Triacylglyceride Accumulation Research Articles

LIFR-α-dependent adipocyte signaling in obesity limits adipose expansion contributing to fatty liver disease

SummaryThe role of chronic adipose inflammation in diet-induced obesity (DIO) and its sequelae including fatty liver disease remains unclear. Leukemia inhibitory factor (LIF) induces JAK-dependent adipocyte lipolysis and altered adipo/cytokine expression, suppressing in vivo adipose expansion in normal and obese mouse models. To characterize LIF receptor (LIFR-α)-dependent cytokine signaling in DIO, we created an adipocyte-specific LIFR knockout mouse model (Adipoq-Cre;LIFRfl/fl). Differentiated adipocytes derived from this model blocked LIF-induced triacylglycerol lipolysis. Adipoq-Cre;LIFRfl/fl mice on a high-fat diet (HFD) displayed reduced adipose STAT3 activation, 50% expansion in adipose, 20% body weight increase, and a 75% reduction in total hepatic triacylglycerides compared with controls. To demonstrate that LIFR-α signals adipocytes through STAT3, we also created an Adipoq-Cre;STAT3fl/fl model that showed similar findings when fed a HFD as Adipoq-Cre;LIFRfl/fl mice. These findings establish the importance of obesity-associated LIFR-α/JAK/STAT3 inflammatory signaling in adipocytes, blocking further adipose expansion in DIO contributing to ectopic liver triacylglyceride accumulation.

ЭНДОКРИННАЯ ФУНКЦИЯ ЖИРОВОЙ ТКАНИ (ОБЗОР ЛИТЕРАТУРЫ)

Adipose tissue serves not only as a place for the accumulation and storage of triacylglycerides as energy substrates, but also produces many hormone-like substances, mediators, cytokines, chemokines that act at the local and systemic level and aff ect metabolism, regulate tissue sensitivity to insulin, reproductive and the immune system. The review presents the current results of scientifi c research on the problem of the physiological role of biologically active substances produced by adipose tissue and their participation in the development of metabolic disorders.

1H-NMR based lipid profiling of Gossypium hirsutum seed oil at different developmental stages

1H-NMR spectroscopy is emerging as powerful robust analytical technique in analyzing bio constituents present in plants. It offers non-invasive method for identification and quantification of several Phyto-constitutes in a single step without much pre-processing of the samples. In the present study, 1H-NMR spectroscopy-based lipid profiling was performed on Gossypium hirsutum seeds, which were collected at different developmental stages. 1H-NMR study reveals the presence of sterols, butyl ester of fatty acids, free fatty acids (FFA), triacyl glycerides (TAG), monounsaturated fatty acids (MUFA), polyunsaturated fatty acids (PUFA) and saturated fatty acids (SFA) at different percentage at different developmental stages. The 1H-NMR result demonstrates the presence of high concentration of FFA in early developmental stage and increased concentration of TAG at maturation stage. The 1H-NMR results suggest that stage III of seed development can be identified as turning point, as at this stage sterols and butyl ester of fatty acids becomes insignificant followed by high accumulation of TAG.

PLHC-1 topminnow liver cells: An alternative model to investigate the toxicity of plastic additives in the aquatic environment

