Docosahexaenoic Acid (DHA) Supplementation Alters Phospholipid Species and Lipid Peroxidation Products in Adult Mouse Brain, Heart, and Plasma

Abstract

The abundance of docosahexaenoic acid (DHA) in phospholipids in the brain and retina has generated interest to search for its role in mediating neurological functions. Besides the source of many oxylipins with pro-resolving properties, DHA also undergoes peroxidation, producing 4-hydroxyhexenal (4-HHE), although its function remains elusive. Despite wide dietary consumption, whether supplementation of DHA may alter the peroxidation products and their relationship to phospholipid species in brain and other body organs have not been explored sufficiently. In this study, adult mice were administered a control or DHA-enriched diet for 3 weeks, and phospholipid species and peroxidation products were examined in brain, heart, and plasma. Results demonstrated that this dietary regimen increased (n-3) and decreased (n-6) species to different extent in all major phospholipid classes (PC, dPE, PE-pl, PI and PS) examined. Besides changes in phospholipid species, DHA-enriched diet also showed substantial increases in 4-HHE in brain, heart, and plasma. Among different brain regions, the hippocampus responded to the DHA-enriched diet showing significant increase in 4-HHE. Considering the pro- and anti-inflammatory pathways mediated by the (n-6) and (n-3) polyunsaturated fatty acids, unveiling the ability for DHA-enriched diet to alter phospholipid species and lipid peroxidation products in the brain and in different body organs may be an important step forward towards understanding the mechanism(s) for this (n-3) fatty acid on health and diseases.

Keywords: Docosahexaenoic acid; Heart; Hippocampus; Lipid peroxidation; Lipidomics; Plasma.

Figure 1.

Body weights of mice in the control and DHA group over the time of the study.

Figure 2.

Levels of 4-HHE and 4-HNE in mouse brain regions after feeding with a control or DHA-enriched (1%) diet for three weeks. Brain tissues including cerebral cortex, hippocampus, striatum, and cerebellum, were dissected and homogenized as described in text. Levels of 4-HHE and 4-HNE were determined by LC-MS/MS protocol as described in text. Data are normalized to tissue weight. Results represent the mean ± SEM of control (n = 5) and DHA (n = 7) samples. Analysis using a two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01.

Figure 3.

Levels of 4-HHE and 4-HNE in mouse (A) heart and (B) plasma after feeding with a control or DHA-enriched diet for three weeks. Procedures for processing the heart and plasma, and protocol for LC-MS/MS determination of 4-HHE and 4-HNE are described in text. Results represent the mean ± SEM of control (n = 5) and DHA (n = 7) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01; *** p < 0.001.

Figure 4.

Levels of phospholipids (diacyl-PE, PEpl, PC, PI, and PS) in mouse (A) cerebral cortex, (B) heart, and (C) plasma after feeding a control and DHA-enriched diet for three weeks. Lipids were extracted and analyzed by the shotgun lipidomics platform as described in text. Data are expressed as nmol/mg protein, and are mean ± SEM of control (n=4) and DHA (n=4) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01; *** p < 0.001. Abbreviations: dPE, diacyl-phosphatidylethanolamine, PEpl, alkenylacyl-phosphatidylethanolamine or ethanolamine plasmalogen, PC, phosphatidylcholine, PI, phosphatidylinositol, PS, phosphatidylserine.

Figure 5.

(A) Levels of (n-3) and (n-6) phospholipid species (dPE, PEpl, PC, PI and PS) in mouse cerebral cortex after feeding a control and DHA-enriched diet for three weeks. Lipids were extracted and analyzed by the shotgun lipidomics platform as described in text. In each phospholipid class, species with fatty acids containing (n-3) or (n-6) were grouped and expressed as nmol/mg protein. (B) Ratios of (n-6)/(n-3) phospholipid species of dPE, PEpl and PC from control and DHA group in the cortex using the data from (A). Data are mean ± SEM of control (n=4) and DHA (n=4) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01.

Figure 5.

(A) Levels of (n-3) and (n-6) phospholipid species (dPE, PEpl, PC, PI and PS) in mouse cerebral cortex after feeding a control and DHA-enriched diet for three weeks. Lipids were extracted and analyzed by the shotgun lipidomics platform as described in text. In each phospholipid class, species with fatty acids containing (n-3) or (n-6) were grouped and expressed as nmol/mg protein. (B) Ratios of (n-6)/(n-3) phospholipid species of dPE, PEpl and PC from control and DHA group in the cortex using the data from (A). Data are mean ± SEM of control (n=4) and DHA (n=4) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01.

Figure 6.

Levels of (n-3) and (n-6) phospholipid species (dPE, PC, PI and PS) in mouse heart after feeding a control and DHA-enriched diet for three weeks. In each phospholipid class, species with fatty acids containing (n-3) or (n-6) were grouped and expressed as nmol/mg protein. Data are mean ± SEM of control (n=4) and DHA (n=4) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01; *** p < 0.001.

Figure 7.

Levels of (n-3) and (n-6) phospholipid species (PC, PI, and PS) in mouse plasma after feeding a control and DHA-enriched diet for three weeks. In each phospholipid class, species with fatty acids containing (n-3) or (n-6) were grouped and expressed as nmol/mg protein. Data are mean ± SEM of control (n=4) and DHA (n=4) samples. Analysis using two-tail unpaired t-test indicated significance between DHA group and controls. * p < 0.05; ** p < 0.01; *** p < 0.001.