Flaxseed Consumption Reduces Blood Pressure in Patients with Hypertension by Altering Circulating Oxylipins via an Alpha Linolenic Acid Induced Inhibition of Soluble Epoxide Hydrolase.
Flaxseed Consumption Reduces Blood Pressure in Patients with Hypertension by Altering Circulating Oxylipins via an Alpha Linolenic Acid Induced Inhibition of Soluble Epoxide Hydrolase.
Year: 2014
Authors: Caliqiuri, S.P. Aukema, H.M. Ravandi, A. Guzman, R. Dibrov, E. Pierce, G.N.
Publication Name: Hypertension
Publication Details: Volume 64; Issue 1; Pages 53 – 59.
Abstract:
In a randomized, double-blinded, controlled clinical trial, participants with peripheral arterial disease (75 per cent hypertensive) consumed 30 g of milled flaxseed/d for 6 months. The flaxseed group exhibited significant reductions in systolic (minus 10 mm Hg) and diastolic (minus 7 mm Hg) blood pressure. Flaxseed contains the n3 fatty acid alpha linolenic acid. Plasma alpha linolenic acid increased with ingestion of flaxseed and was inversely associated with blood pressure. However, the antihypertensive mechanism was unclear. Oxylipins derived from polyunsaturated fatty acids regulate vascular tone. Therefore, the objective was to examine whether flaxseed consumption altered plasma oxylipins in a manner that influenced blood pressure. Plasma of FlaxPAD (Flaxseed for Peripheral Arterial Disease) participants underwent solid phase extraction and high-performance liquid chromatography-mass spectrometry/mass spectrometry analysis. The flaxseed group exhibited significant decreases in 8 plasma oxylipins versus control. Six of these (5, 6, and 8,9 and 11,12 and 14,15 dihydroxyeicosatrienoic acid and 9,10 and 12,13 dihydroxyoctadecenoic acid) were products of soluble epoxide hydrolase, a pharmacological target for antihypertensive treatment. Patients exhibiting a decrease in total plasma soluble epoxide hydrolase derived oxylipins, exhibited a significant decrease in systolic blood pressure (mean [95 per cent confidence interval], minus 7.97 [minus 14.4 to minus 1.50] mm Hg) versus those who exhibited increased plasma soluble epoxide hydrolase derived oxylipins ( plus 3.17 [ minus 4.78 to 11.13] mm Hg). These data suggest that a flaxseed bioactive may have decreased blood pressure via soluble epoxide hydrolase inhibition. Using a soluble epoxide hydrolase inhibitor screening assay, increasing concentrations of alpha linolenic acid decreased soluble epoxide hydrolase activity. In conclusion, alpha linolenic acid in flaxseed may have inhibited soluble epoxide hydrolase, which altered oxylipin concentrations that contributed to the antihypertensive effects in patients with peripheral arterial disease. (Authors Abstract)
The World Health Organization identified high blood pressure as a global public crisis because it is the leading risk factor attributed to death worldwide. Globally, 40% of individuals have hypertension, and a significant proportion of individuals are undiagnosed, untreated, or have uncontrolled high blood pressure. Therefore, it is important to identify effective and desirable treatment strategies to reduce the prevalence and incidence of hypertension globally.
A potential therapeutic strategy for high blood pressure may be through dietary intervention. Recently, patients with peripheral arterial disease (PAD; 75% with hypertension) who consumed 30g of milled flaxseed daily for 6 months exhibited significant reductions in systolic (-10mmHg) and diastolic (-7mm Hg) blood pressure (the FlaxPAD Trial). Flaxseed contains the n3 fatty acid a-linolenic acid (ALA), lignans and fiber. The participants consuming flaxseed exhibited significant increases in plasma ALA and enterolignans, which were inversely associated with blood pressure. However, the biological mechanism of action responsible for the blood pressure-lowering effects was unclear.
Oxylipins are a class of highly bioactive molecules that include the octadecanoids, eicosanoids, and docosanoids. Oxylipins are derived endogenously from polyunsaturated fatty acids. Because plasma concentrations of the poly-unsaturated fatty acids, ALA and eicosapentaenoic acid, increased in the flax group, we hypothesized that concentrations of oxylipins also changed. Oxylipins have a fundamental role in regulating vascular tone and inflammation. For example, some oxylipins produced from arachidonic acid or linoleic acid have been associated with inflammation, tissue damage, vasoconstriction, and oxidative stress. In contrast, the epoxyeicosatrienoic acids (EETs) produced from arachidonic acid are endothelial-derived hyperpolarizing factors that are associated with vasodilation and natriuresis and may indirectly propagate vasodilation by activating endothelial nitric oxide synthase. EETs can be rapidly metabolized to dihydroxyeicosatrienoic acids (DHETs) by the enzyme soluble epoxide hydrolase (sEH), resulting in a concomitant loss of vasodilation. sEH also converts the protoxins, epoxyoctadecenoic acids, to the dihydroxyoctadecenoic acids, which are potent cytotoxic and proinflammatory metabolites. Because products of sEH propagate loss of vascular relaxation and promote inflammation, targeting sEH inhibition pharmacologically has been used successfully to reduce blood pressure, hypertension-associated renal damage, and infarction size in animal models. The proposed relationship between epoxygenase-derived oxylipins and hypertension is shown.