Plasticizers are widespread environmental contaminants that have been described as obesogens in terrestrial vertebrates. However, its effects on fish lipids homeostasis are almost unknown. This work explores the use of PLHC-1 cells as an alternative model to assess the disruption of hepatic lipids by plastic additives and to gather information on the mode of action of these chemicals in fish. PLHC-1 lipid extracts were analyzed by flow injection coupled to high resolution mass spectrometry (FIA-ESI(+/-)-Orbitrap-Exactive) after 24 h exposure of the cells to the selected plasticizers: dibutyl phthalate (DBP), di-(2-ethylhexyl) phthalate (DEHP), bisphenol A (BPA), bisphenol F (BPF), and chlorinated bisphenol A diglycidyl ether (BADGE·2HCl). The analysis of the culture medium and the intracellular concentration of the chemicals revealed the highest bioconcentration of BADGE·2HCl, DBP and DEHP, which was in agreement with the strongest alteration of the cells lipidome. BADGE·2HCl induced a significant depletion of triacylglycerides (TGs), while DEHP and DBP stimulated the accumulation of TGs. Exposure to BPF induced the generation of reactive oxygen species in PLHC-1 cells and a significant depletion of phosphatidylcholine (PC)- and phosphatidylethanolamine (PE)-plasmalogens, and TGs (cell depots of polyunsaturated fatty acids). Overall, this study evidences different modes of action of plastic additives in topminnow liver cells, describes differential lipidomic signatures, and highlights the higher lipotoxicity of BADGE·2HCl and BPF compared to BPA.

Kinetic modelling of the biochemical 9-octadecenedioic acid production and lipid accumulation using differently functionalised C18:1 substrates

The potential use of different hydrophobic substrates (oleic acid, oleyl alcohol, methyl oleate and rapeseed oil) for the production of high concentrations of 9-octadecenedioic acid by the genetically engineered C. tropicalis ATCC-20962 yeast was researched. For comparison purposes, data for growth, intracellular triacylglyceride (TAG) accumulation and dicarboxylic acid (DCA) production were obtained for mathematical description using both known and newly adapted models. Fermentations were carried out in an aerated, stirred 2L-fermenter in which temperature (30°C) and pH (8.2) were controlled automatically. Using oleic acid resulted in the highest DCA production, i.e. 31.3 g.L−1 in 144 h, as well as the highest intracellular TAG concentration (60.5 m%). Mass balance calculations proved that 35 % of the consumed oleic acid was used for TAG accumulation. The oleyl alcohol and rapeseed oil substrates showed high initial DCA production rates as well, however, after 80 h, DCA production stopped to reach a maximum. In contrast, with methyl oleate substrate, a very slow production was observed without approaching a maximum within the experimental time.

A novel strategy for triacylglycerides and polyhydroxyalkanoates production using waste lipids

Lipids are one of the main components of the organic matter present in the effluents of the food-processing industry. These waste streams can be biotransformed into valuable triacylglycerides (TAGs) and polyhydroxyalkanoates (PHAs), precursors of biofuels and biomaterials alternative to petroleum-based products. These compounds are yielded by mixed microbial cultures, and considering that both TAG and PHA accumulators may coexist within the community, it seems crucial to define those operational strategies that might control the selection of the dominant metabolic pathways (TAG or PHA accumulation). In this work, residual fish-canning oil was used as a carbon source in a two-stage process (culture selection and intracellular compounds accumulation) in which the substrate was simultaneously hydrolyzed in these two stages without the need for a previous fermentation unit. It was pretended to maximize preferential TAG or PHA storage in the accumulation reactor by the imposition of certain selective pressures in the enrichment one. Uncoupling C and N feedings and limiting nitrogen availability in the medium, allowed to maximize PHA production (82.3 wt% of PHAs, 0.80 CmmolPHA/CmmolS). Besides, when low pH in the famine phase was considered as additional selective pressure, it was possible to shift the ratio TAG:PHA from 4:96 obtaining 43.0 wt% of TAGs (0.67 CmmolTAG/CmmolS). Therefore, this novel and simplified process demonstrated versatility and efficiency in the storage of TAGs and PHAs from a unique residual feedstock and using an open culture proving that product selection can be harnessed if choosing the right operational conditions in the enrichment stage.

Targeting n-3 Polyunsaturated Fatty Acids in Non-Alcoholic Fatty Liver Disease.

