Milk production of Holstein heifers fed either alfalfa or corn silage diets at two rates of daily gain

OBJECTIVES: L-arginine exerts anti-atherosclerotic effects in hypercholesterolaemic rabbits via modulating endogenous NO production. We investigated whether L-arginine inhibits thromboxane formation in vivo and platelet aggregation ex vivo in this animal model. METHODS: The urinary excretion rates of 2,3-dinor-6-keto-PGF1 alpha (major urinary metabolite of PGI2) and 2,3-dinor-TXB2 (major urinary metabolite of thromboxane A2) were used as indicators of platelet-endothelial cell interactions in vivo. Rabbits were fed 1% cholesterol (Cholesterol group, N = 8), 1% cholesterol plus 2.25% L-arginine (Cholesterol + L-arginine, N = 8), or normal rabbit chow (Control, N = 4) for 12 weeks. Urine samples were collected in weekly intervals. At the end of the study period platelet aggregation ex vivo and endothelium-dependent and -independent vascular function of isolated aortic rings in vitro was assessed. RESULTS: Urinary 2,3-dinor-TXB2 excretion significantly increased in the cholesterol group (p < 0.05), and endogenous NO formation (measured as urinary nitrate excretion) decreased (p < 0.05). Both parameters were significantly correlated with each other (R = 0.48, p < 0.01). L-arginine partly restored urinary nitrate excretion and significantly reduced TXA2 production to values even below those in the control group (p < 0.001). Urinary 2,3-dinor-6-keto-PGF1 alpha excretion increased in early hypercholesterolaemia and returned to control values in the second half of the study period. The early increase in urinary 2,3-dinor-6-keto-PGF1 alpha excretion was attenuated by L-arginine. Platelet aggregation was significantly enhanced in cholesterol-fed rabbits and attenuated by dietary L-arginine. L-arginine also improved the impaired endothelium-dependent relaxations to ADP, and normalized the vasoconstrictor effects of 5-HT in isolated aortic rings. CONCLUSIONS: Cholesterol-feeding enhances platelet aggregation and TXA2 formation, and stimulates platelet-endothelial cell interaction in rabbits. These effects are probably due to impaired NO elaboration, as indicated by decreased urinary nitrate excretion. Chronic dietary supplementation with L-arginine elevates systemic NO elaboration and significantly increases the PGI2/TXA2 ratio. It thus beneficially influences the homeostasis between vasodilator and vasoconstrictor prostanoids in vivo.     

Effects of very low dose and enteric-coated acetylsalicylic acid on prostacyclin and thromboxane formation and on bleeding time in healthy subjects

OBJECTIVE: Low dose acetylsalicylic acid (ASA) is widely used as an anti-aggregatory agent in the primary and secondary prevention of cardiovascular diseases. In an effort to spare prostacyclin formation and to reduce gastrointestinal side-effects, both very low doses and enteric-coated formulations of ASA have been introduced. However, it still remains unclear whether these different formulations and dosages are equally effective with respect to inhibition of platelet aggregation and thromboxane A2 (TXA2) formation. METHODS: In a randomized study, we therefore investigated the effects of 100 mg ASA plain (p), 100 mg ASA enteric-coated (ec) and 40 mg ASA (p) to 36 healthy male subjects given for 7 days on platelet aggregation and endogenous prostanoid formation rates. Platelet aggregation and platelet TXB2 release in platelet rich plasma (PRP) and serum TXB2 and 6-keto-PGF1alpha levels were determined at baseline and after 7 days of each medication. The urinary metabolites of TXA2 (2,3-dinor-TXB2) and prostacyclin (2,3-dinor-6-keto-PGF1alpha) were measured by gas chromatography/tandem mass spectrometry in 24-h-urines at baseline and on day 7 of each medication. RESULTS: Collagen-induced platelet aggregation was 73.1+/-1.6% of maximal aggregation at baseline. It was inhibited by 68.9%, 58.6% and 24.0% by ASA 100 mg plain, 100 mg enteric-coated, and 40 mg plain on day 7, respectively. Platelet TXB2 release was 11592.0+/-367.5 pg x ml(-1) PRP. It was inhibited by 90.1%, 86.5%, and 55.2% by ASA 100 mg plain, 100 mg enteric-coated, and 40 mg plain, respectively. Serum TXB2 was almost completely reduced on day 7 by 100 mg ASA, but not by 40 mg ASA; serum 6-keto-PGF1alpha was slightly, but significantly reduced in all three groups. Urinary 2,3-dinor-TXB, excretion was 196.0+/-41.5 pg x mg(-1) creatinine at baseline. It was reduced by 80.3% and 79.1% by ASA 100 mg plain and enteric-coated, respectively (each P < 0.05 versus baseline), but only by 55.4% by ASA 40 mg plain (P < 0.05 versus both formulations of ASA 100 mg). CONCLUSIONS: Our present data show that the plain and enteric-coated formulations of 100 mg ASA are equally effective in inhibiting platelet aggregation, platelet thromboxane production, and urinary 2,3-dinor-TXB2 excretion rates. In contrast, a very low dose of 40 mg ASA was significantly less effective in inhibiting these indices of platelet activation in healthy human subjects. ASA enteric-coated 100 mg may be a useful alternative to 100 mg ASA (p) in patients with gastrointestinal side-effects, whereas 40 mg ASA (p) may be too low to inhibit sufficiently platelet activity in patients with cardiovascular diseases in whom platelet activity is increased.     

