Diurnal variation of flow-mediated vasodilation in healthy premenopausal women.

The present study was designed to test the hypothesis of a diurnal variation of endothelial function. Sixteen healthy, nonsmoking women were studied, each on four occasions during one 24-h period (2:00 PM, 8:00 PM, 2:00 AM, and 8:00 AM). Endothelial function was assessed by ultrasound determinations of flow-mediated vasodilation (FMD%) in the brachial artery. FMD% was contrasted with endothelium-independent vasodilation, i.e., nitroglycerine-induced vasodilation (NTG%). Additionally, plasma concentrations and urinary excretion of nitrate and cGMP were analyzed. FMD% and NTG% displayed diurnal, albeit not parallel, patterns of variation. Whereas FMD% gradually increased from 2:00 PM and peaked at 2:00 AM (means +/- SE: 3.1 +/- 0.4, 4.4 +/- 0.4, 5.1 +/- 0.9, and 3.9 +/- 0.8%), the NTG% demonstrated a nadir at 2:00 AM. Plasma levels and urinary excretion of nitrate and cGMP did not display diurnal variation and no clear association with the variations seen in FMD% and NTG%. This study demonstrates a diurnal variation in both endothelium-dependent and -independent vasodilation in the brachial artery of healthy women. The background and possible implication of such a variation require further studies.

Intrathecal release of nitric oxide in Alzheimer's disease and vascular dementia.

A growing body of evidence points out the potential role of inflammatory mechanisms in the pathophysiology of brain damage in dementia. We have recently demonstrated that patients with Alzheimer's disease (AD) and vascular dementia (VaD) display an intrathecal production of proinflammatory cytokines. TNF-alpha, one of these cytokines, leads to the production of nitric oxide (NO), a potent inflammatory mediator, by induction of inducible NO synthase. The aim of the present study was to investigate the intrathecal levels of nitrate, one of the main metabolites of NO, and to relate its levels to the degree of intellectual impairment, in patients with AD and VaD. Twenty patients with early AD and 26 patients with VaD were analyzed with respect to cerebrospinal fluid levels of nitrate by gas chromatography/mass spectrometry. Interestingly, in patients with AD but not VaD, the intrathecal levels of nitrate were significantly and inversely correlated (r = -0.68, p = 0.002) to the degree of intellectual impairment. Our study demonstrates an inverse correlation between the intrathecal levels of nitrate and the degree of cognitive impairment in patients with AD, suggesting a neuroprotective effect of NO in AD.

Intrathecal release of nitric oxide and its relation to final brain damage in patients with stroke.

The potential role of inflammatory mechanisms in the pathophysiology of ischemic brain damage has intensely been discussed. We have recently demonstrated that stroke patients display an intrathecal production of proinflammatory cytokines early after onset of symptoms. IL-1beta, one of these cytokines, stimulates the production of nitric oxide (NO), a potent inflammatory mediator. The aim of the present study was to investigate the intrathecal levels of nitrate, one of the main metabolites of NO in acute stroke and to relate its levels to brain damage. Stroke patients were prospectively studied with clinical evaluation, radiological assessment and analysis of intrathecal levels of nitrate by gas chromatography/mass spectrometry. In addition, simultaneous analyses of cytokines and of soluble Fas/APO-1 and bcl-2, two proteins regulating apoptosis, were performed. The intrathecal levels of nitrate were not significantly different in stroke patients compared to controls throughout the observation period. However, the intrathecal levels of nitrate increased significantly 3 months after stroke onset compared with the first 3 days. Interestingly, the levels of nitrate measured at stroke onset were negatively correlated to the final size of infarct volume (r = -0.69, p < 0.05) measured by MRI. In addition, patients with large infarcts displayed significantly (p = 0.008) lower levels of nitrate in cerebrospinal fluid compared to patients with small infarcts during the first 3 days after stroke onset. In contrast, the intrathecal levels of nitrate were significantly positively correlated (r = 0.79, p < 0. 001) to the neurological deficit and negatively correlated (r = -0. 76, p < 0.05) to bcl-2, a protein downregulating neuronal apoptosis, in the late stage of the stroke. Early NO production is associated with a smaller infarct volume, suggesting a protective effect, whereas late NO production is associated with severer neurological deficits, suggesting a neurotoxic effect. Treatment trials pertaining to modulate NO production in stroke should take into consideration the dual effect of NO on ischemic brain damage.