Non-Alcoholic Fatty Liver Disease (NAFLD) is characterized by abnormal hepatic accumulation of triacylglycerides in the absence of alcohol consumption, in association with Oxidative Stress (OS), a pro-inflammatory state and Insulin Resistance (IR), which are attenuated by n-3 long-chain polyunsaturated Fatty Acids (FAs) C20-C22 (LCPUFAs) supplementation. Main causes of NAFLD comprise high caloric intake and a sedentary lifestyle, with high intakes of saturated FAs. The review includes several searches considering the effects of n-3 LCPUFAs in NAFLD in vivo and in vitro models, using the PubMed database from the National Library of Medicine- National Institutes of Health. The LCPUFAs eicosapentaenoic acid (C20:5 n-3, EPA) and docosahexaenoic acid (C22:6 n- 3, DHA) have a positive effect in diminishing liver steatosis, OS, and the levels of aspartate aminotransferase, alanine aminotransferase and pro-inflammatory cytokines, with improvement of insulin sensitivity and adiponectin levels. The molecular pathways described for n-3 LCPUFAs in cellular and animal models and humans include peroxisome proliferator-activated receptor-α activation favouring FA oxidation, diminution of lipogenesis due to sterol responsive element binding protein-1c downregulation and inflammation resolution. Besides, nuclear factor erythroid-2-related factor-2 activation is elicited by n-3 LCPUFA-derived oxidation products producing direct and indirect antioxidant responses, with concomitant anti-fibrogenic action. The discussed effects of n-3 LCPUFA supplementation support its use in NAFLD, although having a limited value in NASH, a contention that may involve n-3 LCPUFA oxygenated derivatives. Clinical trials establishing optimal dosages, intervention times, type of patients and possible synergies with other natural products are needed in future studies.

Integration of proteome and transcriptome refines key molecular processes underlying oil production in Nannochloropsis oceanica

BackgroundUnder nitrogen deficiency situation, Nannochloropsis spp. accumulate large amounts of lipids in the form of triacylglycerides (TAG). Mechanisms of this process from the perspective of transcriptome and metabolome have been obtained previously, yet proteome analysis is still sparse which hinders the analysis of dynamic adaption to nitrogen deficiency. Here, proteomes for 3 h, 6 h, 12 h, 24 h, 48 h and 10th day of nitrogen deplete (N−) and replete (N+) conditions were obtained and integrated with previous transcriptome data for N. oceanica.ResultsPhysiological adaptations to N− not apparent from transcriptome data were unveiled: (a) abundance of proteins related to photosynthesis only slightly decreased in the first 48 h, indicating that photosynthesis is still working efficiently, and protein amounts adjust gradually with reduction in chloroplast size. (b) Most proteins related to the TCA cycle were strongly upregulated after 48 h under N−, suggesting that respiration is enhanced after 48 h and that TCA cycle efflux supports the carbon required for lipid synthesis. (c) Proteins related to lipid accumulation via the Kennedy pathway increased their abundance at 48 h, synchronous with the previously reported diversification of fatty acids after 48 h.ConclusionsThis study adds a proteome perspective on the major pathways for TAG accumulation in Nannochloropsis spp. Temporal changes of proteome exhibited distinct adaptation phases that are usually delayed relative to transcriptomic responses. Notably, proteome data revealed that photosynthesis and carbon fixation are still ongoing even after 48 h of N−. Moreover, sometimes completely opposite trends in proteome and transcriptome demonstrate the relevance of underexplored post-transcriptional regulation for N− adaptation.

Lemon balm and dandelion leaf extract synergistically alleviate ethanol-induced hepatotoxicity by enhancing antioxidant and anti-inflammatory activity.

