Effects of feeding ethyl-dihomo-gamma-linolenate on prostaglandin biosynthesis and platelet aggregation in the rabbit.

1. The ethyl ester of dihomo-gamma-linolenic acid (20:3omega6) (1 g/kg/day) was fed to rabbits for 25 days. Plasma lipids and platelet aggregation were analyzed on day 1, 11, 16, 21 and 26. 2. All plasma lipid classes were greatly enriched with 20:3omega6. Arachidonic acid levels were elevated to a smaller extent. The different platelet phospholipid fractions analyzed were also highly enriched with 20:3omega6, whereas the arachidonic acid content in platelet phospholipids was significantly lower than in control animals. 3. The excretion of 7 alpha-hydroxy-5,11-diketotetranorprostane-1,16-dioic acid, the major urinary metabolite of prostaglandin E1 and E2 was increased 4.6 fold by the treatment. 4. Platelet aggregation in response to ADP, collagen and arachidonic acid did not differ at any time betweeen 20:3omega6 treated rabbits and controls. 5. It is concluded that prostaglandin E biosynthesis can be increased by enriching the prostaglandin precursor pool. Platelet aggregation in vitro is not altered by feeding ethyl 20:3omega6.

Role of oxidant stress in endothelial dysfunction produced by experimental hyperhomocyst(e)inemia in humans.

BACKGROUND: Moderate elevations in plasma homocyst(e)ine concentrations are associated with atherosclerosis and hypertension. We tested the hypothesis that experimental perturbation of homocysteine levels produces resistance and conduit vessel endothelial dysfunction and that this occurs through increased oxidant stress. METHODS AND RESULTS: Oral administration of L-methionine (100 mg/kg) was used to induce moderate hyperhomocyst(e)inemia ( approximately 25 micromol/L) in healthy human subjects. Endothelial function of forearm resistance vessels was assessed by use of forearm vasodilatation to brachial artery administration of the endothelium-dependent dilator acetylcholine. Conduit vessel endothelial function was assessed with flow-mediated dilatation of the brachial artery. Forearm resistance vessel dilatation to acetylcholine was significantly impaired 7 hours after methionine (methionine, 477+/-82%; placebo, 673+/-110%; P=0.016). Methionine did not alter vasodilatation to nitroprusside and verapamil. Flow-mediated dilatation was significantly impaired 8 hours after methionine loading (0.3+/-2.7%) compared with placebo (8. 2+/-1.6%, P=0.01). Oral administration of the antioxidant ascorbic acid (2 g) prevented methionine-induced endothelial dysfunction in both conduit and resistance vessels (P=0.03). CONCLUSIONS: Experimentally increasing plasma homocyst(e)ine concentrations by methionine loading rapidly impairs both conduit and resistance vessel endothelial function in healthy humans. Endothelial dysfunction in conduit and resistance vessels may underlie the reported associations between homocysteine and atherosclerosis and hypertension. Increased oxidant stress appears to play a pathophysiological role in the deleterious endothelial effects of homocysteine.

An effect of estradiol and testosterone on the calcium pump activity and phospholipid fatty acid composition of rat liver microsomes.

Castration of the male rat resulted in a 28% reduction of the specific activity of liver microsomal calcium uptake, three weeks after castration. Treatment of the castrated animals with testosterone during this period returned calcium uptake to control levels. Treatment with estradiol resulted in a reduction of calcium uptake to a level less than 25% of that seen in the normal male. Although testosterone treatment had only a small effect on the fatty acid composition of liver microsomal phospholipids in the castrated male, there were significant changes of linoleic acid (18:2) in phosphatidylcholines and of palmitic acid (16:0) in phosphatidylethanolamines, when compared to the untreated castrated male rat. Administration of estradiol to the castrated male rat resulted in a marked decrease of palmitic acid (16.0) and linoleic acid (18.2) in all three phospholipid fractions studied. Stearic acid (18.0) was significantly increased in the phosphatidylcholines and phosphatidylethanolamines by estradiol treatment. The phospholipid and calcium uptake changes seen after treatment of the castrated rat with testosterone or estradiol are consistent with the sex-related differences observed in the intact, adult rat liver microsomes.

Dietary fatty acids in human thrombosis and hemostasis.

