Polyunsaturated fatty acids, specialized pro-resolving mediators, and targeting inflammation resolution in the age of precision nutrition

Abstract

Chronic inflammation contributes toward the pathogenesis of numerous diseases including, but not limited to, obesity, autoimmunity, cardiovascular diseases, and cancers. The discovery of specialized pro-resolving mediators (SPMs), which are critical for resolving inflammation, has commenced investigation into targeting pathways of inflammation resolution to improve physiological outcomes. SPMs are predominately synthesized from the n-3 polyunsaturated fatty acids (PUFA) eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids. Therefore, one viable strategy to promote inflammation resolution would be to increase dietary intake of EPA/DHA, which are deficient in select populations. However, there are inconsistencies between the use of EPA/DHA as dietary or pharmacological supplements and improved inflammatory status. Herein, we review the literature on the relationship between the high n-6/n-3 PUFA ratio, downstream SPM biosynthesis, and inflammatory endpoints. We highlight key studies that have investigated how dietary intake of EPA/DHA increase tissue SPMs and their effects on inflammation. We also discuss the biochemical pathways by which EPA/DHA drive SPM biosynthesis and underscore mechanistic gaps in knowledge about these pathways which include a neglect for host genetics/ethnic differences in SPM metabolism, sexual dimorphism in SPM levels, and potential competition from select dietary n-6 PUFAs for enzymes of SPM synthesis. Altogether, establishing how dietary PUFAs control SPM biosynthesis in a genetic-and sex-dependent manner will drive new precision nutrition studies with EPA/DHA to prevent chronic inflammation in select populations.

1.0. High n-6 and low n-3 PUFA consumption in the western diet may contribute toward chronic inflammation.

The western diet is high in saturated fat and n-6 (ω−6) polyunsaturated fatty acids (PUFAs) and low in long chain n-3 (ω−3) PUFAs. In addition, more than half of the calories consumed in the western diet come from highly processed foods.¹ During the course of human evolution, the ratio of n-6 to n-3 PUFAs has significantly increased from 1:1 to 10:1 today.² This observed change in dietary fatty acid consumption in favor of n-6 PUFAs is a result of a change in our dietary patterns including, but not limited to, the use of vegetable oils and processed foods.^3,4 In the United States, the increase in dietary n-6 PUFAs is a consequence of dietary policy changes in which saturated fat was replaced with n-6 PUFAs in an effort to lower serum cholesterol and LDL.³ However, whether increased intake of n-6 PUFAs has uniform health benefits is currently debated.^3,5–8

While the optimal ratio of n-6 to n-3 PUFAs for human health and disease remains to be established, the increase in dietary consumption of n-6 PUFAs parallels the increased prevalence of adiposity in western nations.^2,9 Consumption of foods enriched in sugar and high-fat in the western diet and limited physical activity are some of the factors driving the obesity epidemic.¹⁰ Obesity, defined as BMI ≥ 30 kg/m², occurs through the deposition of excess lipids into adipose tissue. Excess adiposity leads to a chronic state of meta-inflammation.¹¹ Individuals with obesity have greater risk for chronic disease and often present with clinical parameters of metabolic syndrome (MetS), such as increased waist circumference, high blood pressure, high fasting triglycerides, decreased high-density lipoprotein (HDL), high low-density lipoprotein (LDL), high total cholesterol and elevated glucose.¹² Since obesity and MetS are associated with increased risk for varying complications, increased dietary intake of n-3 PUFAs may be important for mitigating disease risk.^12–15

There is some debate about the role of n-6 PUFA consumption for differing aspects of human health including chronic inflammation. The prevailing viewpoint is that increased dietary intake of n-6 PUFAs, in comparison to saturated fatty acids, lowers the risk of differing cardiovascular diseases.^3,16 For instance, decreasing the intake of saturated fat or replacement with mono- or polyunsaturated fatty acids, including n-6 PUFAs, reduces the risk of coronary heart disease.¹⁷ However, a 2018 systematic review on n-6 PUFAs on cardiovascular health reported no benefit for increased consumption of n-6 fatty acids to reduce cardiovascular outcomes except for myocardial infarction.¹⁸