We investigated the effect of a 2:1 (w/w) mixture of lemon balm and dandelion extracts (LD) on ethanol (EtOH)-mediated liver injury and explored the underlying mechanisms. Administration of LD synergistically reduced relative liver weight and decreased the levels of serum biomarkers of hepatic injury. Histopathological and biochemical analyses indicated that LD synergistically attenuated hepatic accumulation of triacylglycerides (TGs) and restored the levels of mRNAs related to fatty acid metabolism. In addition, LD significantly reduced EtOH-induced hepatic oxidative stress by attenuating the reduction in levels of nuclear factor E2-related factor 2 (Nrf2) mRNA and enhancing antioxidant activity. Moreover, LD decreased the EtOH-mediated increase in levels of hepatic tumor necrosis factor-α (TNF-α) mRNA. In vitro, LD significantly scavenged free radicals, increased cell viability against tert-butylhydroperoxide (tBHP), and transactivated Nrf2 target genes in HepG2 cells. Furthermore, LD decreased levels of pro-inflammatory mediators in lipopolysaccharide-stimulated Raw264.7 cells. Therefore, LD shows promise for preventing EtOH-mediated liver injury. PRACTICAL APPLICATIONS: There were no approved therapeutic agents for preventing and/or treating alcoholic liver diseases. In this study, a 2:1 (w/w) mixture of lemon balm and dandelion leaf extract (DL) synergistically ameliorated EtOH-induced hepatic injury by inhibiting lipid accumulation, oxidative stress, and inflammation. Our findings will enable the development of a novel food supplement for the prevention or treatment of alcohol-mediated liver injury.

Oleaginous yeasts isolated from traditional fermented foods and beverages of Manipur and Mizoram, India, as a potent source of microbial lipids for biodiesel production

PurposeOleaginous yeasts can accumulate intracellular lipid bodies or triacylglycerides (TAGs) under nutrient limiting conditions. TAGs derived from those yeast strains are considered as an alternative to conventional plant-based oils for biodiesel production. In this study, we attempt to isolate and characterize yeast strains from selected traditional fermented foods of Manipur and Mizoram, India, and study their oleaginous attributes for biodiesel production.MethodFourteen potential oleaginous yeasts were isolated from fermented food samples of Manipur and Mizoram, India. The isolates were identified by 5.8S internal transcribed spacer (ITS) rRNA gene sequencing. Intracellular TAG accumulation by yeast cells were confirmed by Nile red fluorescence microscopy and spectrometry technique. The most promising isolates were evaluated for lipid accumulation having different initial carbon to nitrogen (C/N) ratios and also the full kinetic studies (depicting the glucose consumption, biomass, and lipid production) using optimum C/N ratio were estimated. Fatty acid methyl esters (FAME) profile of the transesterified lipids were analyzed by GC-MS.ResultsThe identified yeast isolates belonged to seven different genera viz. Rhodotorula, Pichia, Candida, Saturnispora, Wickerhamomyces, Zygoascus, and Saccharomyces. Under nitrogen-limiting conditions, maximum biomass concentration of 5.66 ± 0.03 g/L and 4.713 ± 0.03 g/L was produced by Wickerhamomyces anomalus FK09 and Pichia kudriavzevii FK02, respectively. The highest lipid concentration (g lipid/L fermentation broth) of 0.58 g/L was attained by Rhodotorula mucilaginosa R2, followed by Wickerhamomyces anomalus FK09 (0.51 g/L), and Zygoascus hellenicus FC10 (0.41 g/L). Rhodotorula mucilaginosa R2 exhibited the maximum lipid content (% lipid/g dry cell weight) of (21.63 ± 0.1%) after 96 h of growth. The C/N ratio of 40 and 20 was found to be optimum for R. mucilaginosa R2 and W. anomalus FK09 with a lipid content of 22.21 ± 0.4% and 12.83 ± 0.08% respectively.ConclusionNewly isolated yeast strains were obtained from traditional fermented food samples of Manipur and Mizoram, India. FAME analysis of the transesterified lipid extracts suggested the potential use of yeast-derived oil as an alternative to vegetable oil for biodiesel production.

