The role of acyl-CoA thioesterase ACOT8I in mediating intracellular lipid metabolism in oleaginous fungus <Emphasis Type="Italic">Mortierella alpina</Emphasis>

Thioesterases (TEs) play an essential role in the metabolism of fatty acids (FAs). To explore the role of TEs in mediating intracellular lipid metabolism in the oleaginous fungus Mortierella alpina, the acyl-CoA thioesterase ACOT8I was overexpressed. The contents of total fatty acids (TFAs) were the same in the recombinant strains as in the wild-type M. alpina, whilst the production of free fatty acids (FFAs) was enhanced from about 0.9% (wild-type) to 2.8% (recombinant), a roughly threefold increase. Linoleic acid content in FFA form constituted about 9% of the TFAs in the FFA fraction in the recombinant strains but only about 1.3% in the wild-type M. alpina. The gamma-linolenic acid and arachidonic acid contents in FFA form accounted for about 4 and 25%, respectively, of the TFAs in the FFA fraction in the recombinant strains, whilst neither of them in FFA form were detected in the wild-type M. alpina. Overexpression of the TE ACOT8I in the oleaginous fungus M. alpina reinforced the flux from acyl-CoAs to FFAs, improved the production of FFAs and tailored the FA profiles of the lipid species.     

Long-Chain Free Fatty Acid Profiling Analysis by Liquid Chromatography–Mass Spectrometry in Mouse Treated with Peroxisome Proliferator-Activated Receptor α Agonist

A change in the free fatty acid (FFA) profile reflects an alteration in the lipid metabolism of peripheral tissue. A high-throughput quantitative analysis method for individual FFAs therefore needs to be established. We report here an optimized LC-MS assay for a high-throughput and high-sensitivity analysis of the 10 major long-chain FFAs in mouse plasma and liver. This assay enables quantification of individual FFAs by using trace amounts of samples (2 µL of plasma and 10 mg of liver tissue). We apply this method to analyze the FFA profile of plasma and liver samples from an obese mouse model treated with bezafibrate, the peroxisome proliferator-activated receptor α (PPARα) agonist, and show a change in the FFA profile, particularly in the palmitoleic and oleic acid contents. This assay is useful for quantifying individual FFAs and helpful for monitoring the condition of lipid metabolism.     

Deconvolution as a novel approach to analyze moment-to-moment free fatty acid release

Objective: Recent studies have shown that free fatty acid (FFA) release is pulsatile and that this pattern is controlled by the sympathetic nervous system. It is, then, necessary to understand and characterize adipose tissue lipolysis to elucidate its effect on metabolism. In this study, we introduce deconvolution as a method to detect and quantify pulsatile FFA release. Research Methods and Procedures: Octanoate, a medium‐chain fatty acid, was infused in male mongrel dogs (n = 7) to mimic the pulsatile appearance of plasma FFAs. Deconvolution analysis was used to reconstruct the number and timing of infused octanoate pulses from plasma FFA concentrations. Results: Deconvolution analysis was able to reconstruct the exogenously infused pulses of octanoate used to mimic pulsatile appearance of FFAs (pulse frequency, 8 per hour; interpulse interval, 7 minutes). However, determination of pulse mass was less accurate (1.0 ± 0.0 vs. 0.54 ± 0.1 mM). The addition of varying levels of Gaussian noise to non‐oscillatory FFA time series did not lead to detection of extraneous FFA pulses. However, goodness of fit declined with increasing variability. Discussion: These results support the use of deconvolution as an accurate approach to determine the temporal sequence of endogenous FFA release.     

