Accumulation Of Docosahexaenoic Acid Research Articles

Regulatory mechanisms of autophagy on DHA and carotenoid accumulation in Crypthecodinium sp. SUN

BackgroundAutophagy is a crucial process of cellular self-destruction and component reutilization that can affect the accumulation of total fatty acids (TFAs) and carotenoids in microalgae. The regulatory effects of autophagy process in a docosahexaenoic acid (DHA) and carotenoids simultaneously producing microalga, Crypthecodinium sp. SUN, has not been studied. Thus, the autophagy inhibitor (3-methyladenine (MA)) and activator (rapamycin) were used to regulate autophagy in Crypthecodinium sp. SUN.ResultsThe inhibition of autophagy by 3-MA was verified by transmission electron microscopy, with fewer autophagy vacuoles observed. Besides, 3-MA reduced the glucose absorption and intracellular acetyl-CoA level, which resulting in the decrease of TFA and DHA levels by 15.83 and 26.73% respectively; Surprisingly, 3-MA increased intracellular reactive oxygen species level but decreased the carotenoids level. Comparative transcriptome analysis showed that the downregulation of the glycolysis, pentose phosphate pathway and tricarboxylic acid cycle may underlie the decrease of acetyl-CoA, NADPH and ATP supply for fatty acid biosynthesis; the downregulation of PSY and HMGCR may underlie the decreased carotenoids level. In addition, the class I PI3K-AKT signaling pathway may be crucial for the regulation of carbon and energy metabolism. At last, rapamycin was used to activate autophagy, which significantly enhanced the cell growth and TFA level and eventually resulted in 1.70-fold increase in DHA content.ConclusionsOur findings indicate the mechanisms of autophagy in Crypthecodinium sp. SUN and highlight a way to manipulate cell metabolism by regulating autophagy. Overall, this study provides valuable insights to guide further research on autophagy-regulated TFA and carotenoids accumulation in Crypthecodinium sp. SUN.

Influence of Dietary Triacylglycerol Structure on the Accumulation of Docosahexaenoic Acid [22:6(n-3)] in Organs in a Short-Term Feeding Trial with Mildly Omega-3 Deficient Rats.

To study the effect of positional distribution of docosahexaenoic acid (DHA) in dietary triacylglycerols (TAG) on the tissue fatty acid content and composition of mildly (n-3) deficient rats. In a 5-day feeding trial, mildly (n-3) deficient rats received 360mg daily structured TAGs: sn-22:6(n-3)-18:0-18:0, sn-18:0-18:0-22:6(n-3), sn-18:0-22:6(n-3)-18:0, or tristearin. A fifth group receives standard (n-3) adequate feed AIN-93G from birth till the end of the trial. The DHA-fed groups show significantly higher DHA levels in the liver and visceral fat compared to the tristearin or normal feed groups showing that the dose and the short feeding period of DHA were sufficient to restore the DHA content in the organs of (n-3) deficient rats. Feeding sn-1 DHA resulted in higher levels of DHA in the liver TAG compared to sn-3 DHA feeding, although the difference did not reach statistical significance. These findings indicated a possible difference in the tissue accumulation and/or metabolic fate of DHA from the sn-1 and sn-3 positions of TAG.

Knockout of a PLD gene in Schizochytrium limacinum SR21 enhances docosahexaenoic acid accumulation by modulation of the phospholipid profile

BackgroundThe hydrolysis and transphosphatidylation of phospholipase D (PLD) play important roles in the interconversion of phospholipids (PLs), which has been shown to profoundly impact lipid metabolism in plants. In this study, the effect of the PLD1 gene of Schizochytrium limacinum SR21 (S. limacinum SR21) on lipid metabolism was investigated.ResultsPLD1 knockout had little impact on cell growth and lipid production, but it significantly improved the percentage of polyunsaturated fatty acids in lipids, of which docosahexaenoic acid (DHA) content increased by 13.3% compared to the wild-type strain. Phospholipomics and real-time quantitative PCR analysis revealed the knockout of PLD1 reduced the interexchange and increased de novo synthesis of PLs, which altered the composition of PLs, accompanied by a final decrease in phosphatidylcholine (PC) and an increase in phosphatidylinositol, lysophosphatidylcholine, and phosphatidic acid levels. PLD1 knockout also increased DHA content in triglycerides (TAGs) and decreased it in PLs.ConclusionsThese results indicate that PLD1 mainly performs the transphosphatidylation activity in S. limacinum SR21, and its knockout promotes the migration of DHA from PLs to TAGs, which is conducive to DHA accumulation and storage in TAGs via an acyl CoA-independent pathway. This study provides a novel approach for identifying the mechanism of DHA accumulation and metabolic regulation strategies for DHA production in S. limacinum SR21.

Enhancing docosahexaenoic acid production in Aurantiochytrium species using atmospheric and room temperature plasma mutagenesis and comprehensive multi-omics analysis

Aurantiochytrium sp. belongs to marine heterotrophic single-cell protist, which is an important decomposer in marine ecosystem. Aurantiochytrium sp. has gained notoriety because of its ability to accumulate high-value docosahexaenoic acid (DHA), but the key factors of DHA synthesis were unclear at present. In this study, Atmospheric and Room Temperature Plasma technology was applied to the mutagenic breeding of Aurantiochytrium sp., and transcriptomics and proteomics were adopted to analyze the DHA-biosynthesis mechanism. According to the growth and DHA accumulation profiles, the mutant strain Aurantiochytrium sp. R2A35 was selected. The DHA content in total lipids was greatly improved from 49.39 % of the wild strain R2 to 63.69 % of the mutant strain. Moreover, the DHA content in the biomass of Aurantiochytrium sp. R2A35 as 39.72 % was the highest DHA productivity reported so far. The differentially expressed genes distinguished from transcriptome and the TMT-identified differential proteins distinguished from proteome confirmed that the expression of acetyl-CoA carboxylase and ketoacyl reductase was up-regulated by 4.78-fold and 6.95-fold, respectively and the fatty acid synthase was concurrently down-regulated by 2.79-fold, so that more precursor was transported to the polyketide synthase pathway, thereby increasing the DHA yield in Aurantiochytrium sp. R2A35. This research would provide reference for the DHA metabolism process and contribute to the understanding of the decomposer - Aurantiochytrium sp. in marine ecosystems.

Knockdown of Placental Major Facilitator Superfamily Domain Containing 2a in Pregnant Mice Reduces Fetal Brain Growth and Phospholipid Docosahexaenoic Acid Content

Introduction: Docosahexaenoic acid (DHA) is an n-3 long chain polyunsaturated fatty acid critical for fetal brain development that is transported to the fetus from the mother by the placenta. The lysophosphatidylcholine (LPC) transporter, Major Facilitator Superfamily Domain Containing 2a (MFSD2a), is localized in the basal plasma membrane of the syncytiotrophoblast of the human placenta, and MFSD2a expression correlates with umbilical cord blood LPC-DHA levels in human pregnancy. We hypothesized that placenta-specific knockdown of MFSD2a in pregnant mice reduces phospholipid DHA accumulation in the fetal brain. Methods: Mouse blastocysts (E3.5) were transduced with an EGFP-expressing lentivirus containing either an shRNA targeting MFSD2a or a non-coding sequence (SCR), then transferred to pseudopregnant females. At E18.5, fetuses were weighed and their placenta, brain, liver and plasma were collected. MFSD2a mRNA expression was determined by qPCR in the brain, liver and placenta and phospholipid DHA was quantified by LC-MS/MS. Results: MFSD2a-targeting shRNA reduced placental mRNA MFSD2a expression by 38% at E18.5 (n = 45, p < 0.008) compared with SCR controls. MFSD2a expression in the fetal brain and liver were unchanged. Fetal brain weight was reduced by 13% (p = 0.006). Body weight, placenta and liver weights were unaffected. Fetal brain phosphatidyl choline and phosphatidyl ethanolamine DHA content was lower in fetuses with placenta-specific MFSD2a knockdown. Conclusions: Placenta-specific reduction in expression of the LPC-DHA transporter MFSD2a resulted in reduced fetal brain weight and lower phospholipid DHA content in the fetal brain. These data provide mechanistic evidence that placental MFSD2a mediates maternal–fetal transfer of LPC-DHA, which is critical for brain growth.

Low and high temperatures promote docosahexaenoic acid accumulation in Crypthecodinium sp. SUN by regulating the polyunsaturated fatty acid synthase pathway and the expression of saturated fatty acid preferred diacylglycerol acyltransferases

Low (15 °C) and high (35 °C) temperatures significantly increased DHA as a percentage of total fatty acids (TFAs) to 43.6 % and 40.46 %, respectively (1.28- and 1.18-fold of that at 25 °C, respectively). The incompleteness of the FAS pathway indicates that DHA synthesis does not occur via this pathway. Meanwhile, Comparative transcriptome analysis showed that the PUFA synthase pathway might be responsible for DHA synthesis in C. sp. SUN. Additionally, the three diacylglycerol acyltransferases all had a substrate preference for saturated fatty acid (SFA)-CoA, which also contributed to the decreased SFA and increased DHA at both low and high temperatures. Additionally, WGCNA analysis identifies key regulatory genes that may be involved in temperature-regulated DHA proportion. The findings of this study indicate the mechanisms of temperature-regulated DHA accumulation in C. sp. SUN and shed light on the manipulation of DHA proportion by changes in temperature.

Simultaneous production of high-value lipids in Schizochytrium sp. by synergism of chemical modulators.

The study focuses on the simultaneous improvement of biomass, lipid, and docosahexaenoic acid (DHA) productivities in a single reactor using modulator control strategies. The efficacy of three different biochemical modulators, sesamol (Ses), 6-benzylaminopurine (6-BAP), and ethylenediaminetetraacetic acid (EDTA), as potential stimulants in augmenting the biomass, lipid, and DHA production of Schizochytrium sp. MTCC 5890 was elucidated. After 48h of cultivation, among tested modulators, the individual supplementation of 6-BAP and Ses showed improvement in biomass, lipid, and DHA accumulation by 28.2%, 56.1%, and 87.2% and 21.7%, 47.9%, and 91%, respectively, over the non-supplemented group. In addition, the cooperative effect of selected concentrations, i.e., 10 mgL-1 6-BAP and 200 mgL-1 Ses, further increased the productivities of biomass of 13.5 gL-1d-1 ± 0.66, lipid of 7.4 gL-1d-1 ± 0.69, and DHA of 3.2 gL-1d-1 ± 1.09 representing 8%, 39%, and 69% increase over the individual addition of 6-BAP or Ses, respectively, in batch culture. Supplementation with 6-BAP + Ses at 12h of time point eventually increased the lipid yield to 15.6 ± 0.42 gL-1 from 7.88 ± 0.31 gL-1 (control) and DHA yield to 6.4 ± 0.11 gL-1 from 2.23 ± 0.09 gL-1 (control), respectively. Furthermore, the process was optimized in continuous culture supplemented with 6-BAP + Ses for enhanced productivities. Continuous culture resulted in maximum biomass (2.04 ± 1.12 gL-1day-1), lipid (1.0 ± 0.73 gL-1day-1), and DHA (0.386 ± 0.22 gL-1day-1) productivities, which were higher as compared with the batch and fed-batch processes by 26 ± 1.21%, 22 ± 1.01%, and 21 ± 0.98% and 24 ± 0.45%, 16 ± 0.38%, and 14 ± 0.12%, respectively. This work represents the potential application of the combined effect of modulators for the simultaneous enhancement of biomass production and lipid and DHA productivities. KEY POINTS: • The cumulative study of 6-BAP and sesamol proved to be more efficient in the simultaneous production of biomass, lipid, and DHA in a single reactor. • Addition of a combination of 6-BAP + Ses remarkably increased the biomass, lipid, and DHA productivities in tandem in continuous culture.

Enhancing the biomass and docosahexaenoic acid-rich lipid accumulation of Schizochytrium sp. in propionate wastewater.

In order to find a more effective way to obtain docosahexaenoic acid (DHA) rich lipid from Schizochytrium sp., a widespread propionate wastewater (PW) is used. PW is a common industrial and domestic wastewater, and transforming it into valuable products is a potential treatment method. Schizochytrium sp. is a rapidly growing oleaginous organism, which has been used commercially for DHA production. Herein, PW is completely used for DHA production by Schizochytrium sp. by genetic engineering and fermentation optimization, which can alleviate the increasingly tense demand for water resources and environmental pollution caused by industrial wastewater. Firstly, the methylmalonyl-CoA mutase (MCM) was overexpressed in Schizochytrium sp. to enhance the metabolism of propionate, then the engineered strain of overexpressed MCM (OMCM) can effectively use propionate. Then, the effects of PW with different concentration of propionate were investigated, and results showed that OMCM can completely replace clean water with PW containing 5gL-1 propionate. Furthermore, in the fed-batch fermentation, the OMCM obtained the highest biomass of 113.4gL-1 and lipid yield of 64.4gL-1 in PW condition, which is 26.8% and 51.7% higher than that of wild type (WT) in PW condition. Moreover, to verify why overexpression of MCM can promote DHA and lipid accumulation, the comparative metabolomics, ATP production level, the antioxidant system, and the transcription of key genes were investigated. Results showed that ATP induced by PW condition could drive the synthesis of DHA, and remarkably improve the antioxidant capacity of cells by enhancing the carotenoids production. Therefore, PW can be used as an effective and economical substrate and water source for Schizochytrium sp. to accumulate biomass and DHA.

Revealing holistic metabolic responses associated with lipid and docosahexaenoic acid (DHA) production in Aurantiochytrium sp. SW1

Aurantiochytrium sp. SW1, a marine thraustochytrid, has been regarded as a potential candidate as a docosahexaenoic acid (DHA) producer. Even though the genomics of Aurantiochytrium sp. are available, the metabolic responses at a systems level are largely unknown. Therefore, this study aimed to investigate the global metabolic responses to DHA production in Aurantiochytrium sp. through transcriptome and genome-scale network-driven analysis. Of a total of 13,505 genes, 2527 differentially expressed genes (DEGs) were identified in Aurantiochytrium sp., unravelling the transcriptional regulations behinds lipid and DHA accumulation. The highest number of DEG were found for pairwise comparison between growth phase and lipid accumulating phase where a total of 1435 genes were down-regulated with 869 genes being up-regulated. These uncovered several metabolic pathways that contributing in DHA and lipid accumulation including amino acid and acetate metabolism which involve in the generation of crucial precursors. Upon applying network-driven analysis, hydrogen sulphide was found as potential reporter metabolite that could be associated with the genes related to acetyl-CoA synthesis for DHA production. Our findings suggest that the transcriptional regulation of these pathways is a ubiquitous feature in response to specific cultivation phases during DHA overproduction in Aurantiochytrium sp. SW1.

Engineering M1-derived nanovesicles loading with docosahexaenoic acid synergizes ferroptosis and immune activation for treating hepatocellular carcinoma

Ferroptosis represents an innovative strategy to overcome the resistance of traditional cancer therapeutic through lethal lipid peroxidation leading to immunogenic cell death. However, the inefficiency of ferroptosis inducers and mild immunogenicity restrict the further clinical applications. Herein, engineering exosome-mimic M1 nanovesicles (MNV) were prepared by serial extrusion of M1 macrophage and served as an efficient vehicle for docosahexaenoic acid (DHA) delivery. MNV loaded with DHA (MNV@DHA) could promote more DHA accumulation in tumor cells, depletion glutathione and reduction of lipid antioxidant glutathione peroxidase-4 facilitating the occurrence of ferroptosis. Furthermore, MNV were able to induce the polarization of M1 and repolarize M2 macrophages to activate tumor immune microenvironments. The activated immune cells would further trigger the ferroptosis of tumor cells. In a murine orthotopic hepatocellular carcinoma model, MNV@DHA could significantly target tumor tissues, increase the proportion of M1 macrophages and CD8+ T cells and lessen the infiltration of M2 macrophages. Accordingly, MNV@DHA characterized with positive feedback regulation between ferroptosis and immune activation exhibited the strongest in vivo therapeutic effect. The synergism of ferroptosis and immunomodulation based on the dietary polyunsaturated fatty acids and engineered exosome-mimic nanovesicles may serve as a promising modality to efficiently complement pharmacological approaches for cancer management.

Combination of bicarbonate and low temperature stress induces the biosynthesis of both arachidonic and docosahexaenoic acids in alkaliphilic microalgae Dunaliella salina HTBS

High bicarbonate levels and low temperature may have an impact on microalgae cultivation. However, changes in cellular composition in response to the combination of the above stresses are still poorly understood. In this study, the combined effects of bicarbonate and low temperature on biochemical changes in alkaliphilic microalgae Dunaliella salina HTBS were investigated. Comparing to the control condition of 25°C without bicarbonate, the cell density was increased from 0.69 to 1.18 in the treatment condition of 0.15 M bicarbonate and low temperature (16 °C) while the lipid\protein\carbohydrate contents were increased from 34.71% to 43.94%, 22.44% to 26.03%, 22.62% to 29.18%, respectively. Meanwhile, the PUFAs, arachidonic acid (AA) and docosahexaenoic acid (DHA) contents reached to 3.52% and 4.73% with the combination of low temperature and bicarbonate, respectively, whereas they were not detected when the cells were treated with single condition. Moreover, both the chlorophyll and carotenoid contents were also detected with increased profiles in the combined treatments. As a result, the maximum photochemical efficiency but not reduced non-photochemical quenching was strengthened, which enhanced the photosynthetic performance. Additionally, our results indicated that D. salina HTBS could acclimate to the combined stress by up-regulating the activity of SOD\CAT and reducing MDA content. These findings demonstrated that the addition of a certain bicarbonate under low temperature could effectively enhance the biomass production and accumulation of AA and DHA, which would benefit the development of the microalgae industry in value-added products.

Comparative transcriptomic and lipidomic analyses indicate that cold stress enhanced the production of the long C18-C22 polyunsaturated fatty acids in Aurantiochytrium sp.

Aurantiochytrium sp. belonging to Thraustochytrids are known for their capacity to produce long-chain polyunsaturated fatty acids (PUFAs). However, effects of cold stress accompanied with staged-temperature control on the fatty acid metabolism in Aurantiochytrium sp. were rarely studied. In this study, cold stress (15°C, 5°C) was applied for Aurantiochytrium sp., with the physiological responses (morphology, growth, fatty acid profiling) and gene expression related FA synthesis, lipid metabolism, and regulatory processes was observed. Results showed that there is a significant change for the lipid types under 5°C (251 species) and 15°C (97 species) treatment. The 5°C treatment was benefit for the C18–C22 PUFAs with the yield of docosahexaenoic acid (DHA) increased to 1.25 times. After incubation at 15°C, the accumulation of eicosadienoic acid (EA) (20:2) was increased to 2.00-fold. Based on transcriptomic and qPCR analysis, an increase in genes involved in fatty acid synthase (FAS) and polyketide synthase (PKS) pathways was observed under low-temperature treatment. With upregulation of 3-ketoacyl-CoA synthase (2.44-fold), ketoreductase (2.50-fold), and dTDP-glucose 4,6-Dehydratase (rfbB) (2.31-fold) involved in PKS pathway, the accumulation of DHA was enhanced under 5°C. While, FAS and fatty elongase 3 (ELO) involved in the FAS pathway were upregulated (1.55-fold and 2.45-fold, respectively) to accumulate PUFAs at 15°C. Additionally, glycerol-3-phosphate acyltransferase (GPAT), lysophospholipid acyltransferase (LPAT), phosphatidic acid phosphatase (PAP), phosphatidylserine synthase (PSS), and phosphatidylserine decarboxylase (PSD) involved in glycerophospholipid biosynthesis were upregulated at 5°C increasing the accumulation of phosphatidic acid (PA), phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and phosphatidylinositol (PI). However, glycolysis and the TCA cycle were inhibited under 5°C. This study provides a contribution to the application of two-staged temperature control in the Aurantiochytrium sp. fermentation for producing cold stress-enhancing PUFAs, in order to better understand the function of the key genes for future genetic engineering.

Mechanisms of Sodium-Acetate-Induced DHA Accumulation in a DHA-Producing Microalga, Crypthecodinium sp. SUN.

Docosahexaenoic acid (DHA) is an omega-3 polyunsaturated fatty acid (PUFA) that is critical for the intelligence and visual development of infants. Crypthecodinium is the first microalga approved by the Food and Drug Administration for DHA production, but its relatively high intracellular starch content restricts fatty acid accumulation. In this study, different carbon sources, including glucose (G), sodium acetate (S) and mixed carbon (M), were used to investigate the regulatory mechanisms of intracellular organic carbon distribution in Crypthecodinium sp. SUN. Results show that glucose favored cell growth and starch accumulation. Sodium acetate limited glucose utilization and starch accumulation but caused a significant increase in total fatty acid (TFA) accumulation and the DHA percentage. Thus, the DHA content in the S group was highest among three groups and reached a maximum (10.65% of DW) at 96 h that was 2.92-fold and 2.24-fold of that in the G and M groups, respectively. Comparative transcriptome analysis showed that rather than the expression of key genes in fatty acids biosynthesis, increased intracellular acetyl-CoA content appeared to be the key regulatory factor for TFA accumulation. Additionally, metabolome analysis showed that the accumulated DHA-rich metabolites of lipid biosynthesis might be the reason for the higher TFA content and DHA percentage of the S group. The present study provides valuable insights to guide further research in DHA production.

Using field evaluation and systematic iteration to rationalize the accumulation of omega-3 long-chain polyunsaturated fatty acids in transgenic Camelina sativa.

SummaryThe Brassicaceae Camelina sativa (gold of pleasure) is now an established niche crop and being used as a transgenic host for a range of novel seed traits. Most notable of these is the accumulation of omega‐3 long‐chain polyunsaturates such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), fatty acids normally only found in marine organisms. As part of continued efforts to optimize the accumulation of these non‐native fatty acids via seed‐specific expression of algal genes, a new series of iterative constructs was built and introduced into Camelina. Seed fatty acid composition was determined, and the presence of EPA and DHA was confirmed. To provide an additional level of evaluation, full environmental release was carried out on selected events, providing a real‐world gauntlet against which to assess the performance of these novel lines. Composition of the seed oil triacylglycerol was determined by mass spectrometry, allowing for conclusions as to the contribution of different activities to the final accumulation of EPA and DHA. Since these data were derived from field‐grown material, they also represent a robust demonstration of the stability of the omega‐3 LC‐PUFA trait in Camelina. We propose that field trialling should be routinely incorporated in the plant synthetic biology ‘design–build–test–learn’ cycle.

ARTP Mutagenesis of Schizochytrium sp. PKU#Mn4 and Clethodim-Based Mutant Screening for Enhanced Docosahexaenoic Acid Accumulation.

Schizochytrium species are one of the best oleaginous thraustochytrids for high-yield production of docosahexaenoic acid (DHA, 22:6). However, the DHA yields from most wild-type (WT) strains of Schizochytrium are unsatisfactory for large-scale production. In this study, we applied the atmospheric and room-temperature plasma (ARTP) tool to obtain the mutant library of a previously isolated strain of Schizochytrium (i.e., PKU#Mn4). Two rounds of ARTP mutagenesis coupled with the acetyl-CoA carboxylase (ACCase) inhibitor (clethodim)-based screening yielded the mutant A78 that not only displayed better growth, glucose uptake and ACCase activity, but also increased (54.1%) DHA content than that of the WT strain. Subsequent optimization of medium components and supplementation improved the DHA content by 75.5 and 37.2%, respectively, compared with that of mutant A78 cultivated in the unoptimized medium. Interestingly, the ACCase activity of mutant A78 in a medium supplemented with biotin, citric acid or sodium citrate was significantly greater than that in a medium without supplementation. This study provides an effective bioengineering approach for improving the DHA accumulation in oleaginous microbes.

Plant and algal lysophosphatidic acid acyltransferases increase docosahexaenoic acid accumulation at the sn-2 position of triacylglycerol in transgenic Arabidopsis seed oil.

Although docosahexaenoic acid (DHA), an important dietary omega-3 polyunsaturated fatty acid (PUFA), is at present primarily sourced from marine fish, bioengineered crops producing DHA may offer a more sustainable and cost-effective source. DHA has been produced in transgenic oilseed crops, however, DHA in seed oil primarily occupies the sn-1/3 positions of triacylglycerol (TAG) with relatively low amounts of DHA in the sn-2 position. To increase the amount of DHA in the sn-2 position of TAG and in seed oil, putative lysophosphatidic acid acyltransferases (LPAATs) were identified and characterized from the DHA-producing alga Schizochytrium sp. and from soybean (Glycine max). The affinity-purified proteins were confirmed to have LPAAT activity. Expression of the Schizochytrium or soybean LPAATs in DHA-producing Arabidopsis expressing the Schizochytrium PUFA synthase system significantly increased the total amount of DHA in seed oil. A novel sensitive band-selective heteronuclear single quantum coherence (HSQC) NMR method was developed to quantify DHA at the sn-2 position of glycerolipids. More than two-fold increases in sn-2 DHA were observed for Arabidopsis lines expressing Schizochytrium or soybean LPAATs, with one Schizochytrium LPAAT driving DHA accumulation in the sn-2 position to 61% of the total DHA. Furthermore, expression of a soybean LPAAT led to a redistribution of DHA-containing TAG species, with two new TAG species identified. Our results demonstrate that transgenic expression of Schizochytrium or soybean LPAATs can increase the proportion of DHA at the sn-2 position of TAG and the total amount of DHA in the seed oil of a DHA-accumulating oilseed plant. Additionally, the band-selective HSQC NMR method that we developed provides a sensitive and robust method for determining the regiochemistry of DHA in glycerolipids. These findings will benefit the advancement of sustainable sources of DHA via transgenic crops such as canola and soybean.

Towards an Optimized Fetal DHA Accretion: Differences on Maternal DHA Supplementation Using Phospholipids vs. Triglycerides during Pregnancy in Different Models.

Docosahexaenoic acid (DHA) supplementation during pregnancy has been recommended by several health organizations due to its role in neural, visual, and cognitive development. There are several fat sources available on the market for the manufacture of these dietary supplements with DHA. These fat sources differ in the lipid structure in which DHA is esterified, mainly phospholipids (PL) and triglycerides (TG) molecules. The supplementation of DHA in the form of PL or TG during pregnancy can lead to controversial results depending on the animal model, physiological status and the fat sources utilized. The intestinal digestion, placental uptake, and fetal accretion of DHA may vary depending on the lipid source of DHA ingested by the mother. The form of DHA used in maternal supplementation that would provide an optimal DHA accretion for fetal brain development, based on the available data obtained most of them from different animal models, indicates no consistent differences in fetal accretion when DHA is provided as TG or PL. Other related lipid species are under evaluation, e.g., lyso-phospholipids, with promising results to improve DHA bioavailability although more studies are needed. In this review, the evidence on DHA bioavailability and accumulation in both maternal and fetal tissues after the administration of DHA supplementation during pregnancy in the form of PL or TG in different models is summarized.

Two-level factorial analysis of the effect of fructose on DHA biosynthetic capacity of Aurantiochytrium sp. SW1

Thraustochytrids are getting increasingly popular due to their high potential role as alternative producers of the high-valued ω-3 polyunsaturated fatty acids (PUFA), docosahexaenoic acid (DHA). While most thraustochytrids prefer glucose as the major carbon source, few strains have been reported to prefer fructose. One such strain is Aurantiochytrium sp. SW1. In this study, the effect of fructose on DHA accumulation by SW1 was investigated using a two-level full factorial design. Besides, biomass, lipid and DHA accumulation profiles of SW1 cultivated in fructose and glucose media were compared. Results revealed that fructose has a very significant positive effect on the volumetric DHA content. Meanwhile, its involvement in affecting DHA biosynthetic capacity, though significant, is not very profound. It was also found that when cultivated in fructose medium, SW1 had a less steep log phase compared to that of glucose medium. However, after 48h of cultivation, biomass and lipid accumulation in fructose medium outweighed the other. Volumetric DHA content in fructose medium at 96h was 11% higher than that of glucose medium. Overall, fructose was found to be a more suitable substrate for biomass, lipid and DHA accumulation in SW1 compared to the conventional source, glucose.

The Association between the Clinical Severity of Heart Failure and Docosahexaenoic Acid Accumulation in Hypertrophic Cardiomyopathy

Background: Hypertrophic cardiomyopathy (HCM) is a common genetic disease with diverse morphology, symptoms, and prognosis. Hypertrophied myocardium metabolism has not been explored in detail. We assessed the association between myocardium lipid metabolism and clinical severity of heart failure (HF) in HCM using imaging mass spectrometry (IMS). Methods and Results: We studied 16 endomyocardial biopsy (EMB) specimens from patients with HCM. Analysis was conducted using desorption electrospray ionization IMS. The Samples were assigned into two cohorts according to the period of heart biopsy (cohort 1, n=9 and cohort 2, n=7). In each cohort, samples were divided into two groups according to the clinical severity of HF in HCM: clinically severe and clinically mild groups. Signals showing a significant difference between the two groups were analyzed by volcano plot. In cohort 1, the volcano plot identified four signals; the intensity in the clinically severe group was more than twice that of the mild group. Out of the four signals, docosahexaenoic acid (DHA) showed significant differences in intensity between the two groups in cohort 2 (10575.8±2750.3 vs. 19839.3±4803.2, p=0.025). Conclusion: The intensity of DHA was significantly higher in EMB samples from the clinically severe HCM group than in those from the mild group. Funding Information: This study received funding from the Ministry of Education, Science, Sports and Culture of Japan, Grants-in-Aid for Scientific Research (Grant Number 18K08065 [YM]). Declaration of Interests: The authors have no conflicts of interest to disclose. Ethics Approval Statement: All the patients provided informed consent. The institutional review board of Hamamatsu University School of Medicine approved all aspects of this study (application number 17-261). The study was conducted in accordance with the regulations of Declaration of Helsinki.

Low-temperature effects on docosahexaenoic acid biosynthesis in Schizochytrium sp. TIO01 and its proposed underlying mechanism

BackgroundSchizochytrium species are known for their abundant production of docosahexaenoic acid (DHA). Low temperatures can promote the biosynthesis of polyunsaturated fatty acids (PUFAs) in many species. This study investigates low-temperature effects on DHA biosynthesis in Schizochytrium sp. TIO01 and its underlying mechanism.ResultsThe Schizochytrium fatty acid biosynthesis pathway was evaluated based on de novo genome assembly (contig N50 = 2.86 Mb) and iTRAQ-based protein identification. Our findings revealed that desaturases, involved in DHA synthesis via the fatty acid synthase (FAS) pathway, were completely absent. The polyketide synthase (PKS) pathway and the FAS pathway are, respectively, responsible for DHA and saturated fatty acid synthesis in Schizochytrium. Analysis of fatty acid composition profiles indicates that low temperature has a significant impact on the production of DHA in Schizochytrium, increasing the DHA content from 43 to 65% of total fatty acids. However, the expression levels of PKS pathway genes were not significantly regulated as the DHA content increased. Further, gene expression analysis showed that pathways related to the production of substrates (acetyl-CoA and NADPH) for fatty acid synthesis (the branched-chain amino acid degradation pathway and the pentose phosphate pathway) and genes related to saturated fatty acid biosynthesis (the FAS pathway genes and malic enzyme) were, respectively, upregulated and downregulated. These results indicate that low temperatures increase the DHA content by likely promoting the entry of relatively large amounts of substrates into the PKS pathway.ConclusionsIn this study, we provide genomic, proteomic, and transcriptomic evidence for the fatty acid synthesis pathway in Schizochytrium and propose a mechanism by which low temperatures promote the accumulation of DHA in Schizochytrium. The high-quality and nearly complete genome sequence of Schizochytrium provides a valuable reference for investigating the regulation of polyunsaturated fatty acid biosynthesis and the evolutionary characteristics of Thraustochytriidae species.