Pros and Cons of Long-Chain Omega-3 Polyunsaturated Fatty Acids in Cardiovascular Health.

The long-chain omega-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are found in seafood, supplements, and concentrated pharmaceutical preparations. Prospective cohort studies demonstrate an association between higher intakes of EPA+DHA or higher levels of EPA and DHA in the body and lower risk of developing cardiovascular disease (CVD), especially coronary heart disease and myocardial infarction, and of cardiovascular mortality in the general population. The cardioprotective effect of EPA and DHA is due to the beneficial modulation of a number of risk factors for CVD. Some large trials support the use of EPA+DHA (or EPA alone) in high-risk patients, although the evidence is inconsistent. This review presents key studies of EPA and DHA in the primary and secondary prevention of CVD, briefly describes potential mechanisms of action, and discusses recently published RCTs and meta-analyses. Potential adverse aspects of long-chain omega-3 fatty acids in relation to CVD are discussed.

E-series resolvin metabolome, biosynthesis and critical role of stereochemistry of specialized pro-resolving mediators (SPMs) in inflammation-resolution: Preparing SPMs for long COVID-19, human clinical trials, and targeted precision nutrition.

The COVID-19 pandemic has raised international awareness of the importance of rigorous scientific evidence and the havoc caused by uncontrolled excessive inflammation. Here we consider the evidence on whether the specialized pro-resolving mediators (SPMs) are ready to meet this challenge as well as targeted metabololipidomics of the resolution-inflammation metabolomes. Specific stereochemical mechanisms in the biosynthesis of SPMs from omega-3 essential fatty acids give rise to unique local-acting lipid mediators. SPMs possess stereochemically defined potent bioactive structures that are high-affinity ligands for cognate G protein-coupled surface receptors that evoke the cellular responses required for efficient resolution of acute inflammation. The SPMs biosynthesized from the major omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are coined Resolvins (resolution phase interaction products; E series and D-series), Protectins and Maresins (macrophage mediators in resolving inflammation). Their biosynthesis and stereochemical assignments are established and confirmed (>1,441 resolvin publications in PubMed.gov) as well as their functional roles on innate immune cells and adaptive immune cells (both lymphocyte T-cell subsets and B-cells). The resolution of a protective acute inflammatory response is governed mainly by phagocytes that actively clear apoptotic cells, debris, blood clots and pathogens. These resolution phase functions of the acute inflammatory response are enhanced by SPMs, which together prepare the inflammatory loci for homeostasis and stimulate tissue regeneration via activating stem cells and the biosynthesis of novel cys-SPMs (e.g. MCTRs, PCTRs and RCTRs). These cys-SPMs also activate regeneration, are organ protective and stimulate resolution of local inflammation. Herein, we review the biosynthesis and functions of the E-series resolvins, namely resolvin E1 (the first n-3 resolvin identified), resolvin E2, resolvin E3 and resolvin E4 biosynthesized from their precursor eicosapentaenoic acid (EPA), and the critical role of total organic synthesis in confirming SPM complete stereochemistry, establishing their potent functions in resolution of inflammation, and novel structures. The physical properties of each biologically derived SPM, i.e., ultra-violet (UV) absorbance, chromatographic behavior, and tandem mass spectrometry (MS2) fragmentation, were matched to SPMs biosynthesized and prepared by stereospecific total organic synthesis. We briefly review this approach, also used with the endogenous D-series resolvins, protectins and maresins confirming their potent functions in resolution of inflammation, that paves the way for their rigorous evaluation in human tissues and clinical trials. The assignment of complete stereochemistry for each of the E and D series Resolvins, Protectins and Maresins was a critical and required step that enabled human clinical studies as in SPM profiling in COVID-19 infections and experimental animal disease models that also opened the promise of resolution physiology, resolution pharmacology and targeted precision nutrition as new areas for monitoring health and disease mechanisms.

Specialized pro-resolving mediators as modulators of immune responses.