One n-6 PUFA that is of particular relevance to inflammation is linoleic acid (18:2). Current intake of linoleic acid (the most abundant PUFA in the western diet) is ~7% of total energy in the U.S,, which is 14–18 times the amount required to prevent linoleic acid deficiency.^19,20 The role of linoleic acid on inflammation is debated.^21–26 Some studies suggest that linoleic acid is pro-inflammatory, particularly at the pre-clinical level, while others do not support this notion and even suggest an anti-inflammatory role for linoleic acid.^27,28 There is epidemiological evidence to suggest that linoleic acid intake is inversely associated with total mortality.²⁹ The debate may stem from the notion that studies on linoleic acid are from a wide range of systems such as obesity to cancer models.²⁵ Thus, there is a critical need to investigate if lowering excess n-6 PUFAs including linoleic acid in the diet may potentially improve outcomes related to chronic inflammation. It is also important to establish how gene-diet interactions impact circulating levels of n-6 and n-3 PUFAs.⁸ This is discussed below at a molecular level in the context of targeting pathways of inflammation resolution.

2.0. Chronic inflammation is central toward the pathogenesis of obesity and its complications.

Obesity is well established as a state of chronic low-grade inflammation. Adipose tissue depots, liver, muscle and pancreas are primary sites of inflammation in response to diet-induced obesity.³⁰ There is also evidence for inflammation in the brain, which may drive changes in food intake.³¹ In metabolically actives tissues such as the adipose, there is an infiltration of macrophages and other immune cells, as well as a shift away from anti-inflammatory immune cells.³⁰ Adipocytes and macrophages are a source of pro-inflammatory cytokines such as IL-1, TNFα, and IL-6.³⁰ Individuals consuming a western diet can have elevated levels of IL-6, IL-1β, and TNFα, characteristic of obesity, in comparison to healthy individuals.^4,9

Diet-induced obesity is associated with many complications such as cancers, behavioral disorders, non-alcoholic fatty liver disease, cardiovascular complications, and insulin resistance. Notably, MetS is associated with high blood glucose, abnormal cholesterol levels and high triglycerides accompanied by upregulation of pro-inflammatory pathways.¹² High blood glucose and elevated HbA1c is associated with insulin resistance and type 2 diabetes. Adipose tissue in obesity and MetS is characterized by increased lipolysis with the excessive release of free fatty acids, and is also a source of pro-inflammatory cytokines; both of these factors may inhibit insulin action.³² Systemic inflammatory markers are also risk factors for the development of type 2 diabetes and its macrovascular complications. As an example, the pro-inflammatory interleukin-1β is implicated in the pathogenesis of type 2 diabetes through the activation of the NLRP3 inflammasome.³⁰

A prominent feature of MetS is dysregulated lipid metabolism, contributing to increased fasting plasma triglycerides, high LDL cholesterol and low HDL cholesterol.³³ The presence of the small dense LDL phenotype, postprandial hyperlipidemia with accumulation of atherogenic remnants and hepatic overproduction of apoB containing lipoproteins are metabolic risk factors associated with obesity.³³ All these lipid abnormalities are typical features of the MetS and may be associated with a pro-inflammatory gradient.³³

An often-overlooked complication of obesity is increased risk of infection, potentially due to impairments in pathways of inflammation resolution. For instance, the western diet triggers NLRP3-dependent innate immune reprogramming and mice fed a western diet exhibit impaired cellular functionality that is critical for combatting secondary infections.³⁴ Several viruses or virus-like agents such as members of adenoviridae, herpesviridae, slow virus (prion), and hepatitides, have been associated with increased risk for infection in response to obesity.³⁵ Obesity is also a critical factor for vulnerability to influenza A/pdmH1N1 virus infection, and high fat diets rich in saturated fats and cholesterol also negatively impact the pathogenesis of HIV/SIV infection.^5,36 Mechanistically, obesity increases the risk for infection and drives poor responses to vaccinations in part due to compromised humoral immunity and impaired T cell metabolism including poor memory responses from pulmonary T cells upon influenza infection.^37–40