To test the hypothesis that flaxseed decreased the levels of oxylipins associated with inflammation and vasoconstriction, the study objectives were 3-fold: (1) to characterize and quantify the plasma oxylipin profile of participants with PAD, (2) to compare changes in plasma oxylipin concentrations in the control and flaxseed groups, and to determine the mechanism whereby any changes in oxylipins may have influenced blood pressure in the Flax PAD Trial.
Results
Absolute Plasma Oxylipin Concentrations
The plasma oxylipin concentrations of patients with PAD were quantified before and after dietary intervention. Baseline plasma oxylipin profiles did not differ between the flaxseed and control group. After 6 months of dietary intervention, 6 oxylipins were significantly different between the control and flaxseed group. The ALA-derived 9-hydroxyoctadecatrieonic acid was significantly higher in the flax versus placebo group at 6 months, 11, 12 – DHET, 14,15-DHET, 4-hydroxydocosahexanoic acid (HDOHE), 20-HDOHE, and 19,20 – dihydroxydocosapentanoic acid were significantly lower in the flax versus placebo group at 6 months. By comparison of time, the control group did not exhibit any significant differences in the absolute concentrations of plasma oxylipins from baseline to 6 months. The flax group exhibited a significantly lower concentration only in 14,15-DHET at 6 months versus baseline.
Change in Plasma oxylipin Concentrations
Change in plasma oxylipin concentrations were calculated for those individuals who provided blood samples at baseline and 6 months with an adequate volume of plasma of >200uL(n=76).
Nine plasma oxylipins were significantly different between the control and flax group for change from baseline to 6 months. Participants in the flax group exhibited a decrease in docosahexanoic acid-derived 19,20-dihydroxydocosapentanoic acid (-0.248 [0.73 to 0.23] nmol/L) and 20-HDOHE (-0.0239 [0.41 to 0.36] nmol/L) that was significantly different from the control group that exhibited an increase in docosahexanoic acid-derived 19,20-dihydroxydocosapentanoic acid (+0.19 [0.00267 to 0.3728] nmol/L) and (+0.023 [0.13 to 0.17] nmol/L) and 20-HDOHE, respectively. By contrast, the flax group exhibited an increase in the docosahexanoic acid-derived 4-HDOHE (+0.075 [0.55 to 0.71 nmol/L), which was significantly greater than the control group (+0.047 -0.24 to 0.34) nmol/L). The remaining 6 significant oxylipins were all derived from sHE.
Association Between Change in sEH-Derived Oxylipins and Change in Blood Pressure
A reduction in total plasma sEH-derived oxylipins was associated with a reduction in systolic blood pressure (-7.97 [-14.4 to -1.50] mmHg). This was significantly different from those who exhibited an increase in total plasma sEH products who on average exhibited an increase in systolic blood pressure. Change in diastolic blood pressure or mean arterial pressure were not significantly influenced by a change in total plasma sEH-derived oxylipins . When the change in sEH-derived oxylipins was grouped by DHETs and dihydroxyoctadecenoic acids, there was no significant association to change in blood pressure. On analysis of individual sEH-derived oxylipins, 9,10-dihydroxyoctadecenoic acid and 8,9-DHET were significantly associated with change in systolic blood pressure and mean arterial pressure.
