduces Cytokine Production via Heme Iron-Dependent Oxidative Stress

Hemozoin (malaria pigment), a polymer of hematin (ferri-protoporphyrin IX) derived from hemoglobin ingested by intraerythrocytic plasmodia, modulates cytokine production by phagocytes. Mouse peritoneal macrophages (PM) fed with synthetic β-hematin (BH), structurally identical to native hemozoin, no longer produce tumor necrosis factor α (TNFα) and nitric oxide (NO) in response to lipopolysaccharide (LPS). Impairment of NO synthesis is due to inhibition of inducible nitric oxide synthase (iNOS) production. BH-mediated inhibition of PM functions cannot be ascribed to iron release from BH because neither prevention by iron chelators nor down-regulation of iron-regulatory protein activity was detected. Inhibition appears to be related to pigment-induced oxidative stress because (a) thiol compounds partially restored PM functions, (b) heme oxygenase (HO-1) and catalase mRNA levels were up-regulated, and (c) free radicals production increased in BH-treated cells. The antioxidant defenses of the cells determine the response to BH: microglia cells, which show a lower extent of induction of HO-1 and catalase mRNAs and lower accumulation of oxygen radicals, are less sensitive to the inhibitory effect of BH on cytokine production. Results indicate that BH is resistant to degradation by HO-1 and that heme-iron mediated oxidative stress may contribute to malaria-induced immunosuppression. This study may help correlate the different clinical manifestations of malaria, ranging from uncomplicated to severe disease, with dysregulation of phagocyte functions and promote better therapeutic strategies to counteract the effects of hemozoin accumulation.

Role of Nitric Oxide in Modulating the Chronic Cardiovascular and Renal Actions of Leptin

64 Acute studies suggest that leptin has pressor and depressor actions, including stimulation of sympathetic activity as well as increased release of nitric oxide (NO)from the vascular endothelium. The goal of this study was to examine the role of NO in modulating the chronic blood pressure, heart rate, and renal responses to hyperleptinemia, comparable to that found in obesity hypertension. Male Sprague-Dawley rats were implanted with arterial and venous catheters and mean arterial pressure (MAP) and heart rate (HR) were monitored continuously 24h/d. After a 4 day control period, rats were treated with vehicle (n=6) or L-NAME (n=7) for 7 days. After 7 days of vehicle or L-NAME (10 μg/kg/min) treatment to inhibit NO synthesis, rats received a low rate (0.5 μg/kg/min) leptin infusion for 7 days followed by a moderate rate (1.0 μg/kg/min) leptin infusion for 7 days, along with vehicle or L-NAME. A 21 day infusion of L-NAME alone (n=6) served as a control for the leptin+L-NAME rats. The low and moderate leptin infusions reduced food intake by 53±2%and 51±7%, respectively, but L-NAME did not alter the anorectic effect of leptin. Although the low dose of leptin alone did not significantly elevate MAP, it raised HR by 18±3 bpm. The moderate dose of leptin raised MAP from 96±3 to 104±3, but did not increase HR further. In rats pre-treated with L-NAME, the moderate dose of leptin raised MAP by 40±5 mmHg and HR by 73±17 bpm compared to pre-treatment levels. In control L-NAME rats, MAP increased by 31±4 mmHg, while HR was not altered significantly, compared to pre-treatment levels. Chronic leptin infusion alone did not alter glomerular filtration rate (GFR) or renal plasma flow (RPF). However, Leptin+L-NAME reduced GFR by 47±8% and RPF 58±7%; L-NAME alone decreased GFR by 36±11% and RPF 36±17%. Thus, NO does not appear to modulate the anorectic effect of leptin. However, impaired NO synthesis enhances the chronic renal hemodynamic and hypertensive effects of leptin, and greatly amplifies the tachycardia caused by hyperleptinemia, at levels comparable to those found in obesity hypertension.

Role of Nitric Oxide in Modulating the Chronic Cardiovascular and Renal Actions of Leptin

