Eicosapentaenoic Acid Improves Endothelial Nitric Oxide Bioavailability Via Changes in Protein Expression During Inflammation.

BACKGROUND: Endothelial cell (EC) dysfunction involves reduced nitric oxide (NO) bioavailability due to NO synthase uncoupling linked to increased oxidation and reduced cofactor availability. Loss of endothelial function and NO bioavailability are associated with inflammation, including leukocyte activation. Eicosapentaenoic acid (EPA) administered as icosapent ethyl reduced cardiovascular events in REDUCE-IT (Reduction of Cardiovascular Events With Icosapent Ethyl-Intervention Trial) in relation to on-treatment EPA blood levels. The mechanisms of cardiovascular protection for EPA remain incompletely elucidated but likely involve direct effects on the endothelium. METHODS AND RESULTS: In this study, human ECs were treated with EPA and challenged with the cytokine IL-6 (interleukin-6). Proinflammatory responses in the ECs were confirmed by ELISA capture of sICAM-1 (soluble intercellular adhesion molecule-1) and TNF-α (tumor necrosis factor-α). Global protein expression was determined using liquid chromatography-mass spectrometry tandem mass tag. Release kinetics of NO and peroxynitrite were monitored using porphyrinic nanosensors. IL-6 challenge induced proinflammatory responses from the ECs as evidenced by increased release of sICAM-1 and TNF-α, which correlated with a loss of NO bioavailability. ECs pretreated with EPA modulated expression of 327 proteins by >1-fold (P<0.05), compared with IL-6 alone. EPA augmented expression of proteins involved in NO production, including heme oxygenase-1 and dimethylarginine dimethylaminohydrolase-1, and 34 proteins annotated as associated with neutrophil degranulation. EPA reversed the endothelial NO synthase uncoupling induced by IL-6 as evidenced by an increased [NO]/[peroxynitrite] release ratio (P<0.05). CONCLUSIONS: These direct actions of EPA on EC functions during inflammation may contribute to its distinct cardiovascular benefits.

Do patients benefit from omega-3 fatty acids?

Omega-3 fatty acids (O3FAs) possess beneficial properties for cardiovascular (CV) health and elevated O3FA levels are associated with lower incident risk for CV disease (CVD.) Yet, treatment of at-risk patients with various O3FA formulations has produced disparate results in large, well-controlled and well-conducted clinical trials. Prescription formulations and fish oil supplements containing low-dose mixtures of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have routinely failed to prevent CV events in primary and secondary prevention settings when added to contemporary care, as shown most recently in the STRENGTH and OMEMI trials. However, as observed in JELIS, REDUCE-IT, and RESPECT-EPA, EPA-only formulations significantly reduce CVD events in high-risk patients. The CV mechanism of action of EPA, while certainly multifaceted, does not depend solely on reductions of circulating lipids, including triglycerides (TG) and LDL, and event reduction appears related to achieved EPA levels suggesting that the particular chemical and biological properties of EPA, as compared to DHA and other O3FAs, may contribute to its distinct clinical efficacy. In vitro and in vivo studies have shown different effects of EPA compared with DHA alone or EPA/DHA combination treatments, on atherosclerotic plaque morphology, LDL and membrane oxidation, cholesterol distribution, membrane lipid dynamics, glucose homeostasis, endothelial function, and downstream lipid metabolite function. These findings indicate that prescription-grade, EPA-only formulations provide greater benefit than other O3FAs formulations tested. This review summarizes the clinical findings associated with various O3FA formulations, their efficacy in treating CV disease, and their underlying mechanisms of action.

Eicosapentaenoic acid (EPA) reduces pulmonary endothelial dysfunction and inflammation due to changes in protein expression during exposure to particulate matter air pollution.

AIMS: Inhalation of air pollution small particle matter (PM) is a leading cause of cardiovascular (CV) disease. Exposure to PMs causes endothelial cell (EC) dysfunction as evidenced by nitric oxide (NO) synthase uncoupling, vasoconstriction and inflammation. Eicosapentaenoic acid (EPA) has been shown to mitigate PM-induced adverse cardiac changes in patients receiving omega-3 fatty acid supplementation. We set out to determine the pro-inflammatory effects of multiple PMs (urban and fine) on pulmonary EC NO bioavailability and protein expression, and whether EPA restores EC function under these conditions. METHODS AND RESULTS: We pretreated pulmonary ECs with EPA and then exposed them to urban or fine air pollution PMs. LC/MS-based proteomic analysis to assess relative expression levels. Expression of adhesion molecules was measured by immunochemistry. The ratio of NO to peroxynitrite (ONOO-) release, an indication of eNOS coupling, was measured using porphyrinic nanosensors following calcium stimulation. Urban/fine PMs also modulated 9/12 and 13/36 proteins, respectively, linked to platelet and neutrophil degranulation pathways and caused > 50% (p < 0.001) decrease in the stimulated NO/ONOO- release ratio. EPA treatment altered expression of proteins involved in these inflammatory pathways, including a decrease in peroxiredoxin-5 and an increase in superoxide dismutase-1. EPA also increased expression of heme oxygenase-1 (HMOX1), a cytoprotective protein, by 2.1-fold (p = 0.024). EPA reduced elevations in sICAM-1 levels by 22% (p < 0.01) and improved the NO/ONOO- release ratio by > 35% (p < 0.05). CONCLUSION: These cellular changes may contribute to anti-inflammatory, cytoprotective and lipid changes associated with EPA treatment during air pollution exposure.

Omega-3-fatty acids: Do they prevent cardiovascular disease?

Despite cardiovascular disease (CVD) reductions with high-intensity statins, there remains residual risk among patients with metabolic disorders. Alongside low-density lipoproteins (LDL-C), elevated triglycerides (TG) are associated with incident CVD events. Omega-3 fatty acids (n3-FAs), specifically eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), lower TG levels, but their ability to reduce CV risk has been highly inconsistent. Trials using icosapent ethyl (IPE), a purified EPA ethyl ester, produced reductions in CVD events and atherosclerotic plaque regression compared with mixed EPA/DHA formulations despite similar TG-reductions. The separate effects of EPA and DHA on tissue distribution, oxidative stress, inflammation, membrane structure and endothelial function may contribute to these discordant outcomes. Additional mechanistic trials will provide further insights into the role of n3-FAs in reducing CVD risk beyond TG lowering.

Role of Omega-3 Fatty Acids in Cardiovascular Disease: the Debate Continues.

PURPOSE OF REVIEW: The omega-3 fatty acids (n3-FAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have recently undergone testing for their ability to reduce residual cardiovascular (CV) risk among statin-treated subjects. The outcome trials have yielded highly inconsistent results, perhaps attributable to variations in dosage, formulation, and composition. In particular, CV trials using icosapent ethyl (IPE), a highly purified ethyl ester of EPA, reproducibly reduced CV events and progression of atherosclerosis compared with mixed EPA/DHA treatments. This review summarizes the mechanistic evidence for differences among n3-FAs on the development and manifestations of atherothrombotic disease. RECENT FINDINGS: Large randomized clinical trials with n3-FAs have produced discordant outcomes despite similar patient profiles, doses, and triglyceride (TG)-lowering effects. A large, randomized trial with IPE, a prescription EPA only formulation, showed robust reduction in CV events in statin treated patients in a manner proportional to achieved blood EPA concentrations. Multiple trials using mixed EPA/DHA formulations have not shown such benefits, despite similar TG lowering. These inconsistencies have inspired investigations into mechanistic differences among n3-FAs, as EPA and DHA have distinct membrane interactions, metabolic products, effects on cholesterol efflux, antioxidant properties, and tissue distribution. EPA maintains normal membrane cholesterol distribution, enhances endothelial function, and in combination with statins improves features implicated in plaque stability and reduces lipid content of plaques. Insights into reductions in residual CV risk have emerged from clinical trials using different formulations of n3-FAs. Among high-risk patients on contemporary care, mixed n3-FA formulations showed no reduction in CV events. The distinct benefits of IPE in multiple trials may arise from pleiotropic actions that correlate with on-treatment EPA levels beyond TG-lowering. These effects include altered platelet function, inflammation, cholesterol distribution, and endothelial dysfunction. Elucidating such mechanisms of vascular protection for EPA may lead to new interventions for atherosclerosis, a disease that continues to expand worldwide.

Multi-scale analysis of habitat fragmentation on small-mammal abundance and tick-borne pathogen infection prevalence in Essex County, MA.

Habitat fragmentation and heterogeneity transform otherwise contiguous tracks of forest into smaller patches in the northeastern U.S. and likely impact abundances, movement patterns, and disease transmission pathways for small-mammal communities at multiple scales. We sought to determine the structure of a small-mammal community in terms of mammal abundance and infection prevalence of Borrelia burgdorferi sensu stricto (s.s.), Anaplasma phagocytophilum, and Babesia microti within a fragmented landscape in Essex County, Massachusetts, USA. We studied communities at multiple spatial scales, including vegetation, edge type, and landscape (including 200-m, 500-m, and 1000-m radii) scales. A total of 16 study sites were chosen to represent four edge types: interior forest, pasture edge, natural edge, and residential edge. At each site, we trapped small mammals and conducted vegetation surveys and GIS analysis. Upon capture, a tissue sample was collected to analyze for presence of pathogens. Northern short-tailed shrew (Blarina brevicauda) abundance did not differ based on edge type, whereas abundance of the white-footed mouse (Peromyscus leucopus) was greatest at pasture edges, although the relationship was relatively weak. White-footed mouse abundance was negatively associated with amount of forested area within a 500-m radius, whereas northern short-tailed shrew abundance demonstrated a positive relationship with fragmentation indices at the 200-m radius. White-footed mice captured at interior-forest habitat were more likely be infected with B. burgdorferi (s.s.) than individuals from edge habitat. Greater prevalence of B. burgdorferi infection of white-footed mice in forest interiors compared to edge habitats counters previous studies. Reasons for this and implications are discussed.

A biological rationale for the disparate effects of omega-3 fatty acids on cardiovascular disease outcomes.

The omega-3 fatty acids (n3-FAs) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) rapidly incorporate into cell membranes where they modulate signal transduction pathways, lipid raft formation, and cholesterol distribution. Membrane n3-FAs also form specialized pro-resolving mediators and other intracellular oxylipins that modulate inflammatory pathways, including T-cell differentiation and gene expression. Cardiovascular (CV) trials have shown that EPA, administered as icosapent ethyl (IPE), reduces composite CV events, along with plaque volume, in statin-treated, high-risk patients. Mixed EPA/DHA regimens have not shown these benefits, perhaps as the result of differences in formulation, dosage, or potential counter-regulatory actions of DHA. Indeed, EPA and DHA have distinct, tissue-specific effects on membrane structural organization and cell function. This review summarizes: (1) results of clinical outcome and imaging trials using n3-FA formulations; (2) membrane interactions of n3-FAs; (3) effects of n3-FAs on membrane oxidative stress and cholesterol crystalline domain formation during hyperglycemia; (4) n3-FA effects on endothelial function; (5) role of n3-FA-generated metabolites in inflammation; and (6) ongoing and future clinical investigations exploring treatment targets for n3-FAs, including COVID-19.

EPA and DHA containing phospholipids have contrasting effects on membrane structure.

Omega-3 FAs EPA and DHA influence membrane fluidity, lipid rafts, and signal transduction. A clinical trial, Reduction of Cardiovascular Events with Icosapent Ethyl-Intervention Trial, demonstrated that high-dose EPA (4 g/d icosapent ethyl) reduced composite cardiovascular events in statin-treated high-risk patients. EPA benefits correlated with on-treatment levels, but similar trials using DHA-containing formulations did not show event reduction. We hypothesized that differences in clinical efficacy of various omega-3 FA preparations could result from differential effects on membrane structure. To test this, we used small-angle X-ray diffraction to compare 1-palmitoyl-2-eicosapentaenoyl-sn-glycero-3-phosphocholine (PL-EPA), 1-palmitoyl-2-docosahexaenoyl-sn-glycero-3-phosphocholine (PL-DHA), and 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PL-AA) in membranes with and without 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol. Electron density profiles (electrons/Å3 vs. Å) were used to determine membrane structure, including membrane width (d-space). PL-EPA and PL-DHA had similar membrane structures without POPC and/or cholesterol but had contrasting effects in the presence of POPC and cholesterol. PL-EPA increased membrane hydrocarbon core electron density over an area of ±0-10 Å from the center, indicating an extended orientation. PL-DHA increased electron density in the phospholipid head group region, concomitant with disordering in the hydrocarbon core and a similar d-space (58 Å). Adding equimolar amounts of PL-EPA and PL-DHA produced changes that were attenuated compared with their separate effects. PL-AA increased electron density centered ±12 Å from the membrane center. The contrasting effects of PL-EPA, PL-DHA, and PL-AA on membrane structure may contribute to differences observed in the biological activities and clinical actions of various omega-3 FAs.

Omega-3 and omega-6 fatty acids have distinct effects on endothelial fatty acid content and nitric oxide bioavailability.

Treatment with high dose icosapent ethyl (IPE), an ethyl ester of the omega-3 fatty acid eicosapentaenoic acid (EPA), significantly reduced ischemic events in patients with either cardiovascular disease (CV) or diabetes plus other risk factors (REDUCE-IT) but the mechanism is not well understood.  We compared the effects of EPA, docosahexaenoic acid (DHA), and the omega-6 fatty acid arachidonic acid (AA) on bioavailability of nitric oxide (NO) and fatty acid composition. Human umbilical vein endothelial cells (HUVECs) were pretreated with EPA, DHA, or AA (10 µM). Cells were stimulated with calcium ionophore and NO and peroxynitrite (ONOO-) were measured using porphyrinic nanosensors. Levels of EPA, DHA, AA and other fatty acids were measured by gas chromatography (GC). EPA treatment caused the greatest NO release (18%, p < 0.001) and reduction in ONOO- (13%, p < 0.05) compared to control; the [NO]/[ ONOO-] ratio increased by 35% (p < 0.001). DHA treatment increased NO levels by 12% (p < 0.01) but had no effect on ONOO- release. AA did not affect either NO or ONOO- release.  Fatty acid treatments increased their respective levels in endothelial cells.  EPA levels increased 10-fold to 4.59 mg/g protein (p < 0.001) with EPA treatment and the EPA/AA ratio increased by 10-fold (p < 0.001) compared to vehicle.  Only EPA increased docosapentaenoic acid (DPA, omega-3) levels by 2-fold (p < 0.001). AA alone decreased the EPA/AA ratio 4-fold (p<0.001). These findings support a preferential benefit of EPA on endothelial function and omega-3 fatty acid content.

Kinetics of generic tacrolimus in heart transplantation: A cautionary note.

Tacrolimus is a core component of immunosuppressive regimens. This study compared active pharmaceutical ingredient (API) and dissolution kinetics of branded tacrolimus and formulations from three generic manufacturers (Mylan, Dr. Reddy's, Intas) including samples from patients who suffered acute cardiac allograft rejection. Generic samples showed similar API content compared to branded samples with no major impurities. Capsules that underwent uniformity testing had consistent capsule-to-capsule API. Dissolution testing showed similar profiles between branded tacrolimus and Mylan, but notable differences with Dr. Reddy's and Intas. The approximate maximal inhibitory concentration (IC50) was highest in branded tacrolimus (29 minutes), followed by Mylan (26 minutes), Dr. Reddy's (19 minutes), and Intas (14 minutes) (Student-Newman-Keuls Multiple Comparisons Test; overall ANOVA: p = 0.0199, F = 6.469). This study suggests that the bioavailability of certain generic tacrolimus formulations peak significantly earlier than branded tacrolimus. Further study is needed to determine whether these differences are clinically relevant.

Eicosapentaenoic acid (EPA) has optimal chain length and degree of unsaturation to inhibit oxidation of small dense LDL and membrane cholesterol domains as compared to related fatty acids in vitro.

BACKGROUND: Oxidation of small dense low-density lipoprotein (sdLDL) and membranes is causally related to atherosclerosis. The omega-3 fatty acid (FA) eicosapentaenoic acid (EPA, 20:5, ω-3) significantly reduced oxidized LDL in patients with hypertriglyceridemia by unknown mechanisms. We compared EPA effects to related FAs of varying chain length and unsaturation on oxidation of sdLDL and model membranes, and on cholesterol crystal domains. We compared EPA to the FAs: stearic (SA, 18:0), oleic (OA, 18:1, ω-9), linoleic (LA, 18:2, ω-6), alpha-linolenic (ALA, 18:3, ω-3), eicosanoic (EA, 20:0), eicosatrienoic (ETE, 20:3, ω-3), arachidonic (AA, 20:4, ω-6), docosapentaenoic (DPA, 22:5, ω-3), and docosahexaenoic (DHA, 22:6, ω-3). METHODS: Human sdLDL or model membranes of cholesterol and 1,2-Dilinoleoyl-sn-glycero-3-phosphocholine [18:2(cis)PC or DLPC] were preincubated with FAs followed by copper-induced oxidation. Malondialdehyde (MDA) or lipid hydroperoxides (LOOH) levels measured oxidation; small-angle X-ray diffraction assessed cholesterol domain formation. RESULTS: After 40 min, EPA reduced MDA levels 70% compared to vehicle (p < 0.001). Lesser inhibition was observed with DHA, DPA, ETE, and ALA (33%, 34%, 32%, and 16%, respectively; all p < 0.001 versus vehicle). Similar relative FA effects were observed in model membranes where EPA more substantially inhibited cholesterol crystal domain formation. CONCLUSION: We observed relationships between hydrocarbon length and unsaturation with antioxidant activity and membrane cholesterol domain formation. EPA had the most favorable molecular structure, likely contributing to membrane stability, improved lipoprotein clearance, and reduced inflammation. GENERAL SIGNIFICANCE: Insight is provided into FA hydrocarbon length and unsaturation relationships with antioxidant activity in lipoproteins and membranes, and cholesterol crystal domains formation.

Are dietary fish oil supplements appropriate for dyslipidemia management? A review of the evidence.

PURPOSE OF REVIEW: The purpose of this review is to assess whether dietary fish oil supplements can be appropriate for patients with elevated triglycerides and cardiovascular risk based on a comprehensive analysis of their composition, and level of regulatory oversight. RECENT FINDINGS: Approximately 19 million people in the United States take fish oil supplements, many for the purpose of treating or preventing heart disease. Unlike prescription products, fish oil supplements are classified as food by the Food and Drug Administration (FDA) and are not required to undergo manufacturing oversight or clinical testing. Analysis of widely used dietary fish oil supplements show that they may have lower amounts of ω-3 than advertised as well as significant levels of saturated fat and oxidized oils which actually may contribute to dyslipidemia. Clinical outcome trials have failed to show a consistent cardiovascular benefit with fish oil supplements and other low-dose mixed ω-3 fatty acids. SUMMARY: In light of limited regulatory oversight and evidence of quality concerns, dietary fish oil supplements are not an appropriate substitute for FDA approved prescription ω-3 fatty acids for their indicated use in treatment of elevated triglycerides or the prevention of cardiovascular events.

Rosmarinic acid and its esters inhibit membrane cholesterol domain formation through an antioxidant mechanism based, in nonlinear fashion, on alkyl chain length.

BACKGROUND: Under conditions of oxidative stress, cholesterol aggregates into discrete membrane bilayer domains that precipitate the formation of extracellular crystals, a feature of advanced atheroma in cardiovascular disease. Therapeutic interventions using membrane-directed antioxidants, such as polyphenolic esters, may reduce cholesterol domains and crystal formation. In this study, the effects of rosmarinic acid (RC0) and rosmarinic esters, with alkyl chain lengths ranging from 4 to 16­carbons (RC4-RC16), on membrane lipid oxidation and cholesterol domain formation were investigated. METHODS: Model membranes were prepared with 1,2-dilinoleoyl-sn-glycero-3-phosphocholine and cholesterol at different cholesterol-to-phospholipid mole ratios (0.3:1, 0.9:1, and 1.2:1), in the absence or presence of each molecule and exposed to 72 h of oxidation. Changes in lipid hydroperoxide (LOOH) and cholesterol domain formation were measured using iodometric and small angle x-ray diffraction approaches, respectively. RESULTS: Rosmarinic acid and its esters had differential effects on LOOH formation based on alkyl chain length. RC8 exhibited the greatest antioxidant effect, reducing LOOH levels by 82%, and inhibited cholesterol domain formation. By contrast, RC0 and RC16 failed to inhibit either LOOH formation or cholesterol domain formation. CONCLUSION: These data indicate that the membrane antioxidant and cholesterol domain inhibition activities of rosmarinic acid esters are dependent, nonlinearly, on alkyl chain length. The mechanism for this effect is attributed to the influence of alkyl chain length on the optimal depth of the polyphenols into the lipid bilayer for trapping free radicals. GENERAL SIGNIFICANCE: These findings provide insight into novel atheroprotective benefits of polyphenol esters that are dependent on their membrane location.

Eicosapentaenoic acid improves endothelial function and nitric oxide bioavailability in a manner that is enhanced in combination with a statin.

The endothelium exerts many vasoprotective effects that are largely mediated by release of nitric oxide (NO). Endothelial dysfunction represents an early but reversible step in atherosclerosis and is characterized by a reduction in the bioavailability of NO. Previous studies have shown that eicosapentaenoic acid (EPA), an omega-3 fatty acid (O3FA), and statins individually improve endothelial cell function, but their effects in combination have not been tested. Through a series of in vitro experiments, this study evaluated the effects of a combined treatment of EPA and the active metabolite of atorvastatin (ATM) on endothelial cell function under conditions of oxidative stress. Specifically, the comparative and time-dependent effects of these agents on endothelial dysfunction were examined by measuring the levels of NO and peroxynitrite (ONOO-) released from human umbilical vein endothelial cells (HUVECs). The data suggest that combined treatment with EPA and ATM is beneficial to endothelial function and was unique to EPA and ATM since similar improvements could not be recapitulated by substituting another O3FA docosahexaenoic acid (DHA) or other TG-lowering agents such as fenofibrate, niacin, or gemfibrozil. Comparable beneficial effects were observed when HUVECs were pretreated with EPA and ATM before exposure to oxidative stress. Interestingly, the kinetics of EPA-based protection of endothelial function in response to oxidation were found to be significantly different than those of DHA. Lastly, the beneficial effects on endothelial function generated by combined treatment of EPA and ATM were reproduced when this study was expanded to an ex vivo model utilizing rat glomerular endothelial cells. Taken together, these findings suggest that a combined treatment of EPA and ATM can inhibit endothelial dysfunction that occurs in response to conditions such as hyperglycemia, oxidative stress, and dyslipidemia.

Eicosapentaenoic acid and docosahexaenoic acid have distinct membrane locations and lipid interactions as determined by X-ray diffraction.

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) differentially influence lipid oxidation, signal transduction, fluidity, and cholesterol domain formation, potentially due in part to distinct membrane interactions. We used small angle X-ray diffraction to evaluate the EPA and DHA effects on membrane structure. Membrane vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and cholesterol (C) (0.3C:POPC mole ratio) were prepared and treated with vehicle, EPA, or DHA (1:10 mol ratio to POPC). Electron density profiles generated from the diffraction data showed that EPA increased membrane hydrocarbon core electron density over a broad area, up to ±â€¯20 Šfrom the membrane center, indicating an energetically favorable extended orientation for EPA likely stabilized by van der Waals interactions. By contrast, DHA increased electron density in the phospholipid head group region starting at ±â€¯12 Šfrom the membrane center, presumably due to DHA-surface interactions, with coincident reduction in electron density in the membrane hydrocarbon core centered ±â€¯7-9 Šfrom the membrane center. The membrane width (d-space) decreased by 5 Šin the presence of vehicle as the temperature increased from 10 °C to 30 °C due to increased acyl chain trans-gauche isomerizations, which was unaffected by addition of EPA or DHA. The influence of DHA on membrane structure was modulated by temperature changes while the interactions of EPA were unaffected. The contrasting EPA and DHA effects on membrane structure indicate distinct molecular locations and orientations that may contribute to observed differences in biological activity.

Eicosapentaenoic acid inhibits oxidation of high density lipoprotein particles in a manner distinct from docosahexaenoic acid.

The omega-3 fatty acid eicosapentaenoic acid (EPA) reduces oxidation of ApoB-containing particles in vitro and in patients with hypertriglyceridemia. EPA may produce these effects through a potent antioxidant mechanism, which may facilitate LDL clearance and slow plaque progression. We hypothesize that EPA antioxidant effects may extend to ApoA-containing particles like HDL, potentially preserving certain atheroprotective functions. HDL was isolated from human plasma and incubated at 37 °C in the absence (vehicle) or presence of EPA and/or DHA; 5.0 or 10.0 µM each. Samples were then subjected to copper-induced oxidation (10 µM). HDL oxidation was inhibited similarly by EPA and DHA up to 1 h. EPA (10 µM) maintained significant HDL oxidation inhibition of 89% (0.622 ±â€¯0.066 µM MDA; p < .001) at 4 h, with continued inhibition of 64% at 14 h, vs. vehicle (5.65 ±â€¯0.06 to 2.01 ±â€¯0.10 µM MDA; p < .001). Conversely, DHA (10 µM) antioxidant benefit was lost by 4 h. At a lower concentration (5 µM), EPA antioxidant activity remained at 81% (5.53 ±â€¯0.15 to 1.03 ±â€¯0.10 µM MDA; p < .001) at 6 h, while DHA lost all antioxidant activity by 4 h. The antioxidant activity of EPA was preserved when combined with an equimolar concentration of DHA (5 µM each). EPA pretreatment prevented HDL oxidation in a dose-dependent manner that was preserved over time. These results suggest unique lipophilic and electron stabilization properties for EPA as compared to DHA with respect to inhibition of HDL oxidation. These antioxidant effects of EPA may enhance certain atheroprotective functions for HDL.