An Oleaginous Bacterium That Intrinsically Accumulates Long-Chain Free Fatty Acids in its Cytoplasm

Medium- and long-chain fatty acids are present in organisms in esterified forms that serve as cell membrane constituents and storage compounds. A large number of organisms are known to accumulate lipophilic materials as a source of energy and carbon. We found a bacterium, designated GK12, that intrinsically accumulates free fatty acids (FFAs) as intracellular droplets without exhibiting cytotoxicity. GK12 is an obligatory anaerobic, mesophilic lactic acid bacterium that was isolated from a methanogenic reactor. Phylogenetic analysis based on 16S rRNA gene sequences showed that GK12 is affiliated with the family Erysipelotrichaceae in the phylum Firmicutes but is distantly related to type species in this family (less than 92% similarity in 16S rRNA gene sequence). Saturated fatty acids with carbon chain lengths of 14, 16, 18, and 20 were produced from glucose under stress conditions, including higher-than-optimum temperatures and the presence of organic solvents that affect cell membrane integrity. FFAs were produced at levels corresponding to up to 25% (wt/wt) of the dry cell mass. Our data suggest that FFA accumulation is a result of an imbalance between excess membrane fatty acid biosynthesis due to homeoviscous adaptation and limited β-oxidation activity due to anaerobic growth involving lactic acid fermentation. FFA droplets were not further utilized as an energy and carbon source, even under conditions of starvation. A naturally occurring bacterium that accumulates significant amounts of long-chain FFAs with noncytotoxicity would provide useful strategies for microbial biodiesel production.     

Modulating Membrane Composition Alters Free Fatty Acid Tolerance in Escherichia coli

Microbial synthesis of free fatty acids (FFA) is a promising strategy for converting renewable sugars to advanced biofuels and oleochemicals. Unfortunately, FFA production negatively impacts membrane integrity and cell viability in Escherichia coli, the dominant host in which FFA production has been studied. These negative effects provide a selective pressure against FFA production that could lead to genetic instability at industrial scale. In prior work, an engineered E. coli strain harboring an expression plasmid for the Umbellularia californica acyl-acyl carrier protein (ACP) thioesterase was shown to have highly elevated levels of unsaturated fatty acids in the cell membrane. The change in membrane content was hypothesized to be one underlying cause of the negative physiological effects associated with FFA production. In this work, a connection between the regulator of unsaturated fatty acid biosynthesis in E. coli, FabR, thioesterase expression, and unsaturated membrane content was established. A strategy for restoring normal membrane saturation levels and increasing tolerance towards endogenous production of FFAs was implemented by modulating acyl-ACP pools with a second thioesterase (from Geobacillus sp. Y412MC10) that primarily targets medium chain length, unsaturated acyl-ACPs. The strategy succeeded in restoring membrane content and improving viability in FFA producing E. coli while maintaining FFA titers. However, the restored fitness did not increase FFA productivity, indicating the existence of additional metabolic or regulatory barriers.     

Effect of curcumin on glucose and lipid metabolism,FFAs and TNF-α in serum of type 2 diabetes mellitus rat models

ObjectiveTo investigate how curcumin affects the glucose and lipid metabolism in type 2 diabetes mellitus (DM) rat models, and to explore its effect on the free fatty acid (FFA) and tumor necrosis factor α (TNF-α) in serum.MethodsSuccessfully established type 2 DM rats were divided into three groups, i.e. the normal control group, model group and curcumin group, and received the medication for consecutive 8 weeks. Thereafter, we detected the level of fasting blood glucose (FBG), and the blood glucose at 30 min, 60 min and 120 min; besides, we also carried out the insulin tolerance tests to measure the levels of fasting serum insulin (FINS) and blood glucose at 40 min and 90 min; additionally, we also detected the levels of TC, TG, HDL-C, LDL-C, FFA and TNF-α in serum. The results were expected to discover the mechanism of curcumin in decreasing the blood glucose level in DM rats.ResultsCompared with the model group, AUCs of FBG, blood glucose at 30 min, 60 min and 120 min, and glucose were decreased in varying degrees in the curcumin group, and the differences had statistical significance (p < .05). After subcutaneous injection of insulin, we found that the blood glucose at 40 min and 90 min in the curcumin group was decreased, while AUC of glucose level was also decreased (p < .05 or .01). Eight weeks after medication, compared with the rats in the normal group, the levels of HDL-C, LDL-C, TC and TG in rats of the model group and the curcumin group were obviously increased (p < .05). In comparison with the model group, the level of LDL-C in rats of the curcumin group was also decreased significantly (p < .05). In comparison with the normal control group in the same period, we found that the content of FFAs and TNF-α in serum of rats of the model group were elevated significantly, and the differences had statistical significance (p < .05 or .01); the levels in the curcumin group were significantly decreased in comparison with the model group in the same period, and the difference had statistical significance (p < .05 or .01).ConclusionTreatment with curcumin can significantly improve the metabolic disorder of glucose and lipid, enhance the sensitivity to the insulin, and ameliorate the resistance to insulin of the type II DM rats. Meanwhile, this treatment method can also significantly decrease the level of FFA and TNF-α in serum of type II DM rats. Thus, we inferred that the mechanism of curcumin to improve the insulin resistance might be correlated with the decreases of FFA and TNF-α in serum.     