An 18oxygen inhalation method for determination of total body formation of nitric oxide in humans.

The formation of nitric oxide (NO) and the subsequent conversion of the NO formed into nitrate require molecular oxygen. Based on this fact, we have recently developed a method using inhalation of the stable oxygen isotope, i.e. 18O2, to determine total formation of NO in small laboratory animals. The method has now been further developed to be applicable also in humans. Five healthy awake male subjects inhaled a gas mixture of unlabelled and 18-labelled oxygen (approximate ratio 4:1) in nitrogen from a closed breathing system equipped with eliminators for carbon dioxide and water vapour. The ratio of unlabelled to 18-labelled oxygen, as well as the total oxygen concentration during the inhalation, were monitored. Venous blood samples were taken before and after the inhalation for analysis of unlabelled and 18O-labelled nitrate by gas chromatography/mass spectrometry. The procedure was repeated with the same protocol on a later occasion, during ongoing treatment with the NO synthesis inhibitor NG-monomethyl-L-arginine (L-NMMA). The average nitrate level in plasma in the absence of L-NMMA was 26 micromol l-1. The rate of total synthesis of NO was estimated to be 0.38 +/- 0.06 mu mol kg-1 h-1, corresponding to a total body formation of 600-700 mu mol/24 h in an adult male. Infusion of L-NMMA caused an increase in mean arterial blood pressure from 86 +/- 4 to 99 +/- 5 mmHg (P<0.05). The average plasma level of nitrate during infusion of L-NMMA was 24 mu mol l-1. NO formation during infusion of L-NMMA was 0.17 +/- 0.03 mu mol kg-1 h-1, i.e. significantly (P<0.05) lower than in the absence of L-NMMA. We suggest that the described method allows direct determination of total NO formation in man. The method may be useful in the study of various experimental and pathophysiological conditions affecting NO formation.

Detection of an approximately 100 kD protein with strong immunoreactivity to antibodies specific for inducible nitric oxide synthase (iNOS) but without NOS activity in neutrophils of patients suffering from sepsis: results of a preliminary study.

OBJECTIVE: The study was designed to evaluate the expression of inducible nitric oxide synthase (iNOS) in activated human neutrophils. SUBJECTS AND METHODS: By Western blotting and immunocytochemical staining, iNOS expression was analyzed in neutrophils from patients with sepsis who we classified by high plasma nitrate as subjects with activated NO metabolism. For comparison, rats treated with Zymosan documented to induce iNOS were analyzed. RESULTS: Strong immunoreactivity to antibodies for iNOS was detected in leukocytes of Zymosan treated rats and in neutrophils of septic patients. Such immunoreactivity was absent in untreated rats and healthy subjects. In rats, the immunoreactivity was associated to the 130 kD protein as expected for iNOS. In contrast, the intense iNOS staining in neutrophils of septic patients was associated to an approximately 100 kD protein. Despite the iNOS staining, no increased NOS activity could be demonstrated in the neutrophils of septic patients. CONCLUSIONS: The detection of an approximately 100 kD protein with immunoreactivity to antibodies recognizing human iNOS but without NOS activity in neutrophils of patients with sepsis should initiate studies to elucidate the origin and function of this protein.

Urinary albumin excretion and heart rate variability in obese women.

OBJECTIVE: The aim of this work was to examine the relationship between cardiac autonomic function and urinary albumin excretion in obesity. SUBJECTS: These were 27 obese non-diabetic postmenopausal women and 18 non-obese healthy postmenopausal women. MEASUREMENTS: Urinary albumin excretion as well as plasma nitrate, both indices of capillary function, were measured. Power spectral analysis of heart rate variability was performed, as a measurement of vagal function. An oral glucose tolerance test (OGTT) was performed and blood lipids were analysed. RESULTS: The obese women were characterized by higher fasting insulin, sum of glucose, triglycerides and lower high density lipoprotein cholesterol (HDL), the latter of borderline significance, than controls. Urinary albumin excretion (UAE), plasma nitrate and heart rate variability were not different between obese and control women. However, in obese women log UAE correlated positively with systolic and diastolic blood pressure, and inversely with heart rate variability, the latter independent of body mass index (BMI) and the waist/hip circumference ratio. CONCLUSION: It was concluded that this inverse association between UAE and parasympathetic activity in obese women may be an early sign of derangements of endothelial function and autonomic nervous system control, which may contribute to the increased risk of cardiovascular mortality in abdominal obesity.