Very recently, epidemiological research on the impact of obesity on SARS-CoV-2, which is driving the COVID-19 pandemic, suggests that individuals with obesity are at increased risk for infection and complications of viral infection such as increased mechanical ventillation.⁴¹ The effects of obesity are not just limited to viruses as obese mice display increased susceptibility to bacterial infections such as Staphylococcus aureus.⁴² Thus, there may be a potential role for diet-driven inflammation that is contributing toward increased risk for infection in individuals with obesity and/or MetS. In fact, a failure to resolve inflammation may be one major factor that is contributing toward increased risk for infection.⁴³

3.0. Specialized pro-resolving mediators are critical for resolving inflammation.

A plausible approach to combatting the pro-inflammatory state associated with chronic inflammatory diseases such as obesity and its complications is to increase dietary consumption of the long chain n-3 PUFAs eicosapentaenoic (EPA, 20:5) and docosahexaenoic (DHA, 22:6) acids. Here we briefly cover how EPA and DHA give rise to downstream metabolites that are critical regulators of inflammation resolution and highlight key functional roles for these metabolites in the context of inflammation and infection.

EPA and DHA are enzymatically converted into specialized pro-resolving mediators (SPMs).⁴⁴ SPMs act as potent signaling molecules, binding to their receptors on immune cells in various tissues.⁴⁵ The role of SPMs is to promote immunoresolution, a process by which inflammation is resolved to its homeostatic state.⁴⁵ Each SPM has its own functional role through the binding of specific receptors. EPA derived SPMs are known as the E-series resolvins, while DHA derived SPMs comprise of the D-series resolvins, maresins, and protectins.⁴⁶ There is also one class of SPMs produced from arachidonic acid, an n-6 PUFA, known as the lipoxins.⁴⁵ An overview of the biosynthesis of EPA- and DHA-derived SPMs is depicted in Figure 1. In addition, Figure 1 shows the major downstream metabolites of arachidonic acid.

Figure 1: Metabolic pathways by which PUFAs give rise to downstream metabolites of inflammation resolution and their relationship with the obesogenic western diet.

Arachidonic acid (20:4, AA, n-6) gives rise to a range of metabolites such as leukotrienes that drive an inflammatory response (not depicted in detail for simplicity). AA also gives rise to SPMs known as lipoxins. Eicosapentaenoic acid (20:5, EPA, n-3) and docosahexaenoic acid (22:6, DHA, n-3) are precursors for SPMs known as resolvins, protectins, and maresins. Excess consumption of n-6 PUFAs, particularly driven by linoleic acid (18:2, LA, n-6), and low dietary intake of n-3 PUFAs in the western obesogenic diet, may contribute toward a reduction in circulating levels of SPMs. LA may specifically bind key enzymes of SPM biosynthesis. This would limit the ability to resolve inflammation and potentially contribute toward chronic inflammation. In addition, SNPs associated with various enzymes of PUFA metabolism and SPM biosynthesis may be a source of heterogeneity in the concentration of SPMs in the human population.

To produce the E-series resolvins, EPA undergoes modification by either the cytochrome P450 (CYP450) or cyclooxygenase-2 (COX-2) enzymes to produce the intermediate, 18-HpEPE. Further modification of 18-HpEPE by the enzyme arachidonate 15-lipoxygenase (ALOX15) produces resolvin E3 (RvE3), one of the E-series resolvins. 18-HpEPE can be further metabolized into 18-HEPE by 5-lipoxygenase (ALOX5). 18-HEPE is then converted into the other two E-series resolvins, resolvin E1 (RvE1) and resolvin E2 (RvE2) via leukotriene-A4 hydrolase (LTA4-H) or a reduction reaction, respectively.^47–49 RvE1 specifically binds to the Chemerin 23 (ChemR23) receptor and leukotriene B4 (LTB4) receptor, BLT1.⁵⁰

RvE1 binding to ChemR23 on macrophages induces phagocytosis and clearance of debris, and promotes an immunoresolving phenotypic switch from classically activated (M1-like) to non-activated macrophages (M2-like).^45,50,51 On the other hand, RvE1 binding BLT1 blocks LTB4 binding on lymphocytes, macrophages, and neutrophils to decrease immune cell chemotaxis to the area of injury and clearance of inflammatory neutrophils by non-classically activated macrophages.^50,52–54 It is known that antagonistically inhibiting LTB4 on leukocytes, such as B cells, prevents the development of insulin resistance in C57BL6/J mice.⁵⁵ RvE1 also reverses hyperinsulinemia and hyperglycemia in C57BL6/J obese mice through a ChemR23 mediated mechanism independent of a role for adipose tissue inflammation.⁴⁷ Similar to RvE1, RvE2 is a partial agonist to ChemR23 and a BLT1-LTB4 antagonist; ^56,57 furthermore, RvE2 reduces neutrophil infiltration and chemotaxis.⁵⁸ Overexpression of the ChemR23 receptor drives improvements in glucose homeostasis of mice.⁵⁹ It is still unknown which receptors RvE3 binds to; however, it is hypothesized that RvE3 reduces neutrophil infiltration but more research is required to identify RvE3’s specific functions.⁶⁰

The D-series resolvins, protectins and maresins are produced by DHA. DHA is first metabolized by ALOX15 to produce 17S-HpDHA. 17-HpDHA is then a substrate for ALOX5 and a series of peroxidase and hydrolysis reactions to produce the resolvins RvD1, RvD2, RvD3, RvD4, RvD5, and RvD6. RvD1 binds the human G protein-coupled receptor 32 (GPR32) and N-formyl peptide receptor 2 (ALX/FPR2) whereas RvD2 binds GPR18.^61–63 RvD1 and RvD2 rescue diminished adiponectin levels in obese adipose tissue and decrease monocyte chemoattractant protein-1 (MCP1) and LTB4 stimulated monocyte migration.⁶⁴ RvD1 and RvD2 increase macrophage phagocytosis in the context of an infection.^56,65 Furthermore, RvD1 decreases time of epithelial closure and presence of apoptotic cells in diabetic wounds.⁶⁶ RvD3 also exerts antimicrobial actions via GPR32, increases neutrophil clearance, and decreases production of inflammatory prostaglandins and leukotrienes.⁶⁷ Similarly, RvD4 has shown host protection and bacterial clearance against Staphylococcus aureus infections.⁶⁸ RvD4 decreases neutrophil infiltration and increases pro-resolving monocytes in deep vein thrombosis.⁶⁹ RvD5 binds the GPR32 receptor and increases macrophage phagocytosis during E. coli infections.⁵⁶ As for RvD6, it has been hypothesized to aid in corneal wound healing and nerve damage.⁷⁰

17S-HpDHA is also converted into Protectin D1 (PD1) and is an isomer of Protectin DX (PDX). PD1 is neuroprotective by increasing nerve regeneration and decreasing perception of pain.^57,71 Furthermore, PD1 along with RvD1 and RvD5 increase survival and decrease antibiotic usage in response to bacterial infections.⁵⁷ In addition, DHA is converted by ALOX12 into 14S-HpDHA to produce maresin-1 (MaR1) and maresin-2 (MaR2). MaR1 increases macrophage phagocytosis of apoptotic neutrophils and induces tissue regeneration.⁷² Furthermore, the MaR1 precursor 14-HDHA induces a phenotypic switch in macrophages from an M1-like classically activated phenotype to a pro-resolving M2-like phenotype.⁴⁵ Lastly, MaR2 increases phagocytosis of apoptotic neutrophils.⁵⁷ Taken together, SPMs are critical in orchestrating inflammation resolution and targeting the aforementioned pathways of SPM biosynthesis may be a therapeutic approach, particularly for those diseases associated with SPM deficiency.

4.0. SPM biosynthesis is unbalanced in obesity and its metabolic complications.

There is strong literature support for the notion that obesity drives a deficiency in the concentration of n-3 PUFA-derived SPMs in both human and murine models.⁷³ In mice, feeding an obesogenic western diet induces systemic inflammation and functional reprogramming, ³⁴ which is accompanied by decreased levels of RvD1, 17-HDHA, and PD1 in white adipose tissue.⁷³ Strikingly, a high fat diet can drive a reduction in SPM levels within days. For instance, one lab reported that DHA-derived SPMs and their precursors are lowered in white adipose tissue within four days of intervention with a high fat diet compared to mice on a lean diet.⁷³ This raises the possibility that it is the high fat diet and not the state of obesity itself that may be driving a reduction in the concentration of SPMs.

The effects of obesity are not just on DHA-derived SPMs as a recent study reported that 18-hydroxyeicosapentaenoic acid, the precursor for RvE1 was strongly diminished in white adipose tissue and liver.⁴⁷ Administration of dietary EPA or the downstream RvE1 to obese mice strongly improved hyperinsulinemia and hyperglycemia. Another research group reported that SPMs are increased in a model of mouse liver steatosis although the levels of EPA and DHA were decreased in response to the liver steatosis.⁶⁶ Thus, this study suggests that obesity may drive an unbalanced concentration of SPMs rather than a universal deficiency. This study underscores the need to study the kinetics of inflammation resolution as mass spectrometry measurements for SPMs at a single time point do not provide a comprehensive picture of the inflammatory profile.

SPM levels appear to be lowered in circulation and secondary lymphoid organs in response to obesity.³⁷ Notably, the splenic SPM precursors 14-HDHA, 17-HDHA and PDX were decreased in obese male but not female C57BL/6J mice.⁷⁴ This lowering of lipid mediators was associated with an increase in the abundance of n-6 PUFAs.⁷⁴ Administration of dietary DHA can also restore endogenous biosynthesis of SPMs (14-HDHA, 17-HDHA and PDX) in mice consuming a western diet, and treatment with 17-HDHA reduced inflammatory cytokine expression in adipocytes.^37,73

Leukocytes from individuals with obesity also exhibit an impaired SPM signature and display a reduction in select SPMs.^74,75 There is unbalanced production of SPMs (i.e., D- and E-series resolvins, PD1, MaR1, and lipoxins) with respect to inflammatory lipid mediators (i.e., LTB4 and prostaglandins) in omental adipose tissue from study subjects with obesity.⁷⁶ In a 2019 study by López-Vicario et al., individuals with obesity displayed a notable reduction in leukocyte 17-HDHA. The reduction in 17-HDHA was potentially driven by reduced activity of ALOX15, which is critical for the biosynthesis of SPMs.⁷⁵ There is also evidence that MaR1 levels are significantly decreased in type 2 diabetics compared to controls.⁷⁷

A major gap in knowledge is the mechanisms by which components of the western diet may drive an impaired SPM signature. Potential mechanisms include the possibility that n-6 and n-3 PUFAs may compete with each other for esterification into the membrane phospholipid pool.⁷⁸ Thus, competition between differing fatty acids from the diet for membrane phospholipids could impact the bioavailability of fatty acids for downstream SPM biosynthesis. Notably, the n-6 PUFA linoleic acid can bind some of the same enzymes such as 12/15-LOX that generate SPMs from DHA, which could prevent SPM biosynthesis (Figure 1).^79,80 Indeed, a very recent study showed dietary linoleic acid inhibited the synthesis of DHA-derived SPMs in mice.⁸¹ Thus, excess linoleic acid may decrease available substrate for key enzymes used for SPM biosynthesis. Another possibility, which may not be mutually exclusive, is that obesity or the consumption of high fat/high sugar diet may be driving a reduction in the expression of enzymes of SPM biosynthesis and/or a reduction in their activity. The premise for this hypothesis is based on the López-Vicario et al., study described above on enzyme activity. Clearly, more work is needed in this area at a mechanistic level. Finally, there is the possibility that SPM biosynthesis is simply low due to poor intake of dietary n-3 PUFAs in the western diet.⁸² Therefore, future studies should compare the role of the western diet on SPM biosynthesis with other dietary patters such as the Mediterranean diet, which contains higher amounts of in n-3 PUFAs.⁸³

5.0. The use of dietary n-3 PUFAs to mitigate SPM deficiencies in select populations to improve chronic inflammatory outcomes.

There is suggestion in the literature that administration of long chain n-3 PUFAs has a positive role in the prevention and treatment of the pathologies associated with obesity, MetS, and inflammation.⁸⁴ Increased consumption of n-3 PUFAs in the diet can reduce plasma levels of the pro-inflammatory cytokines IL-6 and tumor necrosis factor-alpha (TNFα), as well as plasma C-reactive protein (CRP). ⁸⁴ These effects are thought to be mediated by SPMs.⁸⁴ However, the link between dietary n-3 PUFAs, SPMs, and improvements in inflammation are just starting to emerge. A recent study shows that supplementation with a marine oil known as SPM Active leads to an increase in peripheral blood SPM concentrations and reprograms peripheral blood cells, indicating a role for SPMs in mediating the immune-directed actions of this supplement.⁸⁵ This supplement is formulated to provide specific amounts of SPM precursors.

There are studies that demonstrate a positive correlation between n-3 PUFA supplementation and downstream SPM production in humans. As an example, one study reported that human plasma and serum SPM clusters were increased after n-3 PUFA supplementation.⁸⁶ Studies administering dietary DHA showed increased 12/15-LOX and decreased 5-LOX expression in lymph nodes and isolated lymph node PMN, which correlated with amplified LXA4 formation.⁸⁷ The increase in LXA4 from DHA was unexpected and the underlying mechanism of action remains unclear. Other studies involving n-3 PUFAs have shown increased DHA-derived pro-resolving mediators in women with obesity.⁸⁸ Additionally, patients with coronary artery disease with low or absent levels of specific SPMs had complete restoration with Lovaza, which is a prescription EPA/DHA ethyl ester formulation.⁸⁹

One area of study that requires further attention is the role of dietary docosapentaenoic acid (DPA, 22:5) on biosynthesis of downstream SPMs. Dietary supplementation with DPA can increase downstream SPMs. In a double-blind crossover study, DPA supplementation increased the SPM resolvin RvD5n-3DPA and MaR-1, the vicinal diol 19,20-dihydroxy-DPA and n-6 PUFA derived 15-keto-PGE2.⁹⁰ However, the effects of DPA supplementation on inflammation resolution remain to be established and is an area for future investigation.

It is critical to recognize that not all individuals will display SPM deficiencies and thus the need for precision interventions. A major goal of precision nutrition/medicine is to provide targeted interventions for select populations.⁹¹ For instance, only those individuals with obesity that are SPM deficient may be better candidates for dietary intervention with n-3 PUFAs. In fact, it is increasingly recognized that there is tremendous heterogeneity within the population with obesity and it is better to think about ‘obesities’ rather obesity as a single disease state.^92,93 Therefore, there is likely tremendous heterogeneity in circulating SPM levels across differing inflammatory conditions. Of course, a range of other factors will also impact SPM levels in humans, as discussed in the subsequent sections below.

6.0. Genetic differences in SPM metabolism may be a critical factor in translating n-3 PUFAs for improving inflammation.

There are significant discrepancies in the literature on the potential use of n-3 PUFAs for improving inflammatory outcomes. Of course, there are various causes that may contribute to differences in reported outcomes with n-3 PUFA supplementation studies, particularly through SPM-mediated mechanisms. In Figure 2, we present an overview of these potential causes, which we discuss below in greater detail. Notably, the relative amount of dietary intake of differing n-3 and n-6 PUFAs coupled with differences in each individual’s metabolic and microbiome status, sex, and host genetic background will impact the concentration of SPMs, influence disease risk, and ultimately inflammation resolution. Furthermore, age and environmental exposures (not depicted for simplicity in Figure 2) will also impact the individual’s metabolic and microbiome status and thereby influence inflammatory outcomes.

Figure 2: Framework for future precision nutrition studies with dietary EPA and DHA for inflammation resolution.

The depicted framework underscores the need to establish the amount of dietary intake of differing n-3 and n-6 PUFAs and accounting for each individual’s metabolic status, microbiome status, and sex. The potential influence of age and environmental exposures on metabolic and microbiome status are not depicted but are also of relevance. In addition, the host genetic background will also impact circulating levels of n-3 and n-6 PUFAs and thereby the synthesis of downstream SPMs. Ultimately, these factors will influence the concentration of SPMs, disease risk and inflammation resolution. For simplicity, we depict a few SNPs associated with PUFA metabolism and their related disease risk phenotypes.

One potential source of discrepancy in the literature for n-3 PUFA efficacy for inflammation is that production of EPA- or DHA-derived SPMs can highly depend on the ratio of EPA and DHA in the supplement. Given that both EPA and DHA will compete with arachidonic acid to bind phospholipase A2, the ratio of each n-3 PUFA in the supplement may confound results from study to study.⁹⁴ Moreover, each SPM has differing functions that are tissue and cell specific.⁴⁶ Therefore, the SPMs that are produced from EPA and/or DHA in the supplement will result in targeted activation of the particular receptors that those SPMs bind.⁴⁶ Hence, the formulation of the n-3 PUFA supplement potentially determines the SPM pool that will be produced and actively contribute to the phenotypic outcome of interest measured in the study.

The effects of dietary n-3 PUFAs may be further influenced by the background dietary levels of n-6 PUFAs, notably the n-6 PUFAs linoleic and arachidonic acid (Figure 1). Often, n-6 PUFA levels are not accounted for in clinical trials. Linoleic acid, as discussed above, may have a role in influencing SPM levels, which may a direct effect of linoleic acid or through the biosynthesis of arachidonic acid. Many of the metabolites synthesized from arachidonic acid can be pro-inflammatory. However, SPMs can be synthesized from arachidonic acid. These SPMs are lipoxins, which consist of LXA4 and LXB4.^44,95 LXA4 binds the ALX/FPR2 receptor and has various effects over a wide variety of conditions ranging from ulcerative colitis remission to periodontal disease and hepatic steatosis.^57,96–98 Thus, the role of n-6 PUFAs on dietary n-3 PUFA mediated SPM synthesis is complicated as not all n-6 PUFA-derived metabolites are pro-inflammatory.

Another potential cause for discrepancy in studies with n-3 PUFAs in humans is the lack of accounting for host-genetics. Clinical studies often include a wide range of individuals with a diverse genetic background. There is strong evidence for the role of host genetics with EPA/DHA metabolism but far less is known about SPM biosynthesis and metabolism. For instance, around 80% of African Americans have two alleles associated with higher arachidonic acid levels whereas only about 45% of European Americans have those same alleles.⁹⁹ This suggests that differences between circulating or tissue-specific levels of PUFAs among various genetic or ethnic backgrounds could impact the concentration of downstream SPMs.⁹⁹

Many single nucleotide polymorphisms (SNPs) in the enzymes or receptors of the SPM synthesis pathway have direct clinical translation in metabolic-related diseases in the human population as well. In fact, we have previously shown that when mining the dbSNP and 1000 genomes databases there is a large range of minor allele frequencies present in various enzymes and receptors of SPM production pathways.⁴⁷ When examining the first step in the SPM biosynthesis pathways, the CYP450 enzymes with the capacity to metabolize EPA or DHA have numerous polymorphisms related to various metabolic disorders.¹⁰⁰ CYP2C8, CYP2C9, and CYP2C19 contain SNPs that have been associated with increased susceptibility to type 2 diabetes in Indian, Japanese, and Saudi populations.^101–104 Additionally, two SNPs in CYP4F2 are associated with coronary heart disease in the Chinese population.¹⁰⁵ On the other hand, the etiology and risk of developing type 2 diabetes has been associated with the COX-2 enzyme (rs5275 variant) in type 2 diabetic patients.¹⁰⁶ Furthermore, when examining African Americans with the COX-2 G-765C polymorphism, researchers found a higher risk of stroke with that particular COX-2 variant.¹⁰⁷ Three SNPs in the COX-2 enzyme are associated with coronary/carotid calcified plaques in patients from the Diabetes Heart Study.¹⁰⁸