The effects of fatty acids on hemostasis are controversial. It has been difficult to show convincing effects of saturated or monounsaturated fatty acids that are clearly related to hemostatic variables in humans. Unsaturated fatty acids alter platelet aggregation and processes related to coagulation and fibrinolysis. Indirect evidence exists that n-6 polyunsaturated fatty acids may exert favorable effects on thrombotic processes in vivo, but large clinical trials have failed to show benefits of 5-6 g linoleic acid (18:2n-6) or linolenic acid (18:3n-3)/d. Only long-chain n-3 fatty acids prolong the template bleeding time, and they may exert some beneficial effect on erythrocyte flexibility. It appears unlikely that n-3 fatty acids lower fibrinogen or interact with the fibrinolytic system directly. One prospective secondary prevention trial showed benefits that may have resulted from either an improved hemostatic profile or an antiarrhythmic effect. A similar time course of clinical improvement was noted with reduced rates of cardiac mortality and postoperative thrombosis in Norway during World War II, and this was associated with a drastic dietary alteration involving increased consumption of n-3 fatty acids and reduced consumption of saturated fatty acids. Further work is needed to develop better tools to examine in vivo hemostasis so that the mechanisms and eventual clinical utility of n-3 fatty acids can be elucidated in well-designed clinical trials.

Fatty acid-related functions.

The first recommendations for specific nutrient quantities that must be obtained to support health were made by the US Department of Agriculture before 1939. Hazel Stiebeling was the leader of this effort and the scientific background was published in the Yearbook of Agriculture. The recommendations clearly stated that food must be available to provide the nutrients to support health. The science of nutrition in the United States is engaged in the most thorough review and reexamination of the recommended dietary allowances in at least a generation of nutrition scientists. There is a new awareness of nutrition complexity and the likelihood of identification of new essential nutrients. This meeting was devoted to the search for functional endpoints to reach quantitative estimates of dietary substances needed to support a function. Included in that concept is determining a range of individual needs and identifying factors that alter these needs. We give the rationale for endpoints of fatty acid metabolism related to platelets and the risk of thrombosis, give the rationale for the recommendation for a new nutrient, and show the necessity for including nutrient interaction in the determination of needs for two nutrients.

Pentoxifylline does not alter the response to inhaled grain dust.

Pentoxifylline (PTX) has been shown to reduce sepsis-induced neutrophil sequestration in the lung and inhibit endotoxin-mediated release of tumor necrosis factor-alpha (TNF-alpha). Previously, we have shown that endotoxin appears to be the principal agent in grain dust causing airway inflammation and airflow obstruction following grain dust inhalation. To determine whether PTX affects the physiologic and inflammatory events following acute grain dust inhalation, 10 healthy, nonsmoking subjects with normal airway reactivity were treated with PTX or placebo (PL) followed by corn dust extract (CDE) inhalation (0.08 mL/kg), using a single-blinded, crossover design. Subjects received PTX (1,200 mg/d) or PL for 4 days prior to CDE inhalation and 400 mg PTX or PL on the exposure day. Both respiratory symptoms and declines in FEV1 and FVC occurred following CDE exposure in both groups, but there were no significant differences in the frequency of symptoms or percent declines from baseline in the FEV1 and FVC at any of the time points measured in the study. Elevations in peripheral blood leukocyte and neutrophil concentrations and BAL total cell, neutrophil, TNF-alpha, and interleukin-8 concentrations were measured 4 h following exposure to CDE in both the PTX- and PL-treated subjects, but no significant differences were found between treatment groups. These results suggest that pretreatment with PTX prior to inhalation of CDE, in the doses used in this study, does not alter the acute physiologic or inflammatory events following exposure to inhaled CDE.

Leukotriene B4 synthesis and neutrophil chemotaxis in chronic granulocytic leukaemia.

A sensitive gas chromatography-mass spectrometric method was used to measure the generation in whole blood of leukotriene B4 (LTB4), a potent stimulator of neutrophil chemotaxis, in eight patients with chronic granulocytic leukaemia (CGL) and 12 healthy controls. LTB4 was detectable in unstimulated samples from all the patients (mean 194 (70 SEM) pg/ml), and the capacity for LTB4 production after stimulation with calcium ionophore (A23187) was similar in patients (32.1 (11) ng/10(6) leucocytes) and controls (38.1 (4) ng/10(6) leucocytes). In response to stimuli which induce neutrophil activation, LTB4 production was significantly greater in the patients than in controls: 35.6 (13) v 13.0 (3) ng/ml, p less than 0.05 (f-met-leu-phe); and 42.4 (16) v 14.7 (4) ng/ml, p less than 0.02 (opsonised zymosan). Anti-IgE stimulated considerably more LTB4 production in patients with CGL than in controls (3.86 (1.6) v 0.83 (0.43) ng/ml; p less than 0.005) and this correlated significantly (p less than 0.05) with the basophil count. Neutrophil chemotaxis to LTB4, however, was significantly impaired in the patients with CGL even at the highest concentration of LTB4 (10(-5) M). Chemotaxis to f-met-leu-phe, phagocytosis, and bacterial killing were normal. Thus although LTB4 synthesis is normal or even enhanced in patients with CGL, specific defects in LTB4-mediated responses may contribute to neutrophil dysfunction in this disease.

Effects of low-dose aspirin on endogenous eicosanoid formation in normal and atherosclerotic men.