Effect of nitric oxide gas on platelets during open heart operations.

BACKGROUND: The increased bleeding tendency observed after cardiopulmonary bypass is caused in part by thrombocytopenia and impaired platelet function induced by the procedure. Previous in vitro studies have shown that nitric oxide (NO) added to the oxygenator sweep gas reduces platelet activation during experimental perfusion. We evaluated the effect of 40 ppm of NO, added to the oxygenator sweep gas, on platelet consumption and activation in patients undergoing cardiopulmonary bypass. METHODS: Twenty patients scheduled to undergo cardiopulmonary bypass were randomized to either the control or the NO arm of the study. Their platelet count, plasma beta-thromboglobulin level, platelet membrane glycoprotein Ib and IIb/IIIa levels, adenosine diphosphate-induced platelet aggregation, plasma nitrate level, and plasma hemoglobin were assayed before, during, and after cardiopulmonary bypass. RESULTS: After operation, slightly higher platelet counts were observed in the NO-treated patients than in the control patients, which might indicate a lower degree of platelet adhesion to the artificial surfaces of the extracorporeal circuit. However, this difference did not reach statistical significance. In addition, a difference in platelet membrane expression of glycoprotein Ib was seen between the NO and control groups after operation; the platelets of the control patients had significantly higher glycoprotein Ib expression than those of the NO-treated patients. The results of platelet aggregometry indicated preserved platelet function in both the NO-treated and control patients. The blood methemoglobin levels also were low in both groups. CONCLUSIONS: Nitric oxide might reduce the platelet consumption encountered during cardiopulmonary bypass without having any adverse effect on platelet function, as reflected by the preserved aggregation response seen in our patients. However, the best route of NO administration and the optimum dose remain to be established.

Estimation of total rate of formation of nitric oxide in the rat.

Nitric oxide (NO) is a powerful mediator with important actions in several organ systems. NO is synthesized during the enzymatic conversion of l-arginine and molecular oxygen to L-citrulline. About 90% of the NO formed is degraded to nitrate. Utilizing this information we have developed a method for assessment of the total rate of formation of NO in the rat. Male Wistar rats were kept in a closed-cage system allowing controlled breathing of a mixture of 18O2 and 16O2 in N2 for up to 5h. Blood samples for mass spectrometric analysis of nitrate residues with varying numbers of 18O atoms incorporated were drawn before and during the exposure to 18O2. By comparing the relative incorporation of 18O into nitrate residues to the 16O2/18O2 ratio in the breathing gas mixture in the cage system it was possible to calculate the absolute rate of NO formation in the animal. The rate of formation of NO in anaesthetized rats ranged from 0.33 to 0.85 micromol.kg-1.h-1. The rate of formation did not differ significantly in rats which were awake during the experiment (range 0.36-0.72 micromol.kg-1.h-1). The L-arginine analogue Nomega-nitro-L-arginine methyl ester (L-NAME) dose-dependently inhibited the formation of NO, at a dose of 100mg/kg by more than 99%. The technique presented allows estimation of the total rate of formation of NO in vivo in rats. Application of the technique may yield important information about the physiological and pathophysiological roles of NO. It may also be utilized to evaluate the effect of pharmacological treatment on NO formation.

Increase in nitric oxide formation after chronic voluntary exercise in spontaneously hypertensive rats.

The effect of chronic voluntary exercise on the plasma level of nitrate, a major stable metabolite of nitric oxide (NO) was studied in spontaneously hypertensive rats (SHR). Exercise consisted of spontaneous running in wheels for 3-35 days. Blood samples were collected after 3, 7, 14, 21 and 35 days of exercise and all samples were drawn after the running wheel had been locked during the preceding 12 h. The plasma nitrate level was significantly (P < 0.05) elevated in SHR after 35 days of exercise. Surprisingly after 7 days of exercise a significant (P < 0.001) decrease in the nitrate level in plasma was noted. Further research is needed to elucidate this biphasic change in nitrate seen in this study. The elevated level of plasma nitrate seen after 35 days of voluntary exercise was still present up to 36 h after termination of exercise. We conclude that exercise training in SHR elicits an enhanced formation of NO.

The arginine-nitric oxide pathway in thrombotic microangiopathy.