Inhibition of sEH
A strong inverse relationship was observed for ALA and % initial activity of sEH. From the lowest ALA concentration (0.0075 mg/mL) to the highest (0.11 mg/mL), % initial activity of sEH declined from 43% to 12%. By contrast, enterodiol, enterolactone, and linoleic acid were not significantly associated with sEH activity. Because increasing concentrations of ALA lowered sEH activity in the assay, it was hypothesized that plasma from patients in the flax group who had higher ALA concentrations by 6 months should lower sEH activity more than plasma from patients in the placebo group who had lower ALA concentrations. ALA concentrations in randomly selected plasma samples from patients (n=5/group) in the placebo group were 0.037±0.008 mg/mL at baseline and 0.021 ± 0.007 mg/mL at 6 months. ALA concentrations in plasma samples from patients (n=5/group in the flax group were 0.024 ± 0.006 mg/mL at baseline and 0.067 ± 0.0060 mg/mL at 6 months. Lipid extracts from these plasma samples were applied to the sEH assay to determine whether the patient plasma could inhibit sEH activity. Percent initial activity of sEH fell by -10.2% (-30.7% to 10%3) from baseline to 6 months in the flax group and by only -3.10% (-13.6% to 7.44%) from baseline to 6 months in the placebo group. Although this did not achieve statistical significance, the trend for the response was consistent with the hypothesis that ALA can inhibit sEH activity. Interestingly, the change in plasma ALA concentration from 0.024 to 0.067 mg/mL in the flax group demonstrated the same 10% decrease in sEH activity as obtained using 0.0375 to 0.075 mg/mL ALA. Similarly, the decrease in plasma ALA from 0.037 mg/mL to 0.021 mg/mL in the placebo group resulted in a similar increase in sEH activity as obtained in Figure 4 using 0.0375 mg/mL to 0.01875 mg/mL, as both resulted in comparable 1% to 3% increases in sEH activity.
Inflammatory Markers
Plasma C-reactive protein, monocyte chemoattractant protein-1, tumor necrosis factor-a, and interleukin-10 did not differ significantly by group or time point. None of the inflammatory markers were significantly associated with the sEH-derived oxylipins.
Discussion
The ingestion of flaxseed in the FlaxPAD Trial resulted in a large decrease in systolic and diastolic blood pressure in patients with PAD and hypertension. Identifying the biological mechanism for this effect is critical to provide confidence in the antihypertensive action and insight into therapeutic targets. The present data provide evidence that consumption of flaxseed may reduce blood pressure by altering the plasma oxylipin profile via an inhibition of sEH. All 6 of the sEH-derived oxylipins detected significantly decreased in the flax group and increased in the control.
The relationship between sEH-derived oxylipins and blood pressure was confirmed by the positive relationship between sEH-derived oxylipins and systolic blood pressure. More specifically, the reductions in plasma 9,10-dihydroxyoctadecenoic acid and 8,9-DHET were strongly associated with reductions in systolic and mean arterial blood pressure. A similar trend existed for a positive relationship between sEH products and diastolic blood pressure, but this did not achieve statistical significance. A larger sample size may have been required. The relatively small reduction in diastolic blood pressure in comparison with systolic and mean arterial pressure may also have made it more difficult to detect a significant difference.
Because the flaxseed group exhibited significant reductions in all sEH-derived oxylipins detected, it is reasonable to hypothesize that a bioactive component of flaxseed inhitibted sEH. Of the flaxseed bioactives tested, ALA had the only significant relationship with sEH activity. This data are the first to demonstrate an n3 fatty acid inhibiting sEH. Using lipid extracts from plasma obtained from the placebo and flax groups, the same trend was shown consistent with the hypothesis that plasma ALA had the capacity to lower sEH activity. However, these results warrant further investigation in an vivo animal model or in human tissue biopsy samples where sEH protein levels and sEH activity can be quantified. Fig 5 represents the hypothesized ALA-induced alteration of the oxylipin profile and the influence on blood pressure.
The association between sEH-derived oxylipins and blood pressure in the current trial is consistent with previous animal studies. Pharmacological inhibition of sEH has resulted in a reduction of sEH-derived DHETs and blood pressure in animal studies.
The potential mechanisms behind the relationship between sEH-derived oxylipins and blood pressure are numerous. The sEH substrates, EETs, have known vasodilatory actions. However, because of the short half-life of EETs, these sEH substrates were below the level of detection and it was not possible to confirm whether EETs increased in the patient population. Despite this, it is well established that the products of sEH have deleterious effects, and a decrease in sEH products may provide protection against vasoconstriction and inflammation. Interestingly, C-reactive protein concentrations decreased to half the baseline concentration by 6 months in the flax group. However, none of the inflammatory markers achieved statistical significance. This could have been because of the large variation observed in this population. The above hypothesis for a mechanism of antihypertensive action does not discount the possibility that other oxylipins may have contributed as well. For example, reductions in docosahexanoic acid-derived 19,20-dihydroxydocosapentanoic acid and 20-HDOHE were observed in the flax group. Notably, although the biological actions of 19,20-dihydroxydocosapentanoic acid are unknown, it is also thought to be an sEH product. By contrast, an increase in the docosahexanoic acid-derived 4-HDOHE and 9-hydroxyoctadectrienoic acid was observed in the flax group. However, the influence of these oxylipins on vascular tone or blood pressure has yet to be investigated.
