Testosterone replacement therapy in blood donors modulates erythrocyte metabolism and susceptibility to hemolysis in cold storage.

BACKGROUND: Red blood cells (RBCs) derived from patients who receive testosterone replacement therapy (TRT) may be considered eligible for component production and transfusion. The aim of this study was to identify testosterone-dependent changes in RBC metabolism and to evaluate its impact on susceptibility to hemolysis during cold storage. STUDY DESIGN AND METHODS: We characterized stored RBCs from two cohorts of TRT patients who were matched with control donors (no TRT) based upon sex, age, and ethnicity. We further evaluated the impact of testosterone deficiency (orchiectomy) on RBC metabolism in FVB/NJ mice. RBC metabolites were quantified by ultra-high-pressure liquid chromatography-mass spectrometry. RBC storage stability was determined in RBC units from TRT and controls by quantifying storage, osmotic, and oxidative hemolysis. RESULTS: Orchiectomy in mice was associated with significant (P < 0.05) changes in RBC metabolism as compared with intact males including increased levels of acyl-carnitines, long-chain fatty acids (eg, docosapentaenoic acids), arginine, and dopamine. Stored RBCs from TRT patients exhibited higher levels of pentose phosphate pathway metabolites, glutathione, and oxidized purines (eg, hypoxanthine), suggestive of increased activation of antioxidant pathways in this group. Further analyses indicated significant changes in free fatty acids and acyl-carnitines in response to testosterone therapies. With regard to hemolysis, TRT was associated with enhanced susceptibility to osmotic hemolysis. Correlation analyses identified acyl-carnitines as significant modifiers of RBC predisposition to osmotic and oxidative hemolysis. CONCLUSIONS: These observations provide new insights into testosterone-mediated changes in RBC metabolome and biology that may impact the storage capacity and posttransfusion efficacy of RBCs from TRT donors.

Metabolism of fatty acids and bile acids in plasma is associated with overactive bladder in males: potential biomarkers and targets for novel treatments in a metabolomics analysis.

OBJECTIVES: The present study was conducted to identify metabolites using a metabolomics approach and investigate the relationship between these metabolites and urgency as a major symptom of overactive bladder (OAB). PATIENTS AND METHODS: In 47 male participants without any apparent neurological disease, OAB was defined as an urgency score on the International Prostate Symptom Score of 2 and higher (OAB group, n = 26), while patients with a score of 1 or 0 were placed in a control group (n = 21). A comprehensive study on plasma metabolites was conducted, and metabolites were compared between the OAB and control groups. RESULTS: Age was significantly higher in the OAB group, while prostate volume did not differ between the groups. A 24-h bladder diary revealed that nocturnal urine volume, 24-h micturition frequency, nocturnal micturition frequency, and the nocturnal index were significantly higher in the OAB group, whereas maximum voided volume was significantly lower in this group. The metabolomics analysis identified 79 metabolites from the plasma of participants. The multivariate analysis showed that increases in the fatty acids (22:1), erucic acid and palmitoleic acid, and a decrease in cholic acid correlated with incidence of male OAB. A decrease in acylcarnitine (18:2)-3 and an increase in cis-11-eicosenoic acid also appeared to be associated with OAB in males. CONCLUSIONS: OAB in males may occur through the abnormal metabolism of fatty acids and bile acids. Further studies on these pathways will contribute to the detection of new biomarkers and development of potential targets for novel treatments.

Association of plasma fatty acid alteration with the severity of coronary artery disease lesions in Tunisian patients.