We have investigated the effects of a low-dose aspirin regimen (120 mg orally, then 20 mg twice daily) on the in vivo synthesis of prostacyclin, thromboxane and prostaglandin E in man by measurement of their urinary metabolites (PGI2-M, TxB2-M, PGE-M) using gas chromatography-mass spectrometry. A comparison was made between the selectivity of low-dose aspirin for thromboxane vs prostacyclin synthesis in patients with atherosclerosis, age-matched controls without vascular disease, and young healthy volunteers. After one week of treatment, aspirin reduced TxB2-M synthesis to a similar extent in the three groups (mean declines of 86, 84 and 78% respectively), while there was an unexpected difference in effect on PGI2-M, with only a 27% fall in the young volunteers but 53% and 54% declines in the patients with vascular disease and their age-matched controls. Serum TxB2 was reduced greater than 98% in all groups while PGE-M excretion was unchanged. These results indicate that bioselectivity for inhibition of Tx synthesis by aspirin is more difficult to achieve in older subjects than in the young volunteers previously studied and that very low, frequent dosing, or a sustained-release preparation of aspirin would be needed to achieve bioselectivity for Tx inhibition in patients with vascular disease.

Regulation of the Immune Response by Eicosanoids: Pharmacology and Clinical Effects of Prostaglandin E in Aspirin-Sensitive Syndromes.

Immunomodulatory effects of prostaglandin E(2) (PGE(2)) have been documented both in vitro and in vivo. Our previous studies have examined the effects of intravenously administered PGE(2) in mast-cell-mediated diseases, including aspirin-sensitive asthma and systemic mast-cell-activation syndrome. The basis for investigations of these particular diseases has been the hypothesis that the inhibition of cyclooxygenase removes one of its products, PGE(2), that provides a critical restraint on the activation of the mast cell. Based on the beneficial effects of PGE(2) found in these studies, we have extended our investigations to an evaluation of misoprostol, the orally available analog of PGE(1). Our preliminary studies with this drug are consistent with an inhibition of mast cell activation by misoprostol, an effect observed at doses higher than currently recommended for gastric protection. The findings from these initial trials have led to the development of ex vivo whole-blood assays that assess the pharmacodynamics of misoprostol's immunomodulatory actions, which support the concept of employing higher doses to obtain sustained systemic effects. To extend these results, we have undertaken double-blinded, placebo-controlled clinical investigations to examine the effects of the higher doses of misoprostol (300--600 &mgr;g QID) given chronically in aspirin-sensitive asthma and systemic mast cell activation. Although still ongoing, our studies have confirmed by a variety of clinical evaluations that the higher doses of misoprostol can be tolerated by many patients and appear to be safe. Based on our findings and those of others, further investigation of the therapeutic usefulness of this drug or other PGE analogs in allergic and immunologic diseases appears warranted.

Longitudinal follow-up comparison of educational interventions: multimedia textbook, traditional lecture, and printed textbook.

RATIONALE AND OBJECTIVES: The goal of this prospective, interinstitutional study was to compare the long-term instructional effectiveness of a pediatric multimedia textbook (MMTB) to that of a standard lecture and a printed textbook. MATERIALS AND METHODS: A randomized cohort of 89 3rd-year medical students from two institutions were initially evaluated from June 1992 to June 1993 and reevaluated in May 1994. Students were randomly assigned to one of four instructional groups: computer-aided instruction by means of MMTBs (n = 21), traditional lecture (n = 23), printed textbook (n = 19), and a control group (n = 26). After instruction, all groups were tested by means of a multiple choice test at the end of their pediatric clerkship; they were given this same test 11-22 months later. RESULTS: The long-term instructional effectiveness of the MMTB, printed textbook, and lecture were the same as that in the control group, as determined by analysis of variance of mean test scores. CONCLUSION: The educational advantage of MMTBs observed immediately after instruction was not detected 1 year later. Because attrition reduced statistical power, further research is necessary to determine how educational fading affects these instructional formats.

Effects of feeding ethyl-dihomo-gamma-linolenate on rabbit renomedullary lipid composition and prostaglandin production in vitro.