Eleven patients with thrombotic thrombocytopenic purpura (TTP) or haemolytic uremic syndrome (HUS) were investigated with respect to plasma concentrations of L-arginine, a substrate for nitric oxide (NO) and asymmetrical dimethyl arginine (ADMA), during active disease and after recovery. Plasma concentration of NO3-, the degradation product of NO, was also analyzed. The patients were treated with fresh-frozen plasma and plasmapheresis. One of the patients had experienced relapses of TTP five times during the preceding year. After treatment with p.o. arginine hydrochloride 1.5 g x 3 was started, no relapse has occurred during a 12-month period. During the active phase the plasma concentration of arginine was low and that of NO3- was very high, indicating a high NO-synthesis rate. The arginine concentration normalized on recovery. Plasma levels of ADMA, was twice normal during active disease, and did not return to normal on recovery. In conclusion, patients with TTP/HUS exhibit signs of activation of the NO-synthesis.

Importance of a novel oxidative mechanism for elimination of brain cholesterol. Turnover of cholesterol and 24(S)-hydroxycholesterol in rat brain as measured with 18O2 techniques in vivo and in vitro.

The brain is the most cholesterol-rich organ in the body. Brain cholesterol is characterized by a very low turnover with very little exchange with lipoproteins in the circulation. Very recently we showed that there is a continuous age-dependent flux of 24(S)-hydroxycholesterol from the human brain into the circulation (Lütjohann, D., Breuer, O., Ahlborg, G., Nennesmo, I., Sidén, A., Diczfalusy, U., and Björkhem, I. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 9799-9804). Here we measured the rate of synthesis of cholesterol as well as the conversion of cholesterol into 24(S)-hydroxycholesterol in rat brain in vivo with use of an 18O2 inhalation technique and mass isotopomer distribution analysis. Cholesterol synthesis was found to correspond to 0.03 +/- 0.01% of the pool per h. Conversion of cholesterol into 24(S)-hydroxycholesterol was of a similar magnitude, about 0.02% of the pool per h. Brain microsomes converted endogenous cholesterol into 24(S)-hydroxycholesterol at a similar rate when incubated in the presence of NADPH. When incubated with whole homogenate and subcellular fractions of rat brain, there was no significant conversion of tritium-labeled 24-hydroxycholesterol into more polar products. Plasma from 18O2-exposed rats contained 24(S)-hydroxycholesterol with an enrichment of 18O similar to that in 24(S)-hydroxycholesterol in the brain. 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 (half-life 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.

Seasonal variations in cyclic GMP response on whole-body cooling in women with primary Raynaud's phenomenon.

1. Primary Raynaud's phenomenon (PRP) is characterized by increased vasoconstrictor tone that develops during exposure to cold. The symptoms are most pronounced during the winter months with low outdoor temperature. The L-arginine-nitric oxide (NO)-cyclic GMP (cGMP) pathway plays an important role in counteracting vasospasm. The aim of the present study was to investigate if the venous cGMP response to whole-body cooling in women with PRP varied with the season of the year. 2. The study was performed as an open parallel-group comparison between women with PRP and healthy female controls during the winter months of February 1994 and 1995 and in the summer month of August 1994. Blood samples were drawn just before and 40 min after whole-body cooling. 3. There were no significant changes in venous cGMP after whole-body cooling in women with PRP during the winter months of February 1994 and 1995. Cold exposure in the summer month of August resulted, however, in a significant increase in venous cGMP (P < 0.01). In contrast, the healthy women responded with a significant increase in venous cGMP on all three test occasions: February 1994 (P < 0.05), August 1994 (P < 0.05) and February 1995 (P < 0.01), 4. A seasonal variation in venous cGMP response to whole-body cooling was observed only in women with PRP. Healthy women responded to cold exposure with an increase in venous cGMP during summer and winter, whereas females with PRP showed an increase only during summer. Results from the present study might indicate seasonal variation in the regulation of constitutive nitric oxide synthetase in women with PRP, which may contribute to new therapeutic approaches.

Transformation of subcutaneous nitric oxide into nitrate in the rat.