Further downstream in the E-series biosynthesis pathway, SNPs in FLAP or LTA4-H are correlated with a two-fold risk for myocardial infarction and stroke.^109,110 These particular SNPs in FLAP that were correlated with myocardial infarction and stroke were found to be in greater abundance in the Finnish, English, and Scottish populations.¹¹¹ On the other hand, the enzyme bound to FLAP, ALOX5, has SNPs that are associated with modest increases in body mass index and cardiovascular disease risk.¹¹² Particularly, SNPs in the promoter of ALOX5 are significantly associated with coronary artery disease in white Europeans, with a minor allele frequency of 15%.¹¹³ Polymorphisms in ALOX15, the enzyme that produces RvE3 and 17S-HpDHA, have also been associated with ischemic stroke or coronary artery disease in the Chinese Han population and North Indian population, respectively.^114,115 The ALOX12 polymorphism (rs2073438) is significantly associated with total fat mass and percentage fat mass in obese Chinese men.¹¹⁶ As for polymorphisms in SPM receptors, one group discovered a polymorphism (rs1878022) in ERV1/ChemR23 that increases its expression and leads to reduced levels of pro-inflammatory cytokines in adipose tissue and plasma of individuals with morbid obesity.¹¹⁷ Taken together, these studies highlight the importance of accounting for host-genetics and ethnic differences in future pre-clinical and clinical studies with SPMs and dietary n-3 PUFAs.

7.0. Other factors that could influence SPM availability in humans.

There are likely additional factors that will contribute toward SPM bioavailability in humans. One notable factor is sex. It is established that synthesis of DHA is lower in males compared to females.¹¹⁸ There are also data demonstrating sex differences in SPM bioavailability in mice and humans. In obese mice, SPM levels were generally higher in female mice than males.⁷⁴ In a human study, females were found to have higher levels of SPMs relative to male counterparts. Notably, the sex difference in this study correlated with females having increased protection from inflammation-driven endothelial impairments. The differences in SPM levels between humans may be tissue specific as SPMs were absent in male human tears compared to females.¹¹⁹

Another factor that could influence SPM metabolism is the composition and secretion of metabolites from the microbiome. Specific dietary patterns control microbial composition and thereby tissue-specific and circulating inflammatory status. For instance, a recent longitudinal study demonstrated that a shift toward consumption of a Mediterranean diet in male subjects leads to a unique microbial composition that is associated with improved cardiometabolic outcomes. Therefore, each individual’s dietary pattern and thereby their microbiome profile will likely influence SPM metabolism.¹²⁰

A major step forward will be to establish SPM levels in SPM-deficient populations in response to well defined EPA and/or DHA supplements. Clinical studies will also benefit by genotyping individuals, measuring background levels of n-6 PUFAs, accounting for potential sex differences in SPM metabolism, and establishing a role for the microbiome in SPM bioavailability. This will then set the basis for future precision nutrition randomized clinical trials for specific outcomes related to chronic inflammation, infection, insulin resistance, etc. Ultimately, these studies will lead to precision clinical trials and perhaps even improved dietary recommendations for select clinical populations.

8.0. Conclusions.

The western diet is low in the long chain n-3 PUFAs EPA and DHA. This may contribute toward a chronic pro-inflammatory state that is associated with the pathogenesis of differing diseases such as those complications associated with obesity. One approach to targeting SPM deficiencies in select clinical populations to drive inflammation resolution is to increase dietary consumption of long chain n-3 PUFAs. However, key mechanistic gaps in knowledge remain on the causes of SPM deficiencies and approaches to overcoming these deficiencies. Notably, there is a need to understand how to effectively promote SPM biosynthesis with n-3 PUFAs while accounting for background levels of n-6 PUFAs, sex differences in SPM levels, baseline microbiome profiles, and host genetic differences in SPM biosynthesis/metabolism. Ultimately, this line of research from basic molecular studies to pre-clinical and clinical experimentation will drive precision nutrition trials with EPA and DHA for select populations that are potentially SPM deficient to improve inflammatory outcomes.

Highlights.

• The western diet is generally low in long chain n-3 PUFAs

• Low dietary n-3 PUFAs may contribute to a pro-inflammatory profile

• PUFA-derived SPM levels are unbalanced in chronic inflammatory diseases

• SNPs in enzymes of SPM biosynthesis may have direct clinical translation

• Establishing genetic differences in SPM metabolism will drive precision studies