Echinococcus granulosus infection of farm dogs of Iran

The non-enzymatic metabolites of prostacyclin (PGI2) and thromboxane A2 (TXA2), 6-keto-prostaglandin F1alpha (6-keto-PGF1alpha) and thromboxane B2 (TXB2), and their 2,3-dinor metabolites, 2,3-dinor-6-keto-PGF1alpha and 2,3-dinor-TXB2, were measured in early morning urine samples in 24 in vitro fertilization (IVF) cycles in 24 women and in 27 women who became pregnant after IVF and embryo transfer (ET). The sum of the non-enzymatic metabolites and their 2,3-dinor metabolites was considered to be a reflection of total PGI2 and total TXA2 production in vivo. Both the ratio of 'total' PGI2/'total' TXA2 and the ratio of the 2,3-dinor metabolites were calculated. TXB2 concentrations showed virtually no change and the ratios of the non-enzymatic metabolites of PGI2 and TXA2 versus their 2,3-dinor metabolites remained relatively constant. As a consequence, the ratio of 2,3-dinor-6-keto-PGF1alpha/2,3-dinor-TXB2 was a close reflection of the ratio of 'total' PGI2/'total' TXA2, although the latter ratio was significantly higher all the time. We conclude that for comparative studies on the balance between PGI2 and TXA2 in IVF cycles and during gestation, the determination of the 2,3-dinor metabolites alone can replace the measurement of all four metabolites.     

Interaction of a thrombin inhibitor and a platelet GP IIb/IIIa antagonist in vivo: evidence that thrombin mediates platelet aggregation and subsequent thromboxane A2 formation during coronary thrombolysis

We examined the effect of a specific thrombin inhibitor, Ro 46-6240, alone and combined with an antagonist of the platelet GP IIb/IIIa, Ro44-9883, on the response to tissue-type plasminogen activator in a canine model of thrombolysis. Platelet activity was determined by measuring the excretion of 2,3-dinorthromboxane (TX)B2, an enzymatic metabolite of TXA2. Ro 46-6240 administered before tissue-type plasminogen activator induced a dose-dependent prolongation of the activated partial thromboplastin time and prothrombin time. The time to reperfusion decreased dose-dependently (P < .01) to 10 +/- 6 min vs. 52 +/- 5 min in controls. Ro 46-6240 also prevented reocclusion, which occurred in every case in control experiments. Urinary excretion of 2,3-dinor-TXB2 increased from 3 +/- 1 to 37 +/- 9 ng/mg creatinine in controls after reperfusion. This increase was reduced in a dose-dependent fashion by Ro 46-6240, such that at the highest dose, urinary 2,3-dinor-TXB2 after reperfusion was 5.6 +/- 1 ng/mg creatinine. Similar functional and biochemical effects were seen when a subthreshold dose of Ro 46-6240 was combined with Ro 44-9883. At the dose used, Ro 44-9883 alone abolished platelet aggregation ex vivo but failed to modify the response to tissue-type plasminogen activator or the excretion of 2,3-dinor-TXB2 after reperfusion (51 +/- 6 ng/mg creatinine, n = 3). However, the combination of Ro 44-9883 and Ro 46-6240 reduced the time to reperfusion (40 +/- 8 vs. 68 +/- 15 min; n = 7, P < .05), prevented reocclusion and abolished the rise in urinary 2,3-dinor-TXB2 (5 +/- 1 ng/mg creatinine, n = 4). These findings suggest that thrombin mediates platelet activation during coronary thrombolysis. The increased platelet activity results in platelet aggregation and a subsequent increase in TXA2 formation.     