Following its addition to arterialized blood in vitro, nitric oxide (NO) is transformed into nitrate in the erythrocytes. Inhaled NO is similarly transformed into nitrate in the blood in vivo. These observations suggest that nitrate is a universal end-metabolite of NO, i.e. of endogenously formed NO as well. However, endogenous NO may also be inactivated in tissues, i.e. outside the vascular lumen. To study the fate of NO metabolized with delayed access to the blood, rats were given subcutaneous injections of 15NO or K15NO3, and the plasma concentrations of 15NO3(-) were followed for 450 min after injection. The values for the distribution volume and plasma decay (t12) of 15NO3(-) did not differ between rats given 15N-labelled NO and NO3(-). The area under the plasma decay curve for rats given 15NO amounted to 89% of the corresponding area for animals given K15NO3. This demonstrates that 15NO, when given extravascularly in millimolar concentrations, is mainly transformed into 15N-labelled nitrate. Other rats were kept in an atmosphere containing a mixture of 16O2 and 18O2. Nitrate residues containing either one or two 18O atoms were isolated from the blood, indicating that inhaled oxygen was incorporated during both the formation of NO and the subsequent transformation of NO into nitrate. The fraction of nitrate residues containing two 18O atoms was larger than that containing one 18O atom. We propose that nitrate is a major stable metabolite of endogenous NO that does not primarily diffuse into the vascular lumen following formation. Hence nitrate seems to be the quantitatively most important end-product of the metabolism of endogenous NO. The transformation of endogenous NO into nitrate involves the incorporation of inhaled oxygen.

Plasma nitric oxide metabolite in women with primary Raynaud's phenomenon and in healthy subjects.

Primary Raynaud's phenomenon (PRP) is characterized by cold- or stress-induced transient attacks of impaired skin circulation in fingers and/or toes. PRP displays seasonal variation with less severe symptoms in the summer. The aetiology has not been clarified. The aims of the present study were (a) to assess the influence of cold exposure on the plasma levels of the nitric oxide (NO) metabolite, nitrate, in patients with PRP and in healthy control subjects; and (b) to investigate whether there is a seasonal variation in these plasma levels. In a group of women with PRP and matched control subjects, venous blood was sampled before and at the end of a 40-min period of whole-body cooling. The study was performed with the same protocol on two occasions; once in the winter and once in the summer. A seasonal variation was detected with higher plasma levels of nitrate in the winter than in the summer, both in PRP and in control subjects. However, the plasma level of nitrate was not changed in response to cold exposure on any occasion, either in the patient or in the control group. Our study indicates that NO formation is up-regulated in response to cold weather in both study groups. However, NO formation does not seem to be increased in response to whole-body cooling, either in PRP patients or in healthy subjects. Further investigations are required to reveal whether the observed seasonal variation in NO formation is a universal phenomenon in man.

Both physical fitness and acute exercise regulate nitric oxide formation in healthy humans.

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.

Plasma nitrate as an index of nitric oxide formation in man: analyses of kinetics and confounding factors.

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.

Thromboxane metabolite excretion in patients with hand-arm vibration syndrome.

As chronic exposure to hand-held vibrating tools may cause endothelial injury, a subsequent sustained platelet activation with the increased release of vasoconstricting thromboxane A2 (TxA2) could be of pathophysiological importance in vibration-induced Raynaud's phenomenon. Therefore, the aim of this study was to elucidate whether or not hand-arm vibration syndrome is accompanied by increased endogenous TxA2 biosynthesis. The study involved 64 men, aged 23-61 years, stratified according to the exposure to vibrating tools, the presence of Raynaud's phenomenon, and smoking habit. Forty of them were car mechanics and 24 were age-matched healthy volunteers who served as controls. The assessment of platelet TxA2 formation in vivo was performed by quantification of the urinary excretion of its major metabolite, 2,3-dinorthromboxane B2 (2,3-dinor-TxB2), employing gas chromatography-mass spectrometry. The average urinary excretion rate of 2,3-dinor-TxB2 in patients with Raynaud's phenomenon was 296 +/- 42 pg/mg creatinine and did not differ significantly from the corresponding values in controls (328 +/- 62 pg/mg creatinine) or individuals exposed to vibrating tools, but without any signs of vasospastic disease (232 +/- 29 pg/mg creatinine). The only statistically significant difference was found between smokers and non-smokers (P < 0.001), a finding confirming the existence of chronic platelet dysfunction in cigarette smokers. The present data indicate that chronic exposure to vibrating tools, with or without Raynaud's phenomenon, is not associated with an enhanced platelet function as monitored by the urinary excretion of 2,3-dinor-TxB2. Hence, a possible vibration-induced vascular injury does not seem to provide a stimulus sufficient to induce a persistent platelet activation.