Feeding the ethyl ester of dihomo-gamma-linolenic acid for 25 days to rabbits resulted in increased PGE1 (20 to 30-fold) and PGE2 (1.5-fold) output by a hormone responsive, in vitro, renal papilla preparation. The relative amount of PGE1 increased from less than 5% of PGE2 in controls to 25-35% of PGE2 in the papillae of 20 : 3 omega 6-supplemented animals. During the study renomedullary tri glycerides in the 20 : 3 omega 6-supplemented animals increased 2.8-fold compared to animals fed an equal amount of a control fatty acid mixture, and in addition to a marked enrichment in 20 : 3 omega 6, also contained increased proportions of 20 : 4 omega 6 and longer chain polyenes. The increase in triglyceride content found in the renal medulla was not seen in the renal cortex or liver. There was no increase in renomedullary phospholipid content during the study, and phopholipids of treated animals contained increased proportions of 20 : 3 omega 6 and 20 : 4 omega 6, but not longer chain polyenes. The results indicate that enriching the prostaglandin precursor pool by feeding 20 : 3 omega 6 can alter the type and amount of prostaglandin released by the renal papilla, at least in vitro. Also, the selective changes in amount and long chain polyene content of renomedullary triglycerides during the study suggest some special functions for this lipid class in prostaglandin precursor metabolism.

The effect of linoleic, arachidonic and eicosapentaenoic acid supplementation on prostacyclin production in rats.

We examined the effect of dietary supplementation of linoleic acid (LA), arachidonic acid (AA) or eicosapentaenoic acid (EPA) to rats fed a diet low in linoleic acid on in vitro and in vivo production of prostacyclin. Male Sprague Dawley rats were fed a high-fat diet (50% energy as fat, 1.5% linoleic acid) for two weeks. Three of the groups were then supplemented orally with either 90 mg/d of LA, AA or EPA, all as the ethyl esters, for a further two weeks while remaining on the high-fat diet. Forty-eight hour urine samples were collected at the end of the second and fourth weeks. In vivo prostacyclin production was determined by a stable isotope dilution, gas chromatography/mass spectrometry assay for the major urinary metabolite of prostacyclins (2,3-dinor-6-keto-PGF1 alpha or PGI2-M and delta 17-2,3-dinor-6-keto-PGF1 alpha or PGI3-M). In vitro prostacyclin production was determined by radioimmunoassay of the stable metabolite (6-keto-PGF 1 alpha) following incubation of arterial tissue. Oral supplementation with AA resulted in a rise in plasma and aorta 20:4n-6, and increased in vitro prostacyclin and urinary PGI2-M production. EPA supplementation resulted in a rise in plasma and aorta 20:5n-3 and 22:5n-3, and a decline in plasma 20:4n-6, but not in the aorta. In the EPA-supplemented group, the in vitro prostacyclin and the urinary PGI3-M increased, but urinary PGI2-M decreased. The increase in in vitro prostacyclin production in the EPA-supplemented rats was unexpected and without obvious explanation.(ABSTRACT TRUNCATED AT 250 WORDS)

Polyunsaturated fatty acids and cardiovascular function in man.

Dietary supplementation with either n-6 or n-3 polyunsaturated fatty acids has been reported to lower blood pressure in man, and to confer benefits in atherosclerotic vascular disease. Such effects have been suggested to be due to altered in vivo production of vasoactive prostaglandins, but this hypothesis has not been tested previously. Epidemiologic data are often quoted as showing lower blood pressure in populations with a high consumption of fish, but such conclusions are less clear from the primary data. This review presents some of the population studies on the relationship between fish intake, hypertension and vascular disease, discusses problems in studying blood pressure in dietary intervention trials, and reviews the published literature on reducing blood pressure and vascular reactivity with dietary n-3 supplements in volunteers. Recent work addressing a number of controversial points is presented, with the conclusion that pharmacologic doses of n-3, but not n-6, fatty acids can lower blood pressure in humans, but probably do not do so through directly lowering vascular reactivity to catecholamines or increasing the synthesis of vasodilator prostaglandins.

Polyunsaturates, endogenous eicosanoids, and cardiovascular disease.

The role of dietary polyunsaturated fats in the prevention of human vascular disease has not been defined, but population and intervention studies have suggested that omega-3 fatty acids (FAs) from marine lipids may have a number of potentially beneficial effects. Eicosanoids are extremely potent autacoids made from polyunsaturated fatty acids and have effects on many vascular parameters, so that the physiological effects of dietary supplementation with polyunsaturated fats are often attributed to alterations in endogenous eicosanoid production. Few studies have attempted to correlate in vivo eicosanoid synthesis and functional effects during such dietary maneuvers, however. This article reviews the relationship between dietary polyunsaturates and endogenous eicosanoid synthesis in man, with particular emphasis on recent studies of the effects of omega-3 FAs. Data on omega-3 FAs and platelet-vascular interactions, blood pressure, and vascular reactivity in human subjects are also summarized, with interpretation of recent work addressing a number of controversial points. A discussion of the significance and future direction of such investigations concludes that further clinical trials in selected patient groups are warranted.

n-3 fatty acids and human hypertension.

Clinical studies of the antihypertensive effects of n-3 fatty acids in different groups of patients are summarized, and recent information of possible mechanisms of antihypertensive effects is discussed. Because of marked differences between fatty acid metabolism and blood pressure control of humans and laboratory animals, the focus is on human studies.