Effect of omega-3 fatty acid ethyl esters on the oxylipin composition of lipoproteins in hypertriglyceridemic, statin-treated subjects

Abstract

Background: Oxylipins mediate inflammation, vascular tension, and more. Their presence in lipoproteins could explain why lipoproteins mediate nearly identical activities.

Methods: To determine how oxylipins are distributed in the lipoproteins of hypertriglyceridemic subjects, and whether omega-3 fatty acids alter them in a manner consistent with improved cardiovascular health, we recruited 15 dyslipidemic subjects whose levels of low density lipoprotein cholesterol (LDL-C) were at goal but who remained hypertriglyceridemic (200-499 mg/dL). They were treated them with the indicated dose of 4 g/d omega-3 acid ethyl esters (P-OM3) for 8 weeks. Measured oxylipins included mid-chain alcohols (HETEs, HEPEs and HDoHEs), ketones (KETEs), epoxides (as EpETrEs, EpETEs, and EpDPEs).

Results: At baseline, arachidonate-oxylipins (HETEs, KETEs, and EpETrEs) were most abundant in plasma with the greatest fraction of total abundance (mean |95% CI|) being carried in high density lipoproteins (HDL); 42% |31, 57| followed by very low density lipoproteins (VLDL); 27% |20, 36|; and LDL 21% |16, 28|. EPA- and DHA-derived oxylipins constituted less than 11% of total. HDL carried alcohols and epoxides but VLDL was also rich in ketones. Treatment decreased AA-derived oxylipins across lipoprotein classes (-23% |-33, -12|, p = 0.0003), and expanded EPA-(322% |241, 422|, p<0.0001) and DHA-derived oxylipins (123% |80, 176|, p<0.0001).

Conclusions: Each lipoprotein class carries a unique oxylipin complement. P-OM3 treatment alters the oxylipin content of all classes, reducing pro-inflammatory and increasing anti-inflammatory species, consistent with the improved inflammatory and vascular status associated with the treatment.

Trial registration: ClinicalTrials.gov NCT00959842.

Conflict of interest statement

Competing Interests: JWN, TLP and VRB have nothing to disclose. This was an investigator-initiated study funded by GlaxoSmithKline. GlaxoSmithKline had no role in analysis or interpretation of the study results. GCS and WSH received speakership honoraria from GlaxoSmithKline while the trial was ongoing. This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials; however, in order to protect participant privacy, data are not being made publicly available.

Figures

Figure 1. Distribution of oxylipins in plasma compartments.

The distributions of oxylipins are expressed as a percent of the total baseline abundance before and after treatment. A: Oxylipins in plasma by parent fatty acid and chemistry, regardless of distribution in HDL, LDL, VLDL, or ‘free’. A mixed-model ANOVA was used for each FA. B: Oxylipins in lipoprotein by parent fatty acid, regardless of chemistry. A mixed-model ANOVA was used for each FA pool. n = 14: LDL was not recovered for one subject, making total calculations impossible. *p<0.05, **p<0.0005 versus baseline. Letters (A,B,C) indicate results of Tukey's HSD test for differences between lipoproteins. Those sharing a letter are not different.

Figure 2. Distribution of fatty acid alcohols and ketones.

The per-phospholipid concentration of mid-chain alcohols as AA-derived HETEs (A), EPA-derived HEPEs (B), DHA-derived HDoHEs (B) and AA-derived ketones (C) are shown in lipoproteins and plasma. Regioisomers are shaped and shaded by our best estimate of homologous double-bond positions across all figures. HETEs are mainly products of LOX, but are also products of CYP and autooxidation. KETEs are the HETE-dehydrogenase product of HETEs. Values for EPA and DHA ketones were not available. The effect of P-OM3 adjusted for age and sex was uniform regardless of regioisomer. The mean adjusted effects are shown graphically in Figure S2 in File S1; A mixed model ANOVA was used to test for differences on ln[nM oxylipin/mM PL]. The least-squares mean [95% CI] are shown. Tukey's HSD test was used for post hoc differences between regioisomer levels, and regioisomers sharing a letter are not different. Since no interactions were significant, the term was dropped and the parameter estimate was used to calculate a mean difference [95% CI]. Note that since the test was on log-transformed data, the effect is a proportional one (i.e. percent change). Note the log scale of the y-axis. Note that EPA and DHA oxylipins are shown on the same graphs for brevity, but constitute separate ANOVA tests. a9-HETE is an autooxidative product and is not formed by LOX.

Figure 3. Distribution of fatty acids epoxides and vicinal diols.

The per-phospholipid concentrations of epoxides as AA-derived EpETrEs (A), EPA-derived EpETEs (B), DHA-derived EpDPEs (B) AA-derived DiHETrEs (C), EPA-derived Di HETEs (D), and DHA-derived DiHDPAs (D) are shown in lipoproteins and plasma. Regioisomers are color coded by our best estimate of homologous double-bond positions across all figures. Epoxides are products of CYP-epoxygenase and can also be generated via autooxidation. Vicinal diols are the unique products of soluble epoxide hydrolase on epoxides. The effect of P-OM3 adjusted for age and sex was uniform regardless of regioisomer. The mean adjusted effects are shown graphically in Figure S2 in File S1; A mixed model ANOVA was used to test for differences on ln[nM oxylipin/mM PL]. The least-squares mean [95% CI] are shown. Tukey's HSD test was used for post hoc differences between regioisomer levels, and regioisomers sharing a letter are not different. Since no interactions were significant, the term was dropped and the parameter estimate was used to calculate a mean effect [95% CI]. Note that since the test was on log-transformed data, the effect is a proportional one (i.e. percent change). Note the log scale of the y-axis. Note that EPA and DHA oxylipins are shown on the same graph for brevity, but constitute separate ANOVA tests.

Figure 4. Correlation Heat Map for HDL and LDL.

The Pearson correlation coefficient (r) among lipoprotein-oxylipins for HDL (A) and LDL (B) is shown before treatment (top), after treatment (middle) as correlation of nM oxylipin/mM PL along with the change in oxylipins due to treatment (bottom) expressed as ln(nM oxylipinfinal/mM PLfinal)/ln(nM oxylipinbaseline/mM PLbaseline).

Figure 5. Correlation Heat Map for VLDL and whole plasma-oxylipins.

The Pearson correlation coefficient (r) among lipoprotein-oxylipins for VLDL (A) and whole plasma oxylipins (B) is shown before treatment (top), after treatment (middle) as correlation of nM oxylipin/mM PL along with the change in oxylipins due to treatment (bottom) expressed as ln(nM oxylipinfinal/mM PLfinal)/ln(nM oxylipinbaseline/mM PLbaseline).