64 Acute studies suggest that leptin has pressor and depressor actions, including stimulation of sympathetic activity as well as increased release of nitric oxide (NO)from the vascular endothelium. The goal of this study was to examine the role of NO in modulating the chronic blood pressure, heart rate, and renal responses to hyperleptinemia, comparable to that found in obesity hypertension. Male Sprague-Dawley rats were implanted with arterial and venous catheters and mean arterial pressure (MAP) and heart rate (HR) were monitored continuously 24h/d. After a 4 day control period, rats were treated with vehicle (n=6) or L-NAME (n=7) for 7 days. After 7 days of vehicle or L-NAME (10 μg/kg/min) treatment to inhibit NO synthesis, rats received a low rate (0.5 μg/kg/min) leptin infusion for 7 days followed by a moderate rate (1.0 μg/kg/min) leptin infusion for 7 days, along with vehicle or L-NAME. A 21 day infusion of L-NAME alone (n=6) served as a control for the leptin+L-NAME rats. The low and moderate leptin infusions reduced food intake by 53±2%and 51±7%, respectively, but L-NAME did not alter the anorectic effect of leptin. Although the low dose of leptin alone did not significantly elevate MAP, it raised HR by 18±3 bpm. The moderate dose of leptin raised MAP from 96±3 to 104±3, but did not increase HR further. In rats pre-treated with L-NAME, the moderate dose of leptin raised MAP by 40±5 mmHg and HR by 73±17 bpm compared to pre-treatment levels. In control L-NAME rats, MAP increased by 31±4 mmHg, while HR was not altered significantly, compared to pre-treatment levels. Chronic leptin infusion alone did not alter glomerular filtration rate (GFR) or renal plasma flow (RPF). However, Leptin+L-NAME reduced GFR by 47±8% and RPF 58±7%; L-NAME alone decreased GFR by 36±11% and RPF 36±17%. Thus, NO does not appear to modulate the anorectic effect of leptin. However, impaired NO synthesis enhances the chronic renal hemodynamic and hypertensive effects of leptin, and greatly amplifies the tachycardia caused by hyperleptinemia, at levels comparable to those found in obesity hypertension.

Posttransplantation hypertension related to calcineurin inhibitors.

Calcineurin inhibitors are a mainstay of transplant immunosuppression and commonly induce hypertension. They are highly lipid soluble and penetrate vascular smooth muscle cell membranes readily. Changes in vascular tone are universally observed during administration of these agents, particularly within the kidney, leading to diminished glomerular filtration and enhanced sodium retention. Disturbances of endothelial function are prevalent in many tissues, including stimulation of endothelin and impaired nitric oxide synthesis. Multiple additional pathways produce increased vasoconstriction, leading to an increase in arterial pressure. Clinical manifestations include disturbances in circadian blood pressure patterns, left ventricular hypertrophy, and acceleration of atherosclerotic and renal injury. Rapid increases in pressure occasionally produce accelerated hypertension and microangiopathic tissue damage. Principles of therapy require recognition of hazards of changing arterial pressures during calcineurin use and preferential use of vasodilating drugs, particularly dihydropyridine calcium channel blocking agents. Attention must be paid to interactions between antihypertensive agents and calcineurin inhibitor blood levels.

7-Nitroindazole potentiates the antiseizure activity of some anticonvulsants in DBA/2 mice

7-Nitroindazole, a selective neuronal nitric oxide synthase inhibitor (25–200 mg kg −1, intraperitoneally (i.p.)) antagonized audiogenic seizures in DBA/2 mice in a dose-dependent manner. We investigated the effects of 7-nitroindazole at a dose of 25 mg kg −1 i.p., which per se did not show anticonvulsant activity against audiogenic seizures in DBA/2 mice, on the antiseizure activity of some conventional antiepileptic drugs. 7-Nitroindazole sometimes potentiated the anticonvulsant activity of carbamazepine, diazepam, lamotrigine, phenytoin, phenobarbital and valproate against audiogenic seizures in DBA/2 mice. The degree of potentiation by 7-nitroindazole was greatest for phenobarbital and diazepam, less for valproate and least for carbamazepine, lamotrigine and phenytoin. The increase in anticonvulsant activity was associated with a comparable increase in motor impairment. However, the therapeutic index of combined treatment with diazepam+7-nitroindazole, phenobarbital+7-nitroindazole or valproate+7-nitroindazole was more favourable than that of the diazepam+vehicle, phenobarbital+vehicle or valproate+vehicle treatment. The results indicate that 7-nitroindazole is able to increase the protective activity of some conventional antiepileptics and this effect appears not to result only from the impaired synthesis of nitric oxide. In fact, mice receiving 7-nitroindazole (25 mg kg −1, i.p.) and l-arginine (30 μg/mouse, intracerebroventricularly (i.c.v.) did not show significant changes of ED 50 values in comparison to those of related groups of animals treated with 7-nitroindazole and anticonvulsants. 7-Nitroindazole was able to increase the brain levels of dopamine and noradrenaline and its anticonvulsant effects and changes in catecholamine content were antagonized by pretreatment with α-methyl-paratyrosine, an agent inhibiting the synthesis of catecholamines. The fact that α-methyl-paratyrosine reverses concomitantly both the increase in brain levels of dopamine and noradrenaline and the anticonvulsant properties of 7-nitroindazole strongly suggests an important role of catecholamines in the antiseizure activity of 7-nitroindazole. Since 7-nitroindazole did not significantly influence the total and free plasma levels of the anticonvulsant drugs studied, we suggest that pharmacokinetic interactions, in terms of total or free plasma levels, are not probable. 7-Nitroindazole did not significantly affect the hypothermic effects of the anticonvulsant compounds studied. 7-Nitroindazole showed an additive effect when administered in combination with some classical anticonvulsants, most notably diazepam, phenobarbital and valproate and its activity could be, in part, due to an increase of monoamine levels.