Enrichment of f/2 medium hyperaccumulates biomass and bioactive compounds in the diatom Phaeodactylum tricornutum

Photosynthetic microalgae have garnered research attention due to their inherent advantageous characteristics, however, commercial exploitation of algae is still hampered by their relatively compromised overall productivity. Thus, it is of paramount significance to design a strategy to concurrently enhance the growth rate and compounds production. In this study, we interrogated the impact of fortifying bacterial growth medium components in the conventional f/2-medium for microalgae. Our results showed that supplement of yeast extract and tryptone significantly increased cell growth of the model diatom Phaeodactylum tricornutum. Particularly, cell biomass and triacylglyceride content increased up to 3.48-fold and 2.13-fold in the stationary phase, respectively. Besides, fucoxanthin content increased by 1.7-fold. Transcript abundance of key genes involved in carotenoid synthesis also increased significantly. Overall, the results exemplified a feasible and promising strategy to simultaneously enhance the biomass and accumulation of triacylglyceride and fucoxanthin for industrial applications of microalgae.

Responses of a Newly Evolved Auxotroph of Chlamydomonas to B12 Deprivation.

The corrinoid B12 is synthesized only by prokaryotes yet is widely required by eukaryotes as an enzyme cofactor. Microalgae have evolved B12 dependence on multiple occasions, and we previously demonstrated that experimental evolution of the non-B12-requiring alga Chlamydomonas reinhardtii in media supplemented with B12 generated a B12-dependent mutant (hereafter metE7). This clone provides a unique opportunity to study the physiology of a nascent B12 auxotroph. Our analyses demonstrate that B12 deprivation of metE7 disrupts C1 metabolism, causes an accumulation of starch and triacylglycerides, and leads to a decrease in photosynthetic pigments, proteins, and free amino acids. B12 deprivation also caused a substantial increase in reactive oxygen species, which preceded rapid cell death. Survival could be improved without compromising growth by simultaneously depriving the cells of nitrogen, suggesting a type of cross protection. Significantly, we found further improvements in survival under B12 limitation and an increase in B12 use efficiency after metE7 underwent a further period of experimental evolution, this time in coculture with a B12-producing bacterium. Therefore, although an early B12-dependent alga would likely be poorly adapted to coping with B12 deprivation, association with B12-producers can ensure long-term survival whilst also providing a suitable environment for evolving mechanisms to tolerate B12 limitation better.

Targeting PLIN2/PLIN5-PPARγ: Sulforaphane Disturbs the Maturation of Lipid Droplets.

The effects of sulforaphane (SFN) on the maturation of lipid droplets (LDs)-the storage units for free fatty acids and sterols as triacylglycerides (TAG) and cholesterol esters (CE)-are far from being understood, despite the fact that SFN is known to be beneficial for ameliorating lipid metabolism disorders. High-fat-intake models are established in both HHL-5 hepatocytes and rodents. The numbers and sizes of LDs are decreased by SFN. The accumulation of lipid core components (TAG & CE) is reduced and the expression of their key synthetases, acyl-coenzyme A: diacylglycerol acyltransferases 2 (DGAT2) and acyl-coenzyme A: cholesterol acyltransferases 1 (ACAT1), is also inhibited. Moreover, SFN decreases LD-associated protein PLIN2 and PLIN5 expression, but not that of PLIN1 and PLIN3, both in vivo and in vitro. Furthermore, over-expression of peroxisome proliferator-activated receptor gamma (PPARγ) induces the accumulation of TAG and the up-regulation of PLIN2 and PLIN5, which are not reversed by SFN. These results suggest that PPARγ may be a target of SFN in lipid metabolism. SFN disturbs LD maturation by inhibiting the formation of the neutral lipid core and decreases PLIN2 and PLIN5 via down-regulation of PPARγ.

Palmitic Acid-Enriched Diet Induces Hepatic Steatosis and Injury in Adult Zebrafish.

Palmitic acid (PA) is the most abundant saturated fatty acid in fast foods and is known to induce inflammation and cellular injury in various tissues. In this study, we investigated whether a PA-enriched diet can induce hepatic steatosis and injury in adult zebrafish. The adult zebrafish exhibited increased body weight, hyperlipidemia, hyperglycemia, and steatosis and a hepatic injury phenotype after being fed with a PA-enriched diet for 6 weeks. The quantitative polymerase chain reaction analysis demonstrated that genes associated with hepatic injury were all significantly increased in the liver. Furthermore, livers from the PA-fed group showed an increased messenger RNA (mRNA) expression associated with oxidative stress and endoplasmic reticulum (ER) stress responses. We also found significant upregulation of genes involved in lipid metabolism and triacylglyceride accumulation. Ultrastructural analysis revealed mitochondrial cristae injury and a dilated ER phenotype in the PA-fed hepatocytes, which can be causes of hepatic injury. PA-enriched diet induced hepatic steatosis and injury in adult zebrafish that recapitulated typical metabolic changes and pathophysiological changes as well as increased oxidative stress and ER stress observed in patients with nonalcoholic fatty liver disease.

Drospirenone induces the accumulation of triacylglycerides in the fish hepatoma cell line, PLHC-1.

Drospirenone (DRO) is one of the most commonly used progestins reaching the aquatic environment through wastewater treatment plant effluents. It is a progesterone receptor agonist, and as such, can act primarily in the brain and reproductive organs of fish. In order to better understand and predict its effects, this work evaluates the lipidomic changes induced in PLHC-1 cells after exposure to drospirenone at concentrations below the EC10 (1 and 10 μM) by direct injection of the lipid extracts into a ESI(+/-) Orbitrap mass spectrometer. A significant accumulation of triacylglycerides, particularly long chain ones with unsaturated fatty acid moieties (TGs 46:2, 56:4-7; 58:5-8) and a concomitant decrease of diacylglycerides (DGs 32:1, 34:1-2, 36:1-2, 38:2-4) was observed after 48 h exposure to 10 μM DRO, which corresponded to an intracellular concentration of 8.3 ng·mg-1 protein. No significant alteration of PLHC-1 cell lipids was observed following exposure to 1 μM DRO. EC50 for the cytotoxicity of DRO ranged from 105 to 119 μM (24 h exposure) to 51-58 μM (48 h exposure). The study evidences a dysregulation of neutral lipid metabolism and increased TG/DG ratio in fish hepatic cells exposed to DRO.

Effect of interferon gamma on neutral lipid levels, lipid droplet formation, and antiviral responses in pancreatic islets and INS‐1 β cells

Interferon (IFN)‐regulated pathways play a critical role in the development of type 1 diabetes (T1D) following exposure to environmental insults including viral infection. IFNs have been shown to modulate lipid metabolism in both immune and non‐hematopoietic cells in response to pathogens. Little is known, however, about the impact of IFN signaling on lipid metabolism in pancreatic β cells and whether IFN‐induced changes in β cell lipid metabolism are associated with antiviral responses and β cell dysfunction. To characterize islet lipid changes potentially mediated by IFNs during initiation of β cell autoimmunity in vivo, juvenile LEW.1WR1 rats (an IFN‐dependent, inducible‐T1D model) were treated with the viral analog polyinosinic:polycytidylic (PIC) every other day for 6 days then islets were isolated 16h and 48h post last treatment for gene expression and non‐targeted lipidomics, respectively. Islets from PIC‐treated animals displayed a robust induction of IFN gamma (IFNγ) target genes, e.g. IFNγ‐inducible protein and ubiquitin D, and significantly increased islet triacylglyceride (TAG) and non‐esterified fatty acids (NEFA). Similar increases in TAG and NEFA were observed when isolated human islets were treated ex vivo for 24h with mixtures of IFNγ and IL1β, or IFNγ and TNFα, suggesting that IFNγ drives TAG synthesis or accumulation. To investigate the direct effect of IFNγ, INS‐1 β cells were treated with IFNγ for 24h. Consistent with the in vivo and ex vivo islet results, IFNγ‐treated cells had significantly higher TAG levels. IFNγ also increased mRNA levels of genes involved in lipid droplet (LD) formation and clustering (e.g. perilipins and fat‐induced storage membrane protein 1) and led to the appearance of clusters of large cytoplasmic LDs. Treatment of INS‐1 cells with IFNγ also increased NEFA levels, most notably arachidonic acid. Although the source for increased TAG and NEFA is uncertain, IFNγ induced fatty acid synthase (FASN) gene expression suggesting increased de novo FA synthesis. IFNγ‐mediated changes in lipid metabolism were associated with impaired insulin secretion. IFNγ, however, did not increase the expression of endoplasmic reticulum (ER) stress markers, but sensitized INS‐1 cells to TNFα‐ or IL1β‐induced ER stress and apoptosis. Importantly, pretreatment of INS‐1 β cells with IFNγ for 12 h markedly increased PIC‐induced expression of antiviral genes (e.g. Ifnb, Mx1) and this synergistic response was abrogated by the FASN inhibitor C75. Inhibition of FASN with C75 also led to a redistribution of IFNγ‐mediated LD clusters into more discrete LDs. Overall, these data suggest that IFNγ promotes LD formation and clustering in β cells, and these LD clusters serve as scaffolds to enhance expression of antiviral response genes. Unfortunately, these lipid metabolism changes are also associated with impaired secretory function and increased susceptibility to cytokine‐induced ER stress and β cell death.Support or Funding InformationJDRF grant 3‐SRE‐2014‐43‐Q‐R to LKO.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Unconventional alternative biofuels: Quality assessment of biodiesel and its blends from marine diatom Navicula cincta

The use of microalgae as triacylglycerides (TAG) feedstock for biodiesel is a widely researched field in the development of new technologies. In order to underpin the potential of microalgal biodiesel as a possible substitute in the biofuel industry, more detailed data is therefore required on these aspects. This study assesses the growth, TAG accumulation, kinetics of dissolved nutrients and the quality of biodiesel and its blends from the marine diatom Navicula cincta grown in a photobioreactor, using seawater enriched with nutrients and soil extract. Acid-catalyst transesterification was tested for biodiesel production. This species presented during nutrient depletion: 1) a total lipid content of 49.7% of ash-free dry weight (AFDW) with neutral lipid (mainly TAG) as the dominant fraction (ca. 81.5% of total lipids), and 2) favourable fatty acids for biodiesel, such as palmitic acid (28%) and palmitoleic acid (46%). The yield of biodiesel (B100) derived from diatom N. cincta (BNC) and analysed by Gas Chromatography was 97.6% of FAME, being the FAME composition similar to the fatty acid profile of N. cincta neutral lipids. The fuel properties (including FAME percentage, cetane number, heat of combustion, color and sulfur content) of diatom blends prepared with petrodiesel in a 7% (B7NC) and 10% (B10NC) vol ratio were evaluated using a PetroSpec (PAC) and compared with commercial soybean blends (B7S and B10S). B7 and B10 derived from N. cincta met the requirements of ASTM D6751 and presented improved cetane numbers (50.2 and 51.6, respectively) with respect to soybean blends. Furthermore, biodiesel derived from the studied species showed the lowest sulfur content (0.0056% w/w). The rapid accumulation of neutral lipids in response to natural nutrient deficiency in N. cincta and its capacity of autoflocculate are good selling points for the use of this microalga in the biofuel industry.

Delineating the molecular responses of a halotolerant microalga using integrated omics approach to identify genetic engineering targets for enhanced TAG production