Magnesium inhibits platelet activity--an infusion study in healthy volunteers.

Magnesium (Mg) has shown the ability to inhibit arterial thrombus formation in some experimental animal studies. This effect may be due to an inhibition of platelet reactivity as in vitro studies have demonstrated that Mg inhibits platelet aggregation. In order to evaluate the in vivo effect of Mg in humans measurements of platelet activity, fibrinolytic activity, as well as measurements of prostacyclin (PGI2), and nitric oxide (NO) release were performed after infusion of magnesium sulphate (MgSO4) in healthy volunteers. In a placebo controlled, cross-over study in 14 healthy male subjects, 8 mmol MgSO4 was given as an intravenous bolus over 15 min followed by 3 mmol MgSO4/h. The mean S-Mg concentration increased from 0.85 to 1.50 mM during the Mg infusion period. A transient decrease in blood pressure was observed during the initial bolus infusion of Mg. Haemodynamic parameters were otherwise unstable. The bleeding time increased by 48% during the Mg infusion (p < 0.005), and in accordance with this, ex vivo platelet aggregation in platelet rich plasma was significantly inhibited, both following collagen (p = 0.02) and ADP (p = 0.04) stimulation. There were no significant changes in plasma beta-thromboglobulin concentration or the excretion of 2,3-dinor-thromboxane B2 in the urine. Neither tissue plasminogen activator (t-PA)activity, tissue plasminogen activator (t-PA)antigen nor plasminogen activator inhibitor (PAI)antigen changed during the Mg infusion period. There was no sign of increased release of PGI2 from the vessel wall as judged by urinary concentration of 2,3-dinor-6-keto-prostaglandin F1 alpha. Nor was there any sustained increase in the release of NO, measured as nitrate concentration in urine. However, a transient increase in NO release was observed during one sample period. In conclusion a reduced platelet activity and increased bleeding time, was found during Mg infusion in healthy volunteers. Fibrinolytic activity showed no changes. An anti-platelet effect may in part be responsible for the beneficial effect of Mg, described in patients with acute myocardial infarction (MI) and preeclampsia.

Smoke-derived nitric oxide and vascular prostacyclin are unable to counteract the platelet effect of increased thromboxane formation in healthy female smokers.

The incidence of cigarette smoking tends to be higher in women, justifying directed studies on smoke-related mechanisms of cardiovascular disorder in females. Platelet activity plays an important etiological role in several settings of cardiovascular disease. Cigarette smoking facilitates platelet formation of proaggregatory thromboxane A2. However, cigarette smoke contains nitric oxide (NO), which has antiplatelet activity. Furthermore, the formation of anti-aggregatory prostacyclin (PGI2) may be higher in smokers than in non-smokers. Hence, the concerted action of NO and PGI2 on platelet activity in smoking females is important to elucidate. The metabolites of TxA2, NO, and PGI2, as well as cyclic guanosine 3':5'-monophosphate (cGMP; second messenger for NO in the platelets) and cyclic adenosine 3':5'-monophosphate (cAMP; second messenger for PGI2 in the platelets), were analysed in 23 healthy female smokers (daily consumption 11-20 cigarettes per day) and in 26 matched non-smokers. The urinary excretion of 2,3-dinor TxB2 (metabolite of TxA2) was considerably higher in smokers than in non-smokers (177 vs. 72 pg/mg creatinine, respectively; P<0.001). Plasma and urinary levels of nitrate (metabolite of inhaled NO) did not differ between the groups. Plasma and urinary cGMP were slightly increased (252 vs. 193 nmol/L; P<0.05 and 0.63 vs. 0.51 micromol/24 h; P<0.05, respectively) in smokers compared to non-smokers, while platelet cGMP was lower in smokers than in non-smokers (81 vs. 10.3 pmol/10(6) platelets, respectively; P<0.05). The urinary excretion of 2,3-dinor-6-keto-PGF1a (metabolite of PGI2) did not differ between the groups. Platelet or urinary cAMP did not differ between the groups either, while plasma cAMP was lower in smokers than in non-smokers (19.2 vs. 26.2 nmol/l, respectively; P<0.001). In healthy female smokers NO is not absorbed from the inhaled smoke, and endothelial PGI2 formation is not enhanced to counterbalance the increased platelet formation of proaggregatory TxA2.