Specialized pro-resolving mediators (SPMs) are endogenous small molecules produced mainly from dietary omega-3 polyunsaturated fatty acids by both structural cells and cells of the active and innate immune systems. Specialized pro-resolving mediators have been shown to both limit acute inflammation and promote resolution and return to homeostasis following infection or injury. There is growing evidence that chronic immune disorders are characterized by deficiencies in resolution and SPMs have significant potential as novel therapeutics to prevent and treat chronic inflammation and immune system disorders. This review focuses on important breakthroughs in understanding how SPMs are produced by, and act on, cells of the adaptive immune system, specifically macrophages, B cells and T cells. We also highlight recent evidence demonstrating the potential of SPMs as novel therapeutic agents in topics including immunization, autoimmune disease and transplantation.

Biosynthetic deficiency of docosahexaenoic acid causes nonalcoholic fatty liver disease and ferroptosis-mediated hepatocyte injury.

Exogenous omega-3 fatty acids, particularly docosahexaenoic acid (DHA), have shown to exert beneficial effects on nonalcoholic fatty liver disease (NAFLD), which is characterized by the excessive accumulation of lipids and chronic injury in the liver. However, the effect of endogenous DHA biosynthesis on the lipid homeostasis of liver is poorly understood. In this study, we used a DHA biosynthesis-deficient zebrafish model, elovl2 mutant, to explore the effect of endogenously biosynthesized DHA on hepatic lipid homeostasis. We found the pathways of lipogenesis and lipid uptake were strongly activated, while the pathways of lipid oxidation and lipid transport were inhibited in the liver of elovl2 mutants, leading to lipid droplet accumulation in the mutant hepatocytes and NAFLD. Furthermore, the elovl2 mutant hepatocytes exhibited disrupted mitochondrial structure and function, activated endoplasmic reticulum stress, and hepatic injury. We further unveiled that the hepatic cell death and injury was mainly mediated by ferroptosis, rather than apoptosis, in elovl2 mutants. Elevating DHA content in elovl2 mutants, either by the introduction of an omega-3 desaturase (fat1) transgene or by feeding with a DHA-rich diet, could strongly alleviate NAFLD features and ferroptosis-mediated hepatic injury. Together, our study elucidates the essential role of endogenous DHA biosynthesis in maintaining hepatic lipid homeostasis and liver health, highlighting that DHA deficiency can lead to NAFLD and ferroptosis-mediated hepatic injury.

Biosynthetic mechanisms of omega-3 polyunsaturated fatty acids in microalgae.

Marine microalgae are the primary producers of ω3 polyunsaturated fatty acids (PUFAs), such as octadecapentaenoic acid (OPA, 18:5n-3) and docosahexaenoic acid (DHA, 22:6n-3) for food chains. However, the biosynthetic mechanisms of these PUFAs in the algae remain elusive. To study how these fatty acids are synthesized in microalgae, a series of radiolabeled precursors were used to trace the biosynthetic process of PUFAs in Emiliania huxleyi. Feeding the alga with 14C-labeled acetic acid in a time course showed that OPA was solely found in glycoglycerolipids such as monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) synthesized plastidically by sequential desaturations while DHA was exclusively found in phospholipids synthesized extraplastidically. Feeding the alga with 14C-labeled α-linolenic acid (ALA), linoleic acid (LA) and oleic acid (OA) showed that DHA was synthesized extraplastidically from fed ALA and LA, but not from OA, implying that the aerobic pathway of DHA biosynthesis is incomplete with missing a Δ12 desaturation step. The in vitro enzymatic assays with 14C-labeled malonyl-CoA showed that DHA was synthesized from acetic acid by a PUFA synthase. These results provide the first and conclusive biochemistry evidence that OPA is synthesized by a plastidic aerobic pathway through sequential desaturations with the last step of Δ3 desaturation, while DHA is synthesized by an extraplastidic anaerobic pathway catalyzed by a PUFA synthase in the microalga.

Omega-3 Fatty Acids and Arrhythmias.

The pro- and antiarrhythmic effects of omega-3 polyunsaturated fatty acids (n-3 PUFAs) have been extensively studied in preclinical and human trials. Despite early evidence of an antiarrhythmic role of n-3 PUFA in the prevention of sudden cardiac death and postoperative and persistent atrial fibrillation (AF), subsequent well-designed randomized trials have largely not shown an antiarrhythmic benefit. Two trials that tested moderate and high-dose n-3 PUFA demonstrated a reduction in sudden cardiac death, but these findings have not been widely replicated, and the potential of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) to reduce arrhythmic death in combination, or as monotherapy, remains uncertain. The accumulated clinical evidence does not support supplementation of n-3 PUFA for postoperative AF or secondary prevention of AF. Several large, contemporary, randomized controlled trials of high-dose n-3 PUFA for primary or secondary cardiovascular prevention have demonstrated a small, significant, dose-dependent increased risk of incident AF compared with mineral oil or corn oil comparator. These findings were reproduced with both icosapent ethyl monotherapy and a mixed EPA+DHA formulation. The proarrhythmic mechanism of increased AF in contemporary cohorts exposed to high-dose n-3 PUFA is unknown. EPA and DHA and their metabolites have pleiotropic cardiometabolic and pro- and antiarrhythmic effects, including modification of the lipid raft microenvironment; alteration of cell membrane structure and fluidity; modulation of sodium, potassium, and calcium currents; and regulation of gene transcription, cell proliferation, and inflammation. Further characterization of the complex association between EPA, EPA+DHA, and DHA and AF is needed. Which formulations, dose ranges, and patient subgroups are at highest risk, remain unclear.

Delta-5 elongase knockout reduces docosahexaenoic acid and lipid synthesis and increases heat sensitivity in a diatom.

Recent global marine lipidomic analysis reveals a strong relationship between ocean temperature and phytoplanktonic abundance of omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs), especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are essential for human nutrition and primarily sourced from phytoplankton in marine food webs. In phytoplanktonic organisms, EPA may play a major role in regulating the phase transition temperature of membranes, while the function of DHA remains unexplored. In the oleaginous diatom Phaeodactylum tricornutum, DHA is distributed mainly on extraplastidial phospholipids, which is very different from the EPA enriched in thylakoid lipids. Here, CRISPR/Cas9-mediated knockout of delta-5 elongase (ptELO5a), which encodes a delta-5 elongase (ELO5) catalyzing the elongation of EPA to synthesize DHA, led to a substantial interruption of DHA synthesis in P. tricornutum. The ptELO5a mutants showed some alterations in transcriptome and glycerolipidomes, including membrane lipids and triacylglycerols under normal temperature (22°C), and were more sensitive to elevated temperature (28°C) than wild type. We conclude that PtELO5a-mediated synthesis of small amounts of DHA has indispensable functions in regulating membrane lipids, indirectly contributing to storage lipid accumulation, and maintaining thermomorphogenesis in P. tricornutum. This study also highlights the significance of DHA synthesis and lipid composition for environmental adaptation of P. tricornutum.

Fish and Omega-3 Fatty Acids: Sex and Racial Differences in Cardiovascular Outcomes and Cognitive Function.

Both cardiovascular disease (CVD) and cognitive decline are common features of aging. One in 5 deaths is cardiac for both men and women in the United States, and an estimated 50 million are currently living with dementia worldwide. In this review, we summarize sex and racial differences in the role of fish and its very long chain omega-3 polyunsaturated fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), in preventing CVD events and cognitive decline. In prospective studies, women with higher nonfried and fatty fish intake and women and Black individuals with higher plasma levels of EPA and DHA had a lower risk of CVD. In randomized controlled trials of EPA and DHA supplementation in primary CVD prevention, Black subjects benefited in a secondary outcome. In secondary CVD prevention, both men and women benefited, and Asians benefited as a prespecified subgroup. Fish and omega-3 polyunsaturated fatty acids are associated with prevention of cognitive decline in prospective studies. In randomized controlled trials of EPA and DHA supplementation, women have cognitive benefit. DHA seems more beneficial than EPA, and supplementation is more beneficial when started before cognitive decline. Although studies in women and racial groups are limited, life-long intake of nonfried and fatty fish lowers the risk of CVD and cognitive decline, and randomized controlled trials also show the benefit of EPA and DHA supplementation. These findings should be factored into recommendations for future research and clinical recommendations as dietary modalities could be cost-effective for disease prevention.

DHA-containing phospholipids control membrane fusion and transcellular tunnel dynamics.

Metabolic studies and animal knockout models point to the critical role of polyunsaturated docosahexaenoic acid (22:6, DHA)-containing phospholipids (DHA-PLs) in physiology. Here, we investigated the impact of DHA-PLs on the dynamics of transendothelial cell macroapertures (TEMs) triggered by RhoA inhibition-associated cell spreading. Lipidomic analyses showed that human umbilical vein endothelial cells (HUVECs) subjected to a DHA diet undergo a 6-fold enrichment in DHA-PLs at the plasma membrane (PM) at the expense of monounsaturated oleic acid-containing PLs (OA-PLs). Consequently, DHA-PL enrichment at the PM induces a reduction in cell thickness and shifts cellular membranes towards a permissive mode of membrane fusion for transcellular tunnel initiation. We provide evidence that a global homeostatic control of membrane tension and cell cortex rigidity minimizes overall changes of TEM area through a decrease of TEM size and lifetime. Conversely, low DHA-PL levels at the PM lead to the opening of unstable and wider TEMs. Together, this provides evidence that variations of DHA-PL levels in membranes affect cell biomechanical properties.

DHA and EPA inhibit porcine coronavirus replication by alleviating ER stress.

IMPORTANCE: Porcine epidemic diarrhea caused by porcine coronaviruses remains a major threat to the global swine industry. Fatty acids are extensively involved in the whole life of the virus. In this study, we found that docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) significantly reduced the viral load of porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and porcine delta coronavirus (PDCoV) and acted on the replication of the viruses rather than attachment and entry. We further confirmed that DHA and EPA inhibited PEDV replication by alleviating the endoplasmic reticulum stress. Meanwhile, DHA and EPA alleviate PEDV-induced inflammation and reactive oxygen species (ROS) levels and enhance the cellular antioxidant capacity. These data indicate that DHA and EPA have antiviral effects on porcine coronaviruses and provide a molecular basis for the development of new fatty acid-based therapies to control porcine coronavirus infection and transmission.

Nonalcoholic Fatty Liver Disease and Omega-3 Fatty Acids: Mechanisms and Clinical Use.

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic fatty liver disease worldwide, particularly in obese and type 2 diabetic individuals. Currently, there are no therapies for NAFLD that have been approved by the US Food and Drug Administration. Herein, we examine the rationale for using ω3 polyunsaturated fatty acids (PUFAs) in NAFLD therapy. This focus is based on the finding that NAFLD severity is associated with a reduction of hepatic C20-22 ω3 PUFAs. Because C20-22 ω3 PUFAs are pleiotropic regulators of cell function, loss of C20-22 ω3 PUFAs has the potential to significantly impact hepatic function. We describe NAFLD prevalence and pathophysiology as well as current NAFLD therapies. We also present evidence from clinical and preclinical studies that evaluated the capacity of C20-22 ω3 PUFAs to treat NAFLD. Given the clinical and preclinical evidence, dietary C20-22 ω3 PUFA supplementation has the potential to decrease human NAFLD severity by reducing hepatosteatosis and liver injury.

Plasma Levels of Polyunsaturated Fatty Acids and Adverse Kidney Outcomes.

RATIONALE & OBJECTIVE: Many studies have reported polyunsaturated fatty acids (PUFA) as significant predictors of cardiovascular disease, but little is known about the relationship between PUFA levels and chronic kidney disease (CKD). This study explored this relationship among individuals with and without CKD. STUDY DESIGN: Prospective observational cohort study. SETTING & PARTICIPANTS: 73,419 participants without CKD (cohort 1) and 6,735 participants with CKD (cohort 2) in the UK Biobank Study, with PUFA levels measured between 2007 and 2010. EXPOSURE: Percentage of plasma PUFA, omega-3 fatty acid (FA), omega-6 FA, docosahexaenoic acid (DHA), and linoleic acid relative to total FA. OUTCOME: Incident CKD for cohort 1 and incident kidney failure requiring replacement therapy (KFRT) for cohort 2. ANALYTICAL APPROACH: Cox proportional hazards regression analyses, including a cause-specific competing risk model. RESULTS: In cohort 1, individuals with higher quartiles of plasma PUFA levels had healthier lifestyles and fewer comorbidities. During 841,007 person-years of follow-up (median 11.9 years), incident CKD occurred in 4.5% of participants (incidence rate, 39.1 per 10,000 person-years). For incident CKD in cohort 1, the adjusted cause-specific hazard ratios for quartiles 2, 3, and 4 were 0.83 (95% CI, 0.75-0.92), 0.85 (95% CI, 0.76-0.96), 0.71 (95% CI, 0.62-0.82), respectively, compared with quartile 1. This inverse relationship was consistently observed for all PUFA types. In cohort 2, although total PUFA levels were not associated with KFRT, higher PUFA subtype levels of DHA were associated with a lower risk of KFRT. LIMITATIONS: Observational design and limited generalizability to individuals with higher disease severity; no data on eicosapentaenoic acid. CONCLUSIONS: Among individuals without CKD, higher plasma PUFA levels and all 4 PUFA components were associated with a lower risk of incident CKD. In individuals with CKD, only the omega-3 component of PUFA, DHA, was associated with a lower risk of KFRT. PLAIN-LANGUAGE SUMMARY: Low amounts of polyunsaturated fatty acids (PUFA) in the blood are suspected of increasing the chances of heart disease, but it is not known whether the PUFA relates to kidney disease occurrence. In a large group without kidney disease in the United Kingdom, people with higher levels of PUFA in their blood tended to have a lower risk of developing kidney disease compared to those with lower PUFA levels. This relationship was consistently observed for all PUFA types. However, in the group with kidney disease, only higher levels of docosahexaenoic acid, a subtype of PUFAs, were associated with a lower risk of developing severe kidney problems that required kidney replacement therapy. These findings suggest that higher levels of PUFA, found in certain healthy fats, might protect against the development of kidney disease in the general population. As kidney function declines, only the docosahexaenoic acid, a subtype of PUFA, appears to be associated with preserved kidney function.

Docosahexaenoic acid enhances the treatment efficacy for castration-resistant prostate cancer by inhibiting autophagy through Atg4B inhibition.

Autophagy induction in cancer is involved in cancer progression and the acquisition of resistance to anticancer agents. Therefore, autophagy is considered a potential therapeutic target in cancer therapy. In this study, we found that long-chain fatty acids (LCFAs) have inhibitory effects on Atg4B, which is essential for autophagosome formation, through screening based on the pharmacophore of 21f, a recently developed Atg4B inhibitor. Among these fatty acids, docosahexaenoic acid (DHA), a polyunsaturated fatty acid, exhibited the most potent Atg4B inhibitory activity. DHA inhibited autophagy induced by androgen receptor signaling inhibitors (ARSI) in LNCaP and 22Rv1 prostate cancer cells and significantly increased apoptotic cell death. Furthermore, we investigated the effect of DHA on resistance to ARSI by establishing darolutamide-resistant prostate cancer 22Rv1 (22Rv1/Dar) cells, which had developed resistance to darolutamide, a novel ARSI. At baseline, 22Rv1/Dar cells showed a higher autophagy level than parental 22Rv1 cells. DHA significantly suppressed Dar-induced autophagy and sensitized 22Rv1/Dar cells by inducing apoptotic cell death through mitochondrial dysfunction. These results suggest that DHA supplementation may improve prostate cancer therapy with ARSI.

Docosahexaenoic Acid Controls Pulmonary Macrophage Lipid Raft Size and Inflammation.

BACKGROUND: Docosahexaenoic acid (DHA) controls the biophysical organization of plasma membrane sphingolipid/cholesterol-enriched lipid rafts to exert anti-inflammatory effects, particularly in lymphocytes. However, the impact of DHA on the spatial arrangement of alveolar macrophage lipid rafts and inflammation is unknown. OBJECTIVES: The primary objective was to determine how DHA controls lipid raft organization and function of alveolar macrophages. As proof-of-concept, we also investigated DHA's anti-inflammatory effects on select pulmonary inflammatory markers with a murine influenza model. METHODS: MH-S cells, an alveolar macrophage line, were treated with 50 µM DHA or vehicle control and were used to study plasma membrane molecular organization with fluorescence-based methods. Biomimetic membranes and coarse grain molecular dynamic (MD) simulations were employed to investigate how DHA mechanistically controls lipid raft size. qRT-PCR, mass spectrometry, and ELISAs were used to quantify downstream inflammatory signaling transcripts, oxylipins, and cytokines, respectively. Lungs from DHA-fed influenza-infected mice were analyzed for specific inflammatory markers. RESULTS: DHA increased the size of lipid rafts while decreasing the molecular packing of the MH-S plasma membrane. Adding a DHA-containing phospholipid to a biomimetic lipid raft-containing membrane led to condensing, which was reversed with the removal of cholesterol. MD simulations revealed DHA nucleated lipid rafts by driving cholesterol and sphingomyelin into rafts. Downstream of the plasma membrane, DHA lowered the concentration of select inflammatory transcripts, oxylipins, and IL-6 secretion. DHA lowered pulmonary Il6 and Tnf-α mRNA expression and increased anti-inflammatory oxylipins of influenza-infected mice. CONCLUSIONS: The data suggest a model in which the localization of DHA acyl chains to nonrafts is driving sphingomyelin and cholesterol molecules into larger lipid rafts, which may serve as a trigger to impede signaling and lower inflammation. These findings also identify alveolar macrophages as a target of DHA and underscore the anti-inflammatory properties of DHA for lung inflammation.

How Membrane Phospholipids Containing Long-Chain Polyunsaturated Fatty Acids and Their Oxidation Products Orchestrate Lipid Raft Dynamics to Control Inflammation.

BACKGROUND: Long-chain PUFA (LC-PUFA) influence varying aspects of inflammation. One mechanism by which they regulate inflammation is by controlling the size and molecular composition of lipid rafts. Lipid rafts are sphingolipid/cholesterol-enriched plasma membrane microdomains that compartmentalize signaling proteins and thereby control downstream inflammatory gene expression and cytokine production. OBJECTIVES: This review summarizes developments in our understanding of how LC-PUFA acyl chains of phospholipids, in addition to oxidized derivatives of LC-PUFAs such as oxidized 1-palmitoyl-2-arachidonyl-phosphatidylcholine (oxPAPC), manipulate formation of lipid rafts and thereby inflammation. METHODS: We reviewed the literature, largely from the past 2 decades, on the impact of LC-PUFA acyl chains and oxidized products of LC-PUFAs on lipid raft biophysical organization of myeloid and lymphoid cells. The majority of the studies are based on rodent or cellular experiments with supporting mechanistic studies using biomimetic membranes and molecular dynamic simulations. These studies have focused largely on the LC-PUFA docosahexaenoic acid, with some studies addressing eicosapentaenoic acid. A few studies have investigated the role of oxidized phospholipids on rafts. RESULTS: The biophysical literature suggests a model in which n-3 LC-PUFAs, in addition to oxPAPC, localize predominately to nonraft regions and impart a disordering effect in this environment. Rafts become larger because of the ensuing increase in the difference in order between raft and nonrafts. Biochemical studies suggest that some n-3 LC-PUFAs can be found within rafts. This deviation from homeostasis is a potential trigger for controlling aspects of innate and adaptive immunity. CONCLUSION: Overall, select LC-PUFA acyl chains and oxidized acyl chains of phospholipids control lipid raft dynamics and downstream inflammation. Gaps in knowledge remain, particularly on underlying molecular mechanisms by which plasma membrane receptor organization is controlled in response to oxidized LC-PUFA acyl chains of membrane phospholipids. Validation in humans is also an area for future study.

Lipidomic and Fatty Acid Biomarkers in Whole Blood Can Predict the Dietary Intake of Eicosapentaenoic and Docosahexaenoic Acids in a Danish Population.

BACKGROUND: The intake of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been associated with health benefits. Blood levels of these fatty acids, measured by gas chromatography (GC), are associated with their dietary intake, but the relationships with lipidomic measurements are not well defined. OBJECTIVES: This study aimed to determine the lipidomic biomarkers in whole blood that predict intakes of EPA + DHA and examine the relationship between lipidomic and GC-based n-3 polyunsaturated fatty acid (n-3 PUFA) biomarkers. METHODS: Lipidomic and fatty acid analyses were completed on 120 whole blood samples collected from Danish participants. Dietary intakes were completed using a web-based 7-d food diary. Stepwise multiple linear regression was used to identify the fatty acid and lipidomic variables that predict intakes of EPA + DHA and to determine lipidomic species that predict commonly used fatty acid biomarkers. RESULTS: Stepwise regression selected lipidomic variables with an R2 = 0.52 for predicting EPA + DHA intake compared to R2 = 0.40 for the selected fatty acid GC-based variables. More predictive models were generated when the lipidomic variables were selected for females only (R2 = 0.62, n = 68) and males only (R2 = 0.72, n = 52). Phosphatidylethanolamine plasmalogen species containing EPA or DHA tended to be the most predictive lipidomic variables. Stepwise regression also indicated that selected lipidomic variables can predict commonly used fatty acid GC-based n-3 PUFA biomarkers as the R2 values ranged from 0.84 to 0.91. CONCLUSIONS: Both fatty acid and lipidomic data can be used to predict EPA + DHA intakes, and fatty acid GC-based biomarkers can be emulated by lipidomic species. Lipidomic-based biomarkers appear to be influenced by sex differences, probably in n-3 PUFA and lipoprotein metabolism. These results improve our ability to understand the relationship between novel lipidomic data and GC fatty acid data and will increase our ability to apply lipidomic methods to fatty acid and lipid nutritional research.

Unsaturated fatty acids uniquely alter aggregation rate of α-synuclein and insulin and change the secondary structure and toxicity of amyloid aggregates formed in their presence.

Docosahexaenoic (DHA) and arachidonic acids (ARA) are omega-3 and omega-6 long-chain polyunsaturated fatty acids (LCPUFAs). These molecules constitute a substantial portion of phospholipids in plasma membranes. Therefore, both DHA and ARA are essential diet components. Once consumed, DHA and ARA can interact with a large variety of biomolecules, including proteins such as insulin and α-synuclein (α-Syn). Under pathological conditions known as injection amyloidosis and Parkinson's disease, these proteins aggregate forming amyloid oligomers and fibrils, toxic species that exert high cell toxicity. In this study, we investigate the role of DHA and ARA in the aggregation properties of α-Syn and insulin. We found that the presence of both DHA and ARA at the equimolar concentrations strongly accelerated aggregation rates of α-Syn and insulin. Furthermore, LCPUFAs substantially altered the secondary structure of protein aggregates, whereas no noticeable changes in the fibril morphology were observed. Nanoscale Infrared analysis of α-Syn and insulin fibrils grown in the presence of both DHA and ARA revealed the presence of LCPUFAs in these aggregates. We also found that such LCPUFAs-rich α-Syn and insulin fibrils exerted significantly greater toxicities compared to the aggregates grown in the LCPUFAs-free environment. These findings show that interactions between amyloid-associated proteins and LCPUFAs can be the underlying molecular cause of neurodegenerative diseases.

Long chain acyl-CoA synthetase 6 facilitates the local distribution of di-docosahexaenoic acid- and ultra-long-chain-PUFA-containing phospholipids in the retina to support normal visual function in mice.

Docosahexaenoic acid (DHA) and ultra-long-chain polyunsaturated fatty acids (ULC-PUFAs) are uniquely enriched in membrane phospholipids of retinal photoreceptors. Several studies have shown that di-DHA- and ULC-PUFA-containing phospholipids in photoreceptors have an important role in maintaining normal visual function; however, the molecular mechanisms underlying the synthesis and enrichment of these unique lipids in the retina, and their specific roles in retinal function remain unclear. Long-chain acyl-coenzyme A (CoA) synthetase 6 (ACSL6) preferentially converts DHA into DHA-CoA, which is a substrate during DHA-containing lipid biosynthesis. Here, we report that Acsl6 mRNA is expressed in the inner segment of photoreceptor cells and the retinal pigment epithelial cells, and genetic deletion of ACSL6 resulted in the selective depletion of di-DHA- and ULC-PUFA-containing phospholipids, but not mono-DHA-containing phospholipids in the retina. MALDI mass spectrometry imaging (MALDI-MSI) revealed the selective distribution of di-DHA- and ULC-PUFA-containing phospholipids in the photoreceptor outer segment (OS). Electroretinogram of Acsl6-/- mice exhibited photoreceptor cell-derived visual impairment, whereas the expression levels and localization of opsin proteins were unchanged. Acsl6-/- mice exhibited an age-dependent progressive decrease of the thickness of the outer nuclear layers, whereas the inner nuclear layers and OSs were normal. These results demonstrate that ACSL6 facilitates the local enrichment of di-DHA- and ULC-PUFA-containing phospholipids in the retina, which supports normal visual function and retinal homeostasis.

Genome-scale metabolic modeling of Thraustochytrium sp. RT2316-16: Effects of nutrients on metabolism.

Marine thraustochytrids produce metabolically important lipids such as the long-chain omega-3 polyunsaturated fatty acids, carotenoids, and sterols. The growth and lipid production in thraustochytrids depends on the composition of the culture medium that often contains yeast extract as a source of amino acids. This work discusses the effects of individual amino acids provided in the culture medium as the only source of nitrogen, on the production of biomass and lipids by the thraustochytrid Thraustochytrium sp. RT2316-16. A reconstructed metabolic network based on the annotated genome of RT2316-16 in combination with flux balance analysis was used to explain the observed growth and consumption of the nutrients. The culture kinetic parameters estimated from the experimental data were used to constrain the flux via the nutrient consumption rates and the specific growth rate of the triacylglycerol-free biomass in the genome-scale metabolic model (GEM) to predict the specific rate of ATP production for cell maintenance. A relationship was identified between the specific rate of ATP production for maintenance and the specific rate of glucose consumption. The GEM and the derived relationship for the production of ATP for maintenance were used in linear optimization problems, to successfully predict the specific growth rate of RT2316-16 in different experimental conditions.

Transcriptomics aids in uncovering the metabolic shifts and molecular machinery of Schizochytrium limacinum during biotransformation of hydrophobic substrates to docosahexaenoic acid.

BACKGROUND: Biotransformation of waste oil into value-added nutraceuticals provides a sustainable strategy. Thraustochytrids are heterotrophic marine protists and promising producers of omega (ω) fatty acids. Although the metabolic routes for the assimilation of hydrophilic carbon substrates such as glucose are known for these microbes, the mechanisms employed for the conversion of hydrophobic substrates are not well established. Here, thraustochytrid Schizochytrium limacinum SR21 was investigated for its ability to convert oils (commercial oils with varying fatty acid composition and waste cooking oil) into ω-3 fatty acid; docosahexaenoic acid (DHA). RESULTS: Within 72 h SR21 consumed ~ 90% of the oils resulting in enhanced biomass (7.5 g L- 1) which was 2-fold higher as compared to glucose. Statistical analysis highlights C16 fatty acids as important precursors of DHA biosynthesis. Transcriptomic data indicated the upregulation of multiple lipases, predicted to possess signal peptides for secretory, membrane-anchored and cytoplasmic localization. Additionally, transcripts encoding for mitochondrial and peroxisomal ß-oxidation along with acyl-carnitine transporters were abundant for oil substrates that allowed complete degradation of fatty acids to acetyl CoA. Further, low levels of oxidative biomarkers (H2O2, malondialdehyde) and antioxidants were determined for hydrophobic substrates, suggesting that SR21 efficiently mitigates the metabolic load and diverts the acetyl CoA towards energy generation and DHA accumulation. CONCLUSIONS: The findings of this study contribute to uncovering the route of assimilation of oil substrates by SR21. The thraustochytrid employs an intricate crosstalk among the extracellular and intracellular molecular machinery favoring energy generation. The conversion of hydrophobic substrates to DHA can be further improved using synthetic biology tools, thereby providing a unique platform for the sustainable recycling of waste oil substrates.