BackgroundHarnessing the halotolerant characteristics of microalgae provides a viable alternative for sustainable biomass and triacylglyceride (TAG) production. Scenedesmus sp. IITRIND2 is a fast growing fresh water microalga that has the capability to thrive in high saline environments. To understand the microalga’s adaptability, we studied its physiological and metabolic flexibility by studying differential protein, metabolite and lipid expression profiles using metabolomics, proteomics, real-time polymerase chain reaction, and lipidomics under high salinity conditions.ResultsOn exposure to salinity, the microalga rewired its cellular reserves and ultrastructure, restricted the ions channels, and modulated its surface potential along with secretion of extrapolysaccharide to maintain homeostasis and resolve the cellular damage. The algal-omics studies suggested a well-organized salinity-driven metabolic adjustment by the microalga starting from increasing the negatively charged lipids, up regulation of proline and sugars accumulation, followed by direction of carbon and energy flux towards TAG synthesis. Furthermore, the omics studies indicated both de-novo and lipid cycling pathways at work for increasing the overall TAG accumulation inside the microalgal cells.ConclusionThe salt response observed here is unique and is different from the well-known halotolerant microalga; Dunaliella salina, implying diversity in algal response with species. Based on the integrated algal-omics studies, four potential genetic targets belonging to two different metabolic pathways (salt tolerance and lipid production) were identified, which can be further tested in non-halotolerant algal strains.

Innovative screening approach for the identification of triacylglycerol accumulating oleaginous strains

Currently, triacylglycerides (TAG) accumulation in the form of lipid droplets (LDs) in oleaginous microorganisms is of immense importance due to their ability to get transesterified into value-added products in the form of biodiesel. Hence, in order to search for oleaginous microorganisms having high lipid content among a wide range of samples from different niches, there is a compulsive need to develop simple, reliable and rapid methods for screening of TAG accumulating strains. Conventional methods require multistep processes for the isolation, cultivation, extraction and estimation of lipids to identify oleagenic strains. To overcome these challenges, we are proposing an easy, live cell imaging technique for the estimation of lipids via visualization of TAG accumulation in probable strains at the single cell level that gives real-time monitoring of intracellular lipid accumulation in yeasts. In this screening technique, only 100 μl of specific neutral lipid accumulating medium was used to grow the isolated culture in the microtiter plate. The harvested cells were stained with LipidTOX™ Green and visualized by a LED based digital inverted fluorescence microscope. Among 446 yeast colonies screened, maximum lipid producing yeast strains Rhodosporidium kratochvilovae HIMPA1 and Rhodotorula minuta, having supersized lipid body of 5.05 ± 0.87 μm and 4.46 ± 0.61 μm, respectively, were identified as potential candidates for biodiesel production. To the best of our knowledge, this is the first report of using LipidTOX™ Green for the staining of lipid droplets present in yeast cells as per the literature.

Chlorogenic Acid Functions as a Novel Agonist of PPARγ2 during the Differentiation of Mouse 3T3-L1 Preadipocytes.

Rosiglitazone (RG) is a well-known activator of peroxisome proliferator-activated receptor-gamma (PPARγ) and used to treat hyperglycemia and type 2 diabetes; however, its clinical application has been confounded by adverse side effects. Here, we assessed the roles of chlorogenic acid (CGA), a phenolic secondary metabolite found in many fruits and vegetables, on the differentiation and lipolysis of mouse 3T3-L1 preadipocytes. The results showed that CGA promoted differentiation in vitro according to oil red O staining and quantitative polymerase chain reaction assays. As a potential molecular mechanism, CGA downregulated mRNA levels of the adipocyte differentiation-inhibitor gene Pref1 and upregulated those of major adipogenic transcriptional factors (Cebpb and Srebp1). Additionally, CGA upregulated the expression of the differentiation-related transcriptional factor PPARγ2 at both the mRNA and protein levels. However, following CGA intervention, the accumulation of intracellular triacylglycerides following preadipocyte differentiation was significantly lower than that in the RG group. Consistent with this, our data indicated that CGA treatment significantly upregulated the expression of lipogenic pathway-related genes Plin and Srebp1 during the differentiation stage, although the influence of CGA was weaker than that of RG. Notably, CGA upregulated the expression of the lipolysis-related gene Hsl, whereas it did not increase the expression of the lipid synthesis-related gene Dgat1. These results demonstrated that CGA might function as a potential PPARγ agonist similar to RG; however, the impact of CGA on lipolysis in 3T3-L1 preadipocytes differed from that of RG.