Despite the increasing insight in the clinical importance of nitric oxide (NO), formerly known as endothelium-derived relaxing factor (EDRF), there is limited information about the metabolism and elimination of this mediator in humans. We studied the degradation of NO in healthy subjects inhaling 25 ppm for 60 minutes and in patients with severe heart failure inhaling 20, 40, and 80 ppm in consecutive 10-minute periods. In other healthy subjects, the renal clearance of NO metabolite was measured. The metabolism ex vivo was evaluated by direct incubation of nitrite, the NO oxidation product, in blood from healthy humans. During inhalation of NO, the plasma levels of nitrate increased progressively, both in the healthy subjects (from 26 to 38 jpmol/L, P<.001) and in the patients (from 72 to 90 !Lmol/L, P<.001).Methemoglobin (MetHb) also increased in the healthy subjects (from 7 to 13 ,mol/L, P<.001) as well as in the patients (from 19 to 42 ,umol/L, P<.01). No change in nitrosohemoglobin (HbNO) was detected, either in the healthy subjects or in the patients. In arterialized blood (02 saturation, 94% to 99%), incubated nitrite was semiquantitatively converted to nitrate and MetHb. In venous blood (02 saturation, 36% to 85%) moderate amounts of HbNO were also formed. Plasma and urinary clearance of nitrate in healthy subjects averaged 20 mL/min. We conclude that uptake into the red blood cells with subsequent conversion to nitrate and MetHb is a major metabolic pathway for endogenously formed NO. Nitrate may then enter the plasma to be eliminated via the kidneys. decreased formation of this compound,9'0 seems to constitute an important functional component in atherosclerotic vascular disease. An increased formation of NO in activated macrophages has been suggested to be responsible for the hypotension in endotoxin shock." Recently, the application of NO inhalation as a beneficial therapeutic principle was reported in patients with primary pulmonary hypertension12 or adult respiratory distress syndrome.13We assumed that quantitative methods to estimate NO formation might facilitate further evaluation of some of its physiological, pathophysiological, and therapeutic roles. The development of such methods relies on proper knowledge of the inactivation and elimination of NO from the intact organism. However, only little is known concerning the in vivo metabolism of NO and the excretion of its metabolite(s). The amino acid L-arginine, which is a precursor for NO both in macrophages and in endothelial cells,314 is also a precursor for nitrate biosynthesis in humans.15 Furthermore, nitrate is a normal constituent of human urine.16 Based on these observations, it might be speculated that NO is metabolized to nitrate and subsequently excreted as such into the urine. Preliminary support for this hypothesis was obtained earlier in a study on NO degradation in human blood ex vivo.'7 In the present report, the proposed metabolic route for NO is shown to be operative in vivo, in healthy subjects as well as in patients with severe...

SUMMARY1. Forearm blood flow was measured bilaterally in healthy young male and female volunteers, in the basal state and after upper-arm occlusion of arterial or venous blood flow for 1-20 min. The investigations were repeated after pre-treatment with drugs affecting vascular prostaglandins and/or adenosine.2. Simultaneous arterial occlusion in one arm and venous occlusion in the contralateral arm for up to 20 min elicited a considerable reactive hyperaemia in the arm subjected to arterial occlusion, but completely failed to elevate the post-occlusive flow in the arm subjected to venous occlusion above the pre-occlusive level.3. When the arterial occlusion was increased from 1 to 20 min there was a progressive increase in the subsequent reactive hyperaemia, up to 30 ml 100 ml tissue-1. The time dependence following 1-3 min of arterial occlusion was based on a facilitation of the peak post-occlusive flow, while prolongation of the arterial occlusion from 3 to 20 min augmented the reactive hyperaemia mainly by increasing its duration.4. Inhibition of prostaglandin synthesis with ibuprofen reduced the total reactive hyperaemia following 3-5 min of arterial occlusion by up to 70 %. This attenuation was due both to a reduction of peak post-occlusive flow and to a shortening of the duration of the post-occlusive hyperaemia.5. The adenosine receptor antagonist theophylline reduced the reactive hyperaemia following 5 min of arterial occlusion by about 35 %. Combined treatment with ibuprofen and theophylline did not reduce the reactive hyperaemia more than either drug alone.6. Infusion of dipyridamole, a drug which inhibits the elimination of adenosine, reinforced the reactive hyperaemia by about 45 %. This effect of dipyridamole was completely inhibited by administration of theophylline, and also by ibuprofen.7. Plasma levels of adenosine, hypoxanthine and uric acid were maintained during the reactive hyperaemia, indicating increased production of purines during or immediately after the ischaemia.