Effects of the Suxiao Jiuxin pill on acute myocardial infarction assessed by comprehensive metabolomics

BackgroundSJP is the commercial Chinese medicine included in the Chinese Pharmacopoeia, with well-established cardiovascular protective effects in the clinic. However, the mechanisms underlying the protective effects of SJP on cardiovascular disease have not yet been clearly elucidated.AimsTo investigate the underlying protective mechanisms of SJP in an acute myocardial infarction (AMI) rat model using comprehensive metabolomics.Materials and methodsThe rat model of AMI was generated by ligating the left anterior descending coronary artery. After 2 weeks treatment with SJP, the entire metabolic changes in the serum, heart, urine and feces of the rat were profiled by HPLC-QTOF-MS/MS.ResultsThe metabolic profiles in different biological samples (heart, serum, urine and feces) were significantly different among groups, in which a total of 112 metabolites were identified. AMI caused comprehensive metabolic changes in amino acid metabolism, galactose metabolism and fatty acid metabolism, while SJP reversed more than half of the differential metabolic changes, mainly affecting amino acid metabolism and fatty acid metabolism. Correlation analysis found that SJP could significantly alter the metabolic activity of 12 key metabolites, regarded as potential biomarkers of SJP treatment. According to the results of network analysis, 6 biomarkers were considered to be hub metabolites, which means that these metabolites may have a major relationship with the SJP therapeutic effects on AMI.ConclusionThe combined comprehensive metabolomics and network analysis, indicated that the protective effect of SJP on cardiovascular disease was associated with systemic metabolic modulation, in particular regulation of amino acid and fatty acid metabolism.     

Engineering Escherichia coli for odd straight medium chain free fatty acid production

Microbial biosynthesis of free fatty acids (FFAs) can be achieved by introducing an acyl–acyl carrier protein thioesterase gene into Escherichia coli. The engineered E. coli usually produced even chain FFAs. In this study, propionyl-CoA synthetase (prpE) from Salmonella enterica was overexpressed in two efficient even chain FFAs producers, ML103 (pXZM12) carrying the acyl-ACP thioesterase gene from Umbellularia californica and ML103 (pXZ18) carrying the acyl-ACP thioesterase gene from Ricinus communis combined with supplement of extracellular propionate. With these metabolically engineered E. coli, the odd straight chain FFAs, undecanoic acid (C11:0), tridecanoic acid (C13:0), and pentadecanoic acid (C15:0) were produced from glucose and propionate. The highest total odd straight chain FFAs produced by ML103 (pXZM12, pBAD-prpE) reached 276 mg/l with a ratio of 23.43 % of the total FFAs. In ML103 (pXZ18, pBAD-prpE), the highest total odd straight chain FFAs accumulated to 297 mg/l, and the ratio reached 17.68 % of the total FFAs. Due to the different substrate specificