Supplementation with omega-3 or omega-6 fatty acids attenuates platelet reactivity in postmenopausal women

Abstract

Postmenopausal women are at increased risk for a cardiovascular event due to platelet hyperactivity. There is evidence suggesting that ω-3 polyunsaturated fatty acids (PUFAs) and ω-6 PUFAs have cardioprotective effects in these women. However, a mechanistic understanding of how these fatty acids regulate platelet function is unknown. In this study, we supplemented postmenopausal women with fish oil (ω-3 fatty acids) or evening primrose oil (ω-6 fatty acids) and investigated the effects on their platelet activity. The effects of fatty acid supplementation on platelet aggregation, dense granule secretion, and activation of integrin αIIbβ3 at basal levels and in response to agonist were tested in postmenopausal women following a supplementation and washout period. Supplementation with fish oil or primrose oil attenuated the thrombin receptor PAR4-induced platelet aggregation. Supplementation with ω-3 or ω-6 fatty acids decreased platelet dense granule secretion and attenuated basal levels of integrin αIIbβ3 activation. Interestingly, after the washout period following supplementation with primrose oil, platelet aggregation was similarly attenuated. Additionally, for either treatment, the observed protective effects post-supplementation on platelet dense granule secretion and basal levels of integrin activation were sustained after the washout period, suggesting a long-term shift in platelet reactivity due to fatty acid supplementation. These findings begin to elucidate the underlying mechanistic effects of ω-3 and ω-6 fatty acids on platelet reactivity in postmenopausal women. Hence, this study supports the beneficial effects of fish oil or primrose oil supplementation as a therapeutic intervention to reduce the risk of thrombotic events in postmenopausal women. https://ichgcp.net/clinical-trials-registry/NCT02629497.

Conflict of interest statement

Dr. Holinstat is a consultant and equity holder for Veralox therapeutics and a consultant for Cereno Scientific. All other authors declared no competing interests for this work.

© 2022 The Authors. Clinical and Translational Science published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.

Figures

FIGURE 1

Fish oil or evening primrose oil supplementation enhances ω‐3 and ω‐6 fatty acid content in platelets. (a) Schematic figure of major fatty acids produced by the precursor ω‐3 α‐linolenic fatty acid (ALA) and the precursor ω‐6 linoleic fatty acid (LA). (b) Platelets from subjects supplemented with fish oil and (c) platelets from subjects supplemented with evening primrose oil were measured at pre‐supplementation (baseline), after 60 days of supplementation (fish oil or primrose oil) and after 14‐day washout period (washout) for docosapentaenoic acid ω‐3 (DPAω‐3), docosahexaenoic acid (DHA), γ‐linolenic acid (GLA), and dihomo‐γ‐linolenic acid (DGLA). Numbers are given in parenthesis. The data represent mean ± SD. A mixed‐effects analysis with Dunnett's multiple comparison post‐hoc test was performed. Asterisks indicate statistically significant differences between groups (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).

FIGURE 2

Fish oil supplementation in postmenopausal women attenuates PAR4‐induced platelet aggregation. Platelet aggregation was tested at pre‐supplementation (baseline), after 60 days of supplementation (fish oil), and after a 14‐day washout period (washout) in the presence of a variety of agonists (a) PAR1‐AP, (b) PAR4‐AP, (c) thrombin, or (d) collagen. Numbers are given in parenthesis. The data represent mean ± SEM. A mixed‐effects analysis with Dunnett's multiple comparison post‐hoc test was performed. Asterisks indicate statistically significant differences between groups (*p < 0.05).

FIGURE 3

Primrose oil supplementation in postmenopausal women attenuates PAR4‐induced platelet aggregation. Platelet aggregation was tested at pre‐supplementation (baseline), after 60 days of supplementation (primrose oil), and after a 14‐day washout period (washout) in the presence of a variety of agonists (a) PAR1‐AP, (b) PAR4‐AP, (c) thrombin, or (d) collagen. Numbers are given in parenthesis. The data represent mean ± SEM. A mixed‐effects analysis with Dunnett's multiple comparison post‐hoc test was performed. Asterisks indicate statistically significant differences between groups (*p < 0.05, **p < 0.01).

FIGURE 4

Basal activity of integrin αIIbβ3 is lowered following supplementation and washout, but attenuation in agonis‐induced αIIbβ3 activation is only observed post‐washout. Basal activation of αIIbβ3 was measured before supplementation (baseline), after 60 days of supplementation with either (a) fish oil (post fish oil supplementation) or (b) evening primrose oil (post primrose oil supplementation) and (c, d) after a 14‐day washout period (post fish oil washout AND post primrose oil washout). Agonist‐induced αIIbβ3 activation was assessed at same timepoints for (e) fish oil and (f) primrose oil supplementation. Numbers are given in parenthesis. The data represent mean ± SEM. An unpaired two‐tailed t‐test was performed to determine significance. Asterisks indicate statistically significant differences between groups (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001). MFI refers to mean fluorescence intensity.

FIGURE 5

Postmenopausal women exhibit attenuated dense granule secretion when supplemented with fish oil. ATP secretion were assessed at pre‐supplementation (baseline), post‐supplementation (fish oil), and after a 14‐day washout period (washout) in the presence of (a) PAR1‐AP, (b) PAR4‐AP, (c) thrombin, or (d) collagen. Numbers are given in parenthesis. The data represent mean ± SEM. A mixed‐effects analysis with Dunnett's multiple comparison post hoc test was performed. Asterisks indicate statistically significant differences between groups (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).

FIGURE 6

Postmenopausal women exhibit attenuated dense granule secretion when supplemented with evening primrose oil. ATP secretion were assessed at pre‐supplementation (baseline), post‐supplementation (primrose oil), and after a 14‐day washout period (washout) in the presence of (a) PAR1‐AP, (b) PAR4‐AP, (c) thrombin, or (d) collagen. Numbers are given in parenthesis. The data represent mean ± SEM. A mixed‐effects analysis with Dunnett's multiple comparison post hoc test was performed. Asterisks indicate statistically significant differences between groups (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001).