Die Bedeutung von Stickstoffmonoxid in der Pathophysiologie und Therapie der Präeklampsie*

The spectrum of the hypertensive disorders of pregnancy result from an imbalance between vasoconstrictive agglutinating substances and vasodilating anticoagulative substances. Nitric oxide is the most potent endogenous vasodilator and plays a central role in the regulation of vascular tone. Impaired synthesis of nitric oxide by endothelial cells is assumed to be a central mechanism in the pathophysiology of preeclampsia. Nitric oxide also decreases the sensitivity of thrombocytes to procoagulative substances, inhibits the adhesion of leucozytes. and reduces the release of endothelin and the proliferation of vascular smooth muscle cells. The role of the endothelial nitric oxide synthetase gene and the expression of endothelial nitric oxide synthetase in placental tissue and umbilical vessels in preeclampsia and normal pregnancy is unclear. Nitric oxide donors have been used experimentally in attempts to treat hypertensive disorders of pregnancy but their efficacy is unproved and there are no recommendations for clinical use.

The nitric oxide synthase cofactor tetrahydrobiopterin reduces allograft ischemia-reperfusion injury after lung transplantation

Objective: Exogenous nitric oxide reduces ischemia-reperfusion injury after solid organ transplantation. Tetrahydrobiopterin, an essential cofactor for nitric oxide synthases, may restore impaired endothelium-dependent nitric oxide synthesis. We evaluated whether tetrahydrobiopterin administration to the recipient attenuates lung reperfusion injury after transplantation in swine. Methods: Unilateral left lung transplantation was performed in 15 weight-matched pigs (24-31 kg). Donor lungs were flushed with 1.5 L cold (1°C) low-potassium-dextran solution and preserved for 20 hours. Group I animals served as controls. Group II and III animals were treated with a bolus of tetrahydrobiopterin (20 mg/kg). In addition, in group III a continuous infusion of tetrahydrobiopterin (10 mg/kg per hour over 5 hours) was given. One hour after reperfusion, the recipient right lung was occluded. Cyclic guanosine monophosphate levels were measured in the pulmonary venous and central venous blood. Extravascular lung water index, hemodynamic variables, lipid peroxidation, and neutrophil migration to the allograft were assessed. Results: In group III a significant reduction of extravascular lung water was noted in comparison with the controls ( P = .0047). Lipid peroxidation in lung allograft tissue was significantly reduced in group II ( P = .0021) and group III ( P = .0077) in comparison with group I. Pulmonary venous levels of cyclic guanosine monophosphate increased up to 23 ± 1 pmol/mL at 5 hours in group II and up to 40 ± 1 pmol/mL in group III (group I, 4.1 ± 0.5 pmol/mL [I vs III]; P < .001), whereas central venous levels of cyclic guanosine monophosphate were unchanged in all groups. Conclusion: Tetrahydrobiopterin administration during lung allograft reperfusion may reduce posttransplantation lung edema and oxygen-derived free radical injury in the graft. This effect is mediated by local enhancement of the nitric oxide/cyclic guanosine monophosphate pathway. (J Thorac Cardiovasc Surg 1999;118: 726-32)

Dietary Protein or Arginine Deficiency Impairs Constitutive and Inducible Nitric Oxide Synthesis by Young Rats

Effects of dietary protein or arginine deficiency on constitutive and lipopolysaccharide (LPS)-induced nitric oxide (NO) synthesis were determined in young rats by quantifying urinary nitrate excretion. In Experiment 1, 30-d-old rats (n = 16) were divided randomly into two groups (n = 8/group) and pair-fed on the basis of body weight semipurified isocaloric diets containing 20 or 5% casein. In Experiment 2, 30-d-old rats (n = 24) were divided randomly into three groups (n = 8) and pair-fed on the basis of body weight purified isonitrogenous and isocaloric diets (composed of amino acids) containing 0.0, 0.3 or 1.0% L-arginine. In both experiments, daily collection of urine was initiated 10 d after the start of pair-feeding. On d 17 after the pair-feeding was initiated, LPS (1 mg/kg body wt) was injected intraperitoneally into rats, and urine was collected daily for an additional 7 d. In Experiments 3 and 4, activities of constitutive and inducible NO synthases were measured in macrophages and various tissues from protein- or arginine-deficient rats (n = 6). Body weight was lower in rats fed the 5% casein diet or the 0.0 and 0.3% arginine diets than in those fed 20% casein or 1% arginine, respectively. Dietary protein or arginine deficiency decreased serum concentrations of arginine and urinary nitrate excretion before and after LPS treatment, indicating impaired constitutive and inducible NO synthesis. Protein malnutrition reduced constitutive and inducible NO synthase activities in brain, heart, jejunum, lung, skeletal muscle and spleen, and inducible NO synthase activity in macrophages. Because NO is a mediator of the immune response and is the endothelium-dependent relaxing factor, impaired NO synthesis may help explain immunodeficiency and cardiovascular dysfunction in protein- or arginine-deficient subjects.