We analyzed nitrate, a major stable end product of nitric oxide (NO) metabolism in vivo in plasma and urine from groups of healthy subjects with different working capacities. Resting plasma nitrate was higher in athletic subjects than in nonathletic controls [45 +/- 2 vs. 34 +/- 2 (SE) microM; P < 0.01]. In other subjects, both the resting plasma nitrate level (r = 0.53; P < 0.01) and the urinary excretion of nitrate at rest (r = 0.46; P < 0.01) correlated to the subjects' peak work rates, as determined by bicycle ergometry. Two hours of physical exercise elevated plasma nitrate by 18 +/- 4 (P < 0.01) and 16 +/- 6% (P < 0.01), respectively, in athletes and nonathletes, compared with resting nitrate before exercise. We conclude that physical fitness and formation of NO at rest are positively linked to each other. Furthermore, a single session of exercise elicits an acute elevation of NO formation. The observed positive relation between physical exercise and NO formation may help to explain the beneficial effects of physical exercise on cardiovascular health.

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Nitric oxide (NO) is metabolized to nitrate in humans. Accordingly, plasma nitrate has been proposed as an index of the in vivo formation of NO. Such an application requires knowledge about the possible influence of nitrate from sources other than endogenous NO formation, as well as of the kinetics of nitrate in plasma. In the present study, plasma nitrate increased from 32 +/- 4 to 205 +/- 27 mumol/l (mean +/- SE) following intake of nitrate-rich food. It dropped during the intake of nitrate-restricted diet and stabilized at a level of 29 +/- 1 mumol/l. The urinary excretion of nitrate during nitrate restriction was 840 +/- 146 mumol/24 h. Plasma nitrate was not affected following the intake of a gastrointestinal antibiotic drug for a period of four days. Smoking three cigarettes in succession did not affect the plasma nitrate levels significantly. The oral intake of potassium nitrate (500 mg approximately 4950 mumol) elevated plasma nitrate from 29 +/- 3 to 313 +/- 12 mumol/l within 60 min. The subsequent drop in plasma nitrate, with a t1/2 of 451 +/- 42 min, was probably a reflection of the redistribution of nitrate within the body fluids and the renal excretion of nitrate. The plasma clearance of nitrate was 30 +/- 2 ml/min/1.73 m2 BSA. The distribution volume for nitrate was 28 +/- 1% of the bodyweight (BW). We conclude that plasma nitrate can be used as an index of the endogenous formation of NO, provided that the oral intake of nitrate is restricted for at least 48 h. Due to the large distribution volume and the low clearance of the ion wide-spread, marked, and chronic changes in NO formation are required to significantly affect the levels of nitrate in samples of mixed blood.

SummaryEndothelial cells regulate vascular tone by secreting paracrine mediators that control the contractility of arterial smooth muscle cells. Nitric oxide (NO) is an important vasodilating agent that is generated from t-arginine by the enzyme nitric oxide synthase (NOS), which is expressed constitutively by the endothelium. NO also inhibits platelet aggregation, contributing to the antithrombotic properties of the endothelial surface. It would therefore be expected that loss of the endothelium during arterial injury would lead to vasospasm and thrombosis but instead, the neointima formed after injury has a nonthrombogenic surface and a maintained vascular patency. We report here that arterial smooth muscle cells in the neointima formed after a deendothelializing balloon injury to the rat carotid artery express the cytokine-inducible isoform of NOS. Expression was detectable by reverse transcription-polymerase chain reaction from day 1-14 after injury and in situ hybridization showed expression of NOS mRNA by neointimal smooth muscle cells, particularly at the surface of the lesion. This was associated with systemically detectable NO production as revealed by electron paramagnetic resonance spectroscopic analysis of nitrosylated red cell hemoglobin. Local NO production by intimal smooth muscle cells after endothelial injury could represent an important mechanism for the maintenance of arterial patency and nonthrombogenicity in the injured artery.

The results suggest that the present 24(S)-hydroxylase mediated mechanism is most important for elimination of cholesterol from the brain of rats. There is a slow conversion of brain cholesterol into 24(S)-hydroxycholesterol with a rapid turnover of the small pool of the latter oxysterol due to leakage to the circulation (halflife of brain 24(S)-hydroxycholesterol is about 0.5 days as compared with 2-4 months for brain cholesterol). It is evident that the 24(S)-hydroxylation greatly facilitates transfer of cholesterol over the blood-brain barrier and that this hydroxylation may be critical for cholesterol homeostasis in the brain.

Background. Cigarette smoking is a risk factor for cardiovascular disease. The present study addressed the effect of tobacco use on the formation of two eicosanoids, thromboxane A2 and prostacyclin, which have been implicated in both acute and chronic cardiovascular disorders.Methods and Results. In 577 randomly sampled 18-19-year-old men, the urinary excretion of the 2,3 -dinor metabolites of thromboxane A2 and prostacyclin (Tx-M and PGI-M, respectively)