BACKGROUND: Some factors related to diet are known to be involved in the progression of atherosclerosis in humans. METHODS: The relationship between plasma fatty acid (FA) levels and the severity of coronary artery disease (CAD), evaluated by Gensini score (GS), was investigated in CAD Tunisian patients compared to controls. Lipid profiles were analyzed, GS was calculated in CAD and non-CAD patients and compared to controls. RESULTS: CAD patients showed an alteration of conventional lipid parameters. In fact, a significant increase of plasmatic triglycerides (TG) level, atherogenic lipid ratios (TC/HDL-C,TG/HDL-C, LDL-C/HDL-C); and ApoB/ApoA1 was observed in the CAD group comparatively to controls (p < 0.001). Gensini score was showed to be a good indicator to evaluate cholesterol metabolism disorders associated with HDL-C since a negative association was found between HDL-C levels and GS for the two groups of patients. In addition, in the relation with FA and classes of FA, a negative association was found as expected, between Gensini score and total MUFA, PUFA n-3, total PUFA, GLA, DGLA and DHA. Furthermore, a positive association with stearic and erucic acid was found. Suggests that, GS is also a good indicator to evaluate FA metabolic disorders. Higher elongation index and modifications of desaturation index (D5D, D6D and D9D) were observed in patients compared to controls, supporting FA metabolism modifications. CONCLUSIONS: In conclusion, although that Tunisian population appears to follow the Mediterranean diet, variations of plasmatic FA levels and desaturase activities in CAD patients highlights an alteration of FA metabolism and suggests an important implication of certain FA in the development of atherosclerosis.

A hierarchical Bayesian approach for combining pharmacokinetic/pharmacodynamic modeling and Phase IIa trial design in orphan drugs: Treating adrenoleukodystrophy with Lorenzo's oil.

X-linked adrenoleukodystrophy (X-ALD) is a rare, progressive, and typically fatal neurodegenerative disease. Lorenzo's oil (LO) is one of the few X-ALD treatments available, but little has been done to establish its clinical efficacy or indications for its use. In this article, we analyze data on 116 male asymptomatic pediatric patients who were administered LO. We offer a hierarchical Bayesian statistical approach to understand LO pharmacokinetics (PK) and pharmacodynamics (PD) resulting from an accumulation of very long-chain fatty acids. We experiment with individual- and observational-level errors and various choices of prior distributions and deal with the limitation of having just one observation per administration of the drug, as opposed to the more usual multiple observations per administration. We link LO dose to the plasma erucic acid concentrations by PK modeling, and then link this concentration to a biomarker (C26, a very long-chain fatty acid) by PD modeling. Next, we design a Bayesian Phase IIa study to estimate precisely what improvements in the biomarker can arise from various LO doses while simultaneously modeling a binary toxicity endpoint. Our Bayesian adaptive algorithm emerges as reasonably robust and efficient while still retaining good classical (frequentist) operating characteristics. Future work looks toward using the results of this trial to design a Phase III study linking LO dose to actual improvements in health status, as measured by the appearance of brain lesions observed via magnetic resonance imaging.

A model-based approach to assess the exposure-response relationship of Lorenzo's oil in adrenoleukodystrophy.

AIMS: X-linked adrenoleukodystrophy (X-ALD) is a peroxisomal disorder, most commonly affecting boys, associated with increased very long chain fatty acids (C26:0) in all tissues, causing cerebral demyelination and adrenocortical insufficiency. Certain monounsaturated long chain fatty acids including oleic and erucic acids, known as Lorenzo's oil (LO), lower plasma C26:0 levels. The aims of this study were to characterize the effect of LO administration on plasma C26:0 concentrations and to determine whether there is an association between plasma concentrations of erucic acid or C26:0 and the likelihood of developing brain MRI abnormalities in asymptomatic boys. METHODS: Non-linear mixed effects modelling was performed on 2384 samples collected during an open label single arm trial. The subjects (n = 104) were administered LO daily at ~2-3 mg kg(-1) with a mean follow-up of 4.88 ± 2.76 years. The effect of erucic acid exposure on plasma C26:0 concentrations was characterized by an inhibitory fractional Emax model. A Weibull model was used to characterize the time-to-developing MRI abnormality. RESULTS: The population estimate for the fractional maximum reduction of C26:0 plasma concentrations was 0.76 (bootstrap 95% CI 0.73, 0.793). Our time-to-event analyses showed that every mg l(-1) increase in time-weighted average of erucic acid and C26:0 plasma concentrations was, respectively, associated with a 3.7% reduction and a 753% increase in the hazard of developing MRI abnormality. However, the results were not significant (P = 0.5344, 0.1509, respectively). CONCLUSIONS: LO administration significantly reduces the abnormally high plasma C26:0 concentrations in X-ALD patients. Further studies to evaluate the effect of LO on the likelihood of developing brain MRI abnormality are warranted.