Prognostic significance of normal dobutamine-atropine stress sestamibi scintigraphy in women with chest pain

The high risk of vaso-occlusive events in children younger than 4 years with cyanotic congenital heart disease and polycythaemia has been attributed to increased thromboxane (Tx) A2 formation. In older children with cyanotic congenital heart disease, however, the risk of vaso-occlusive events is much lower. We therefore hypothesized that the formation of TxA2 and prostacyclin is not disturbed in this age group. We measured urinary excretion of stable index metabolites of in vivo TxA2 and prostacyclin formation by gas chromatography-mass spectrometry in nine children (age 5.9-14.4, median 8.7 years) with cyanotic congenital heart disease, and in nine healthy, age-matched control subjects. The patients excreted less 2,3-dinor-TxB2 (systemic TxA2 formation, P = 0.03), 2,3-dinor-6-keto-PGF1 alpha (systemic prostacyclin formation. P = 0.03) and TxB2 (renal TxA2 formation, P = 0.01) than the control subjects. We conclude that in children older than 5 years with cyanotic congenital heart disease, endogenous synthesis of TxA2 and prostacyclin is not stimulated. This result may explain the lower risk of vaso-occlusive events in this age group as compared with younger children. In addition, our results suggest that chronic hypoxaemia may affect the in vivo formation of TxA2 and prostacyclin and the metabolic disposition of TxB2.     

Measurements of urinary prostaglandins in young ovulatory women during the menstrual cycle and in postmenopausal women

The purpose of the present work was to study the prostaglandin excretion in young nonpregnant ovulatory women during the menstrual cycle on the one hand and in postmenopausal women on the other hand and to investigate the influence of female sex hormones (estradiol, progesterone) on urinary prostanoid excretion. Urinary excretion rates of prostaglandin E2 (PGE2), 6-keto-PGF1 alpha, thromboxane B2 (TxB2) and their metabolites PGE-M (11 alpha-hydroxy-9, 15-dioxo-2,3,4,5,20-pentanor-19-carboxyprostanoic acid), 2,3-dinor-6-keto-PGF1 alpha, 2,3-dinor-TxB2 and 11-dehydro-TxB2 were determined by gas chromatography-triple stage quadrupole mass spectrometry (GC/MS/MS) in 41 young non-pregnant women during the follicular phase and during the luteal phase and in 23 postmenopausal women. Excretion rates of all urinary prostanoids were not significantly different in the follicular phase when compared with the luteal phase. In contrast to the young ovulatory women, PGE2 and TxB2 were significantly higher in postmenopausal women. Concerning the other prostaglandins significant differences between these groups of women did not exist. Although serum levels of estradiol and progesterone were different in young and postmenopausal women, sex hormones have not been shown to correlate with prostaglandins. Our data do not suggest sex hormones to be responsible for the difference in the prostaglandin excretion in women of reproductive age and in women in the menopause. Further systematic investigations into age dependency of prostaglandin excretion in women are necessary.     

Correction of increased prostacyclin synthesis in Bartter's syndrome by indomethacin treatment

The role of prostacyclin and thromboxane A2 in the pathogenesis of Bartter's syndrome was investigated by measurement of the urinary excretion of 6-keto-PGF1 alpha and thromboxane B2, respectively, in five patients. The prostaglandin metabolites were extracted from urine by a reproducible method and measured by specific radioimmunoassays. The patients with Bartter's syndrome excreted about four-times as much 6-keto-PGF1 alpha as the normal controls. In contrast, there was no significant difference in the urinary excretion of thromboxane B2 between the patients and the controls. In a second part of the study, three patients were treated with indomethacin (150 mg/day for four days), an inhibitor of prostaglandin synthesis. This regimen suppressed urinary excretion of 6-keto-PGF1 alpha by 43% and that of thromboxane B2 by 46%. It is suggested that the increase in prostacyclin production is responsible for both the hyperreninemia and and the other endocrine derangements as well as the hyporesponsiveness of blood pressure to intravenous infusion of vasopressors in patients with Bartter's syndrome.     

Effects of acetylsalicylic acid on peripheral hemodynamics in patients with chronic heart failure treated with angiotensin-converting enzyme inhibitors

Cyclooxygenase inhibitors may affect the hemodynamic status of patients with heart failure adversely and may also block the vasodilatory effects of angiotensin-converting enzyme (ACE) inhibitors in such patients. Relatively low doses of the cyclooxygenase inhibitor acetylsalicylic acid (ASA) are now used routinely in ischemic heart disease, the most important cause of heart failure. Therefore, we investigated the hemodynamic interaction between ASA and captopril in heart failure. In a randomized, cross-over study, 13 patients with congestive heart failure (CHF) who were already receiving maintenance treatment with an ACE inhibitor received a single dose of 25 mg captopril combined with 236 mg ASA or placebo. Peripheral blood flow was studied noninvasively by venous occlusion plethysmography of the calves. Liver blood flow was estimated from indocyanine green (ICG) clearance. Administration of captopril alone significantly decreased blood pressure (BP), and ICG clearance. Calf blood flow remained unchanged. However, after arterial occlusion, hyperemic calf blood flow persisted for longer. Captopril alone did not significantly change the plasma levels of the vasodilating prostaglandins PGI2 and PGE2 or the vasoconstricting thromboxane A2 (TXA2). In contrast, captopril combined with ASA reduced the plasma levels of these vasoactive substances, with significant decreases in PGE2 and TXA2 as compared with captopril alone, yet the hemodynamic alterations after captopril plus ASA were similar to those observed after captopril alone. A single antithrombotic dose of ASA (236 mg) in 13 patients with CHF [New York Heart Association (NYHA) class II-IV] undergoing chronic treatment with ACE inhibitors had no discernible effect on hemodynamic status.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of prostacyclin and thromboxane in lupus pregnancies: effect of small dose of aspirin

OBJECTIVES: To find out whether the tendency toward poor outcome in lupus pregnancies could be explained by changes in prostacyclin/thromboxane production, to relate these changes to the presence of antiphospholipid antibodies, and to study the potential benefits of low-dose aspirin. METHODS: We followed the urinary output of prostacyclin metabolites (6-keto-prostaglandin [PG]F1 alpha, 2,3-dinor-6-keto-PGF1 alpha) and thromboxane metabolites (thromboxane B2, 2,3-dinor-thromboxane B2) using high-pressure liquid chromatography followed by radioimmunoassay. We studied 14 pregnant women with systemic lupus erythematosus (SLE), of whom six had detectable antiphospholipid antibodies. The patients were randomized by a computerized program to receive either 50 mg aspirin daily (six women) or placebo (eight women). Nine healthy pregnant women served as controls. RESULTS: The production of prostacyclin was normal in early pregnancy in SLE patients but was reduced during late gestation in those without antiphospholipid antibodies. The production of thromboxane was increased in SLE patients compared with controls, and this increase was highest (two-to threefold rise) when antiphospholipid antibodies were detectable. Aspirin eliminated thromboxane dominance without affecting prostacyclin production. CONCLUSION: These data suggest that the presence of antiphospholipid antibodies in SLE patients may trigger thromboxane dominance, possibly contributing to the adverse outcome of these pregnancies. This thromboxane dominance can be eliminated with aspirin.     

Simultaneous solid phase extraction, derivatization, and gas chromatographic mass spectrometric quantification of thromboxane and prostacyclin metabolites, prostaglandins, and isoprostanes in urine

The current analytical methods for the various prostanoids require a separate and extended sample workup, derivatization, and gas chromatographic/mass spectrometric detection of each compound. Therefore, we developed and validated a rapid method for the common purification, derivatization, and GC/MS determination of 11-dehydrothromboxane B2, 2,3-dinor-6-keto-PGF1a, PGF2A, PGE2, PGD2, and isoprostanes in urine. A single reversed-phase solid-phase extraction step and modified reaction conditions yielded excellent sample purification at high recoveries and efficient derivatization for all compounds in one vial. The method allows, for the first time, the simultaneous quantification of these index metabolites of systemic thromboxane and prostacyclin synthesis, renal prostaglandin formation, and nonenzymatic in vivo lipid peroxidation in a single GC/MS run with high sensitivity and precision.     

Establishment of microanalysis of prostaglandin metabolites by GC/MS and its clinical application

We established microdetermination methods of prostaglandin (PG) metabolites by GC-selected ion monitoring (GC-SIM) and applied them to the clinical investigations. At first the microdetermination of delta 17-6-keto-PGF1 alpha, a hydrolyzed metabolite of PGI2, is described. An authentic delta 17-6-keto-PGF1 alpha was prepared from eicosapentaenoic acid (EPA) incubated with a homogenate from the bovine aortic intima. [18O] delta 17-6-Keto-PGF1 alpha was synthesized to obtain an internal standard for GC-SIM of delta 17-6-keto-PGF1 alpha. A good linear response over the range of 10 pg-5 ng was demonstrated. Chromatographic conditions using a MP-65HT column presented nearly baseline separation of delta 17-6-keto-PGF1 alpha and 6-keto-PGF1 alpha. Furthermore, a monoclonal antibody against cis-3-hexen-1-ol was prepared and used to separate and/or concentrate delta 17-6-keto-PGF1 alpha in the human blood sera. Using the prepared immunoaffinity columns of this antibody, delta 17-6-keto-PGF1 alpha was clearly detected in the human blood sera by GC/MS analysis. We were able to detect delta 17-6-keto-PGF1 alpha of the amount ranging from 6 to 26 pg/ml in the human blood plasma. The present method can be applied to the determination of delta 17-6-keto-PGF1 alpha in the human urine and plasma. Diabetes mellitus induces platelet alterations such as hyperaggregation. Variations in PG production seem to be related to this phenomenon but the changes in PG levels remain unclear. So we microanalyzed the 11-dehydrothromboxane B2 (TXB2) and 2,3-dinor-6-keto-PGF1 alpha, which were stable metabolites of TXA2 and PGI2, in the urine and investigated the relationship between the thromboxane/prostacyclin (TX/PGI) ratio and diabetes mellitus. The TX/PGI ratio in the urine of diabetics was higher than that of healthy volunteers. In murine, the TX/PGI ratio of STZ-induced mice was also higher than that of non-induced mice. The ratio of db/db mice also increased with the progress of diabetes mellitus. Furthermore, we investigated the relationship between the retinal vein occlusion (RVO), a thrombotic disease in which the retinal vein is blocked by blood aggregations, and the TX/PGI ratio. The TX/PGI level in patients with the RVO, who were not combine diabetes, was significantly higher than that in healthy volunteers. One of the causes of the RVO may be due to the variation of thromboxane production. This GC-SIM method can be used to determine the TX/PGI ratio in the urine.     

Nitric oxide as a regulator of prostacyclin synthesis in cultured rat heart endothelial cells

The effects of nitric oxide (NO) and its second messenger cyclic guanosine monophosphate (cGMT) on prostacyclin (PGI2) synthesis were studied in cultured rat heart endothelial cells using three different non-enzymatic nitric oxide releasing substances as well as inhibitors of nitric oxide synthase and of soluble guanylate cyclase. Production of prostacyclin, measured as 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha), was stimulated up to 1.7 fold in endothelial cells treated with the NO donors SIN-1 (3-morpholino sydnonimine), GEA 3162 (3-aryl-substituted oxatriazole imine) and GEA 3175 (3-aryl-substituted oxatriazole sulfonyl), chloride). In each case the synthesis of cGMP increase as much as 40-100 fold. An inhibitor of NO synthase, NG-nitro-L-arginine methyl ester (L-NAME), decreased the basal production of 6-keto-PGF1 alpha in non-stimulated endothelial cells, an effect that could be reversed by the NO donors SIN-1, GEA 3162 and GEA 3175. cGMP formation in the L-NAME treated endothelial cells was unaltered. The guanylate cyclase inhibitors, methylene blue (100 mumol/l) and LY83583 (100 mumol/l), caused a 1.5-10 fold increase in 6-keto-PGF1 alpha production while NO-donor-stimulated endothelial cGMP production was decreased by 10 to 90%. However, when SIN-1 was used as a stimulant, LY83583 had no significant effect on the production of cGMP. These findings support the hypothesis that NO stimulates prostacyclin production directly by activating cyclooxygenase. The results also suggest that NO could have an indirect effect on prostacyclin production via cGMP.     

Determination of seven prostanoids in 1 ml of urine by gas chromatography-negative ion chemical ionization triple stage quadrupole mass spectrometry

In an isotope dilution assay, prostaglandin (PG) E2, 6-keto-PGF1 alpha, thromboxane (Tx) B2 and their metabolites PGE-M (11 alpha-hydroxy-9,15-dioxo-2,3,4,5,20-pentanor-19-carboxyprostano ic acid), 2,3-dinor-6-keto-PGF1 alpha, 2,3-dinor-TxB2 and 11-dehydro-TxB2 were determined in urine by gas chromatography-triple stage quadrupole mass spectrometry (GC-MS-MS). After addition of deuterated internal standards, the prostaglandins were derivatized to their methoximes and extracted with ethyl acetate-hexane. The sample was further derivatized to the pentafluorobenzylesters and purified by thin-layer chromatography (TLC). Three zones were scraped from the TLC plate. The prostanoid derivatives were converted to their trimethylsilyl ethers and the products were quantified by GC-MS-MS. In each run, two or three prostanoids were determined.     

Effects of captopril related to increased levels of prostacyclin and angiotensin-(1-7) in essential hypertension

OBJECTIVE: To evaluate the contribution of angiotensin-(1-7) [Ang-(1-7)] and prostaglandins to the acute and long-term antihypertensive actions of captopril in mild-to-moderate essential hypertensive patients. DESIGN AND METHODS: Blood pressure, cardiac rate and the plasma concentrations of angiotensin I (Ang I), angiotensin II (Ang II), Ang-(1-7), prostaglandin E2 and 6-keto prostaglandin F1 alpha (the breakdown product of prostacyclin) were determined in the peripheral venous blood of 24 essential hypertensive subjects before and 3 h after administration of 50 mg captopril. Eleven of 24 patients completed a 6-month treatment period with captopril monotherapy (50 mg twice a day). The hemodynamic and hormonal response produced by a last 50 mg dose of captopril was determined once again in the 11 subjects who maintained blood pressure control with captopril monotherapy for 6 months. RESULTS: The fall in blood pressure produced 3 h after drug intake was comparable for the first and the last 50 mg captopril dose. Although the first response to captopril increased plasma levels of Ang I only, the response to the last dose of the drug (6 months after) caused significantly higher levels of Ang I and Ang-(1-7). Neither acute nor chronic therapy with captopril had a significant effect on plasma concentrations of Ang II. Although plasma levels of prostaglandin E2 and 6-keto prostaglandin F1 alpha were not modified by a first exposure to captopril, the concentrations of 6-keto prostaglandin F1 alpha but not prostaglandin E2 rose significantly in subjects treated with the inhibitor for 6 months. A negative correlation was also demonstrated between diastolic blood pressure and plasma Ang-(1-7) levels in the 11 essential hypertensive subjects in whom blood pressure was controlled with captopril monotherapy. CONCLUSIONS: Inhibition of angiotensin converting enzyme with captopril had a significant effect on blood pressure that was not directly accounted for by a suppression of plasma Ang II levels. Continuous therapy with captopril unmasked a contribution of Ang-(1-7) and prostacyclin to the antihypertensive actions of this drug.     

Effects of dietary fat saturation on eicosanoid production, platelet aggregation and blood pressure

The effects of dietary fat saturation on eicosanoid urinary excretion, platelet aggregation (PA) and blood pressure (BP) were studied in 42 healthy subjects. They consumed four consecutive diets differing in their fat saturation [saturated (SFA); monounsaturated (MUFA); polyunsaturated n-6 (PUFA n-6); and polyunsaturated n-6/n-3, (PUFA n-3)]. Each diet period lasted 5 weeks. There were no differences in 24-h 2,3-dinor-6- keto-prostaglandin F1 alpha excretion among dietary periods. A significant effect was noted regarding the excretion of 11-dehydro-thromboxane B2 (P < 0.0001). During the PUFA n-6 phase the excretion was significantly higher than during SFA and MUFA periods. Dietary fatty acid composition had a significant effect on ADP (1 mumolL-1) and collagen (2 mgL-1) induced PA. Dietary fat also had a significant effect on systolic and diastolic blood pressure (P < 0.0001). Both were significantly higher during the SFA period than during the other three periods. Our findings suggest that changes in dietary fatty acids may have mild, but significant, effects on eicosanoid production, platelet aggregation and blood pressure.     

Prostacyclin formation elicited by endothelin-1 in rat aorta is mediated via phospholipase D activation and not phospholipase C or A2

Endothelin-1 (ET-1) is a potent vasoconstrictor peptide that also stimulates production of prostacyclin (PGI2) from arachidonic acid. The purpose of this study was to determine the contribution of phospholipases (PLs) A2, C, and/or D in ET-1-induced PGI2 formation in the rat aorta, measured as immunoreactive 6-ketoprostaglandin (PG) F1 alpha. ET-1 increased 6-keto-PGF1 alpha formation, which was not affected by a PLA2 inhibitor, 7,7-dimethyl eicosadienoic acid (DEDA). Furthermore, ET-1 failed to stimulate PLA2 activity measured in the cytosol (cPLA2), using phosphatidylcholine, L-a-1-palmitoyl-2-arachidonyl[14C] as a substrate. However, the adrenergic agonist norepinephrine increased 6-keto-PGF1 alpha formation, which was attenuated by DEDA, and enhanced PLA2 activity. ET-1 enhanced PLC activity, as indicated by increased inositol phosphate production, which was prevented by a PLC inhibitor, U-73122. However, ET-1-induced 6-keto-PGF1 alpha production was not altered by U-73122. An inhibitor of PLD activation, C2-ceramide, attenuated ET-1-induced PLD activity, as indicated by the production of phosphatidylethanol. Furthermore, ET-1-induced 6-keto-PGF1 alpha formation was inhibited by C2-ceramide as well as by ethanol treatment. Moreover, inhibitors of phosphatidate phosphohydrolase (propranolol) and diacylglycerol lipase (RHC-80267), attenuated ET-1-induced 6-keto-PGF1 alpha formation. Finally, ET-1-induced activation of PLD was not attenuated by a selective PKC inhibitor, bisindolylmaleimide I. These data suggest a novel pathway for ET-1-induced PGI2 formation in the rat aorta involving activation of PLD but not cPLA2 and independent of PLC or PKC activation.     

Chronic inhibition of nitric oxide synthase in Heymann nephritis

Effects of nitric oxide (NO) synthase inhibition on blood pressure and on the course of Heymann nephritis was examined in rats. L-NG-nitroarginine-methylester (L-NAME, 10 mg/100 ml in the drinking water for 12 weeks) was used as an inhibitor of NO synthase. Urinary excretion of guanosine 3',5'-cyclic monophosphate (cGMP), a second messenger of NO, was used as an indirect estimate of NO activity. Rats were divided into the following groups: control, nephritis, L-NAME, and nephritis-L-NAME. Urinary cGMP excretion was lower in the nephritis group (p < 0.05) and in the nephritis-L-NAME group (p < 0.005) compared with controls. Plasma atrial natriuretic peptide (ANP) levels were elevated in the nephritis (p < 0.001) and in the nephritis-L-NAME groups (p < 0.05. L-NAME treatment alone did not have any effect on plasma ANP levels. Blood pressure rose progressively in all L-NAME-treated rats. Most marked albuminuria developed in the nephritis-L-NAME group. No differences in the immunohistological findings were observed between the nephritis and the nephritis-L-NAME groups. NO synthase inhibition causes hypertension and aggravates albuminuria in chronic nephritis. Moreover, nephritis itself may decrease then production of cGMP either as a consequence of blunted NO activity or, in addition, because of ANP resistance. It appears that NO synthase inhibition does not change the immunological course of Heymann nephritis but rather the increased hemodynamic load makes the course of nephritis worse.     

Aspirin inhibits both lipid peroxides and thromboxane in preeclamptic placentas

Preeclampsia is a hypertensive disorder of human pregnancy that is a leading cause of premature delivery and fetal growth retardation. It is characterized by hypertension, reduced uteroplacental blood flow, proteinuria, and edema. Preeclampsia is associated with an imbalance of increased thromboxane and decreased prostacyclin, as well as with an imbalance of increased lipid peroxides and decreased antioxidants. Low-dose aspirin (ASA) therapy (60-150 mg/day) is being evaluated for the prevention of preeclampsia. The rationale for this is that low-dose ASA selectively inhibits thromboxane synthesis without affecting prostacyclin synthesis. We hypothesized that ASA might also inhibit the synthesis of lipid peroxides. The purpose of this study was to examine the effects of aspirin on lipid peroxide, thromboxane, and prostacyclin production rates in placentas obtained from women with preeclampsia. Placentas were obtained from five preeclamptic women. Placental tissues (350 mg) were incubated in Dulbecco's Modified Eagles Medium (DMEM) for 48 h, alone and with varying concentrations of aspirin: 1 x 10(-6) M, 1 x 10(-5) M, 5 x 10(-5) M, 1 x 10(-4) M, and 5 x 10(-4) M. Samples were collected at 0, 2, 6, 16, 28, and 48 h of incubation, and analyzed for thromboxane and prostacyclin by RIA of their stable metabolites, thromboxane B2 and 6-keto-PGF1 alpha, and for lipid peroxides by peroxide equivalents. As compared to control, an aspirin concentration of 5 x 10(-5) M significantly inhibited (p < 0.05) both lipid peroxides (3.15 +/- 0.49 vs. 1.90 +/- 0.31 pmol/microgram/h) and thromboxane (0.66 +/- 0.11 vs. 0.32 +/- 0.10 pg/microgram/h), but not prostacyclin (0.24 +/- 0.05 vs. 0.17 +/- 0.02 pg/microgram/h, p > 0.05). Lower aspirin doses (1 x 10(-6) M, 1 x 10(-5) M) had no effect, whereas higher doses (1 x 10(-4) M and 5 x 10(-4) M) inhibited all three compounds. We conclude that aspirin inhibits lipid peroxides, as well as thromboxane and prostacyclin, in preeclamptic placentas. The inhibitory effects are dose dependent. Low-dose aspirin (5 x 10(-5) M) selectively inhibits lipid peroxides and thromboxane without affecting prostacyclin. We speculate that the selective inhibitory effect of low-dose aspirin may account for its effectiveness in the prevention of preeclampsia.     

Change in nasopharyngeal carriage of Streptococcus pneumoniae resulting from antibiotic therapy for acute otitis media in children

Although theophylline has been used in the treatment of asthma for decades, it is not a first line choice any more. It is a well-known bronchodilator, but was recently discovered also to be an anti-inflammatory, immunomodulatory and bronchoprotective agent. Therefore we wanted to establish the role of theophylline on prostaglandin and leukotriene production, which plays a part in the pathogenesis of asthma. Theophylline was infused (bolus 5 mg/kg in 15 min and infusion 0.4 mg/kg/h for 1 h 45 min) into healthy volunteers. Thromboxane B2, prostaglandin E2 and leukotriene E4 were measured from the A23187-stimulated whole blood samples and stable metabolites of thromboxane A2; prostacyclin and leukotriene E4 were measured from urine. Theophylline increased prostaglandin E2 production and decreased leukotriene E4 production ex vivo in whole blood, thus increasing the prostanoid/leukotriene ratio. It did not change thromboxane B2 production stimulated by either spontaneous clotting or A23187 in the whole blood. Theophylline had hardly any effect on in vivo thromboxane, prostacyclin and leukotriene E4 production measured as urinary metabolites, 11-dehydro-thromboxane B2, 2,3-dinor-6-keto-prostaglandin F1alpha and leukotriene E4, respectively. Serum theophylline concentrations were at the lower level of normal therapeutic range during the infusion. The increase in PGE2 and the decrease in LTE4 synthesis ex vivo may offer a new explanation for the mode of antiasthmatic action of theophylline. It is notable that this phenomenon occurs at low serum theophylline concentrations. These results confirm the idea that theophylline has an anti-inflammatory and bronchoprotective action and support the use of theophylline as a therapeutic agent in asthmatic patients.     

Phospholipid antibodies and resistance to activated protein C in women with thrombophilia

The influence of captopril and ritanserin (5-HT2 receptor antagonist) on the serotonin-induced contraction of isolated tail artery of two-kidney, one clip hypertensive (2K, 1C-RHR) and normotensive rats was studied. Captopril, administrated in a single dose of 100 mg/kg, po or in a dose of 30 mg/kg/day, po for one week, diminished the systolic blood pressure in both groups of rats. Ritanserin (0.01 mg/kg/day, sc for one week) administrated alone or in combination with captopril (30 mg/kg/day) evoked such effect only in 2K, 1C-RHR. The reactivity of rat tail artery isolated from 2K, 1C-RHR to serotonin was significantly higher than that obtained from normotensive ones. Captopril given in a single dose or chronically did not change the serotonin-induced contraction of rat tail artery in normotensive rats. The attenuation of serotonin contractile effect was registered in hypertensive rats pre-treated with captopril. Administration ritanserin with or without captopril also inhibited the response to serotonin in both groups. The stronger effect was observed after administration of ritanserin with captopril in 2K, 1C-RHR than in control group. Our results indicate, that in 2K, 1C-RHR the reactivity of rat tail arteries to serotonin is enhanced and this effect can be reduced by chronic captopril administration.