Herring and Beef Meals Lead to Differences in Plasma 2-Aminoadipic Acid, ß-Alanine, 4-Hydroxyproline, Cetoleic Acid, and Docosahexaenoic Acid Concentrations in Overweight Men.

BACKGROUND: Dietary guidelines generally recommend increasing fish intake and reducing red meat intake for better long-term health. Few studies have compared the metabolic differences between eating meat and fish. OBJECTIVE: The objective of this study was to determine whether there are differences in the postprandial plasma metabolic response to meals containing baked beef, baked herring, and pickled herring. METHODS: Seventeen overweight men (BMI 25-30 kg/m(2), 41-67 y of age) were included in a randomized crossover intervention study. Subjects ate baked herring-, pickled herring-, and baked beef-based meals in a randomized order and postprandial blood plasma samples were taken over 7 h. Plasma metabolomics were measured with the use of gas chromatography-mass spectrometry and areas under the curve for detected metabolites were compared between meals. RESULTS: The plasma postprandial response of 2-aminoadipic acid, a suggested marker of diabetes risk, was 1.6 times higher after the beef meal than after the baked herring meal (P < 0.001). Plasma ß-alanine and 4-hydroxyproline both were markedly greater after beef intake than after herring intake (16 and 3.4 times the response of baked herring, respectively; P < 0.001). Herring intake led to a greater plasma postprandial response from docosahexaenoic acid (DHA) and cetoleic acid compared with beef (17.6 and 150 times greater, respectively; P < 0.001), whereas hippuric acid and benzoic acid were elevated after pickled herring compared with baked herring (5.4 and 43 times higher; P < 0.001). CONCLUSIONS: These results in overweight men confirm that DHA and cetoleic acid reflect herring intake, whereas ß-alanine and 4-hydroxyproline are potential biomarkers for beef intake. The greater postprandial rise in 2-aminoadipic acid after the beef meal, coupled to its proposed role in stimulating insulin secretion, may have importance in the context of red meat intake and increased diabetes risk. This trial was registered at clinicaltrials.gov as NCT02381613.

Fatty acid correlates of temperament in adolescent boys with attention deficit hyperactivity disorder.

Atypical fatty acid metabolism has been reported in attention deficit hyperactivity disorder (ADHD), however, its relationship with temperament in this population is unclear. The current study investigated the association between blood levels of fatty acids implicated in brain structure and function (omega-3, omega-6, omega-9) and personality traits of stability (neuroticism, conscientiousness and agreeableness) and plasticity (extraversion and openness). Twenty right-handed adolescent boys with ADHD completed a self-report NEO-FFI personality questionnaire, and had fatty acid content assessed from red blood using gas chromatography. Pearson's correlations showed no significant associations between omega-3 levels and personality. After correction for multiple comparisons, Adrenic Acid (C22:4n6) was inversely associated with stability. Oleic acid (C18:1n9) was positively associated with plasticity. Results are in line with a role of fatty acids in brain function. They suggest that those fatty acids that are involved in myelination (Adrenic, Oleic) have the strongest associations with temperament in adolescents with ADHD.

Exploratory study into the relation between plasma phospholipid fatty acid status and cognitive performance.

Previous research found a negative association between DHA status and selective attention in pregnant women. Goal of the present exploratory study is to investigate the potential relationship between essential fatty acid status and cognitive performance in a healthy non-pregnant population. Cognitive performance of 54 non-pregnant women was determined at baseline, 3, 15, and 22 weeks later with an objective neurocognitive test battery covering different brain domains. Plasma phospholipid fatty acid status was determined at baseline and at 22 weeks. The fatty acids of primary interest (arachidonic acid, adrenic acid, Osbond acid, eicosapentaenoic acid, and docosahexaenoic acid) were no significant predictors of cognitive performance at baseline or 22 weeks later. However, they attributed significantly (26.3%) to the amount of explained variance of the learning effect on the Stroop task, measuring general speed of information processing. Higher docosahexaenoic acid levels were associated with a slower learning curve. For arachidonic acid the opposite was found. In conclusion, this study provides a preliminary indication that a higher DHA status might be associated with slower learning curves. However, additional studies are necessary.

Uptake and metabolism of plasma-derived erucic acid by rat brain.

We examined the ability of erucic acid (22:1n-9) to cross the blood-brain barrier (BBB) by infusing [14-14C]22:1n-9 (170 microCi/kg, iv and icv) into awake, male rats. [1-14C]arachidonic acid (20:4n-6) [intravenous (i.v.)] was the positive control. After i.v. infusion, 0.011% of the plasma [14-14C]22:1n-9 was extracted by the brain, compared with 0.055% of the plasma [1-14C]20:4n-6. The [14-14C]22:1n-9 was extensively beta-oxidized (60%), compared with 30% for [1-14C]20:4n-6. Although 20:4n-6 was targeted primarily to phospholipid pools, 22:1n-9 was targeted to cholesteryl esters, triglycerides, and phospholipids. When [14-14C]22:1n-9 was infused directly into the fourth ventricle of the brain [intracerebroventricular (i.c.v.)] for 7 days, 60% of the tracer entered the phospholipid pools, similar to the distribution observed for [1-14C]20:4n-6. This demonstrates plasticity in the ability of the brain to esterify 22:1n-9 in an exposure-dependent manner. In i.v. and i.c.v. infused rats, a significant amount of tracer found in the phospholipid pools underwent sequential rounds of chain shortening and was found as [12-14C]20:1n-9 and [10-14C]oleic acid. These results demonstrate for the first time that intact 22:1n-9 crosses the BBB, is incorporated into specific lipid pools, and is chain-shortened.

Erucamide as a modulator of water balance: new function of a fatty acid amide.

The aim of this study was to isolate a compound from blood plasma that inhibits intestinal diarrhea and that appears also to regulate fluid volumes in other organs. The isolation procedure included lipid extraction, liquid chromatography, and gas chromatography. The active substance was identified by mass spectrometry as erucamide (MW 337 Da). The biological effect was reproduced with authentic erucamide. Erucamide is a fatty acid amide, such as oleamide and anandamide, which modulate other physiological functions in a receptor-mediated fashion. All the exact biological functions of erucamide are as yet to be defined, but it is already known to stimulate angiogenesis. Erucamide concentrations were determined in body organs from the pig. The blood plasma level was 3 ng/g, and those of lung, kidney, liver, and brain were 12, 2.5, 1.0, and 0.5 ng/g, respectively. Erucamide was below detection level in the intestine, but is known to be present in the cerebrospinal fluid. In the rat, 3H-erucamide was accumulated in vivo into lung, liver, and spleen and in vitro into lung, liver, brain, and intestine. The in vitro uptake was time and temperature dependent, but not saturable.

Effect of erucic acid on platelets in patients with adrenoleukodystrophy.

In a clinical trial for the management of adrenoleukodystrophy, we analyzed the effect of erucic acid (a component of Lorenzo's oil) on platelet number, fatty acid composition, and function. Analysis of variance was performed to compare platelet counts before starting treatment with Lorenzo's oil and at 6 and 12 months. We measured platelet fatty acid composition in subjects and control patients and correlated these values with their platelet counts using discriminant analysis. After 6 months, the mean platelet count decreased from 247,000/mm3 to 169,000/mm3 (+/- 1 standard deviation 58,000,n =39), P < 0.0001 compared to 18 subjects on a control diet having a mean baseline platelet count of 259,000/mm3 (+/- 1 standard deviation 67,000, n = 19) and at 6 months 267,000/mm3 (+/- 1 standard deviation 71,000). We found at P < 0.05 that the platelet counts showed a strong inverse relationship with erucic acid levels and other omega 9 fatty acids that form from the administration of the erucic acid component of Lorenzo's oil. Morphologic and platelet sizing measurements suggest that the physical properties of platelets may also be affected by erucic acid. Our studies show that the ingestion of erucic acid affects platelet biology. This indicates that platelet counts and properties are influenced by monounsaturated fatty acids, in addition to the well-known effects of polyunsaturated fatty acids. In areas of the world where erucic acid is widely ingested, the biology of platelets in these populations may be affected.

Relationship between dietary fatty acid, selenium, and degenerative cardiomyopathy.

With the finding of an increasing number of cases of degenerative cardiomyopathy (DCM) amongst patients in Chongqing, Sichuan, People's Republic of China, an attempt has been made to delineate possible etiological factors. In this province endemic for Keshan disease and with considerable consumption of oils high in erucic acid, the latter does not appear to be an operative noxious agent in DCM. Additionally, it does not appear to be caused by excessive oxygen radicals, low levels of antioxidants or low selenium levels. However, lower omega-3 fatty acid levels along with higher serum lipids may be the mechanism, via higher thromboxane levels, of the production of the myocardial degeneration seen in DCM.

Fatty acid composition of blood lipids in Chinese children consuming high erucic acid rapeseed oil.

The fatty acid patterns of oils, blood plasma and erythrocyte lipids from 28 children in the Shaanxi province of the People's Republic of China were determined by capillary gas liquid chromatography. The main source of fat in this region is rapeseed oil. The analysis of locally available rapeseed oil shows a high erucic acid content (mean 43.83%, range 33.91-50.48%). According to protocol data, about 3% of the daily nutrient energy is provided by erucic acid. Despite a low fat intake in Chinese children, the composition of the fatty acids of the fractions analyzed showed normal patterns. However, erucic acid was found in all fractions analyzed. Data on erucic acid in human tissue are scarce. Although there are no indications of erucic acid toxicity in man, it is known to cause cardiac lipidosis and necrosis in rats. The question remains open if erucic acid aggravates selenium deficiency symptoms which are known to be associated with Keshan disease, an endemic cardiomyopathy.

22-Carbon polyenoic acids. Incorporation into platelet phospholipids and the synthesis of these acids from 20-carbon polyenoic acid precursors by intact platelets.

The types of unsaturated fatty acids found in platelet phospholipids must be regulated by a series of controls which include specificity for activation and acylation as well as modification of circulating fatty acids by platelets prior to incubation into phospholipids. In this study we show that washed human platelets not only incorporate [1-14C]6,9,12-18:3, [1-14C]6,9,12,15-18:4, [1-14C]5,8,11-20:3, [1-14C]5,8,11,14-20:4, and [1-14C]5,8,11,14,17-20:5 into their phospholipids but also chain elongate each of these acids with subsequent acylation of the chain elongated products into phospholipids. Platelets incubated alone with 1-14C-labeled 5,8,11-20:3, 5,8,11,14-20:4, 5,8,11,14,17-20:5, 7,10,13,16,19-22:5, or 4,7,10,13,16,19-22:6 incorporated each of these acids into individual phosphoglycerides with phosphatidylinositol having the highest specific activity followed by phosphatidylcholine with phosphatidylserine approximately equal to phosphatidylethanolamine. The incorporation specificity of 4,7,10,13,16,19-22:6 was atypical since it was a relatively poor substrate for acylation into all phospholipids except phosphatidylethanolamine. The 20-carbon acids were better substrates for incorporation into phospholipids than were the 22-carbon compounds. Simultaneous incubation of 10 microM [1-14C]5,8,11,14-20:4 with increasing levels (5 to 15 microM) of each of the above five other 1-14C-labeled acids showed a concentration-dependent increase in the amount of the second fatty acid incorporated into platelet phospholipids. Dietary fat modification thus has the potential of increasing the plasma pool of 22-carbon acids for incorporation into platelets. In addition the activation of 20-carbon eicosanoid precursors by the high affinity platelet activating enzyme (Wilson, D. B., Prescott, S. M. and Majerus, P. W. (1982) J. Biol. Chem. 257, 3510-3515) will yield an acyl-CoA for both acylation and chain elongation followed by subsequent incorporation of 22-carbon acids into phosphoglycerides.

Metabolism of 7,10,13,16-docosatetraenoic acid to dihomo-thromboxane, 14-hydroxy-7,10,12-nonadecatrienoic acid and hydroxy fatty acids by human platelets.

Human platelets metabolize 7,10,13,16-docosatetraenoic acid (22:4(n - 6)) into dihomo-thromboxane B2 and 14-hydroxy-7,10,12-nonadecatrienoic acid at about twenty percent of the rate they convert arachidonic acid to thromboxane B2 and 12-hydroxy-5,8,10-heptadecatrienoic acid. 14-Hydroxy-7,10,12,16-docosatetraenoic was the major metabolite produce via the lipoxygenase pathway. Several other hydroxy acids were also produced in small amounts via an indomethacin-insensitive pathway. Incubation of 20 microM arachidonic acid with various levels of 22:4(n - 6) resulted in a dose-dependent inhibition of both thromboxane B2 and 12-hydroxy-5,8,10-heptadecatrienoic acid production. Conversely, 12-hydroxy-5,8,10,14-eicosatetraenoic acid synthesis was stimulated because of substrate shunting to the lipoxygenase pathway. These results show that 22:4(n - 6) may modify platelet function both by serving as a precursor for a 22-carbon thromboxane and by suppressing the synthesis of thromboxane A2 from arachidonic acid. In addition, our results suggest that simultaneous release of 22:4(n - 6) and arachidonic acid from platelet phospholipids will result in an elevation of both 12-hydroxy-5,8,10,14-eicosatetraenoic acid levels as well as simultaneous synthesis of 14-hydroxy-7,10,12,16-docosatetraenoic acid.

[Metabolism of fatty acids in the healthy myocardium and in ischemia]. / Metabolizm zhirnykh kislot v miokarde v norme i pri ishemii (obzor)

Data on absorption and consumption in heart tissue of free and ester-bound fatty acids from blood lipids are discussed. Preferential utilization of individual fatty acids in heart tissue from blood lipids is considered. Dependence of fatty acid metabolism on the activity of tricarboxylic acid cycle, interrelationship between metabolism of endogenous and exogenous fatty acids in heart muscle are considered. The data are analyzed on pathways of exogenous fatty acids turnover in tissues, their conversion into endogenous fatty acids, specific for individual tissue, cells and their organelles. Development of syndrome of unsaturated fatty acids deficiency, induced by incomplete fatty diet, is discussed. Metabolism of fatty acids in heart under conditions of oxygen deficiency is considered. The data are reviewed on the effects of hypoxia on metabolism of fatty acids in myocardium. Carbohydrate and fatty acid consumption in heart muscle, typical alterations in fatty acid incorporation into heart lipids, effect of fatty acids excess on functioning of sarcoplasmic reticulum and mitochondrial membranes are discussed.

Lipoxygenase activity of intact human platelets.

The oxygenation by lipoxygenase of different icosaenoic and docosaenoic acids by intact human platelets was studied. The HPLC analysis of the hydroxy compound (s) derived from icosaenoic acids showed that the 12-derivatives predominate. The increase of the fatty acid concentration markedly enhanced their oxygenation except for icosapentaenoic acid. The conversion of this acid into its hydroxy derivative rose in the presence of arachidonic acid, probably through both its cyclo-oxygenase and lipoxygenase product formation. Since 12-hydroxy-icosaenoic acids are modulators of PGH2-induced platelet aggregation, we conclude that the interactions between polyunsaturated fatty acids during their oxygenation by platelet lipoxygenase could be relevant to the regulating activity of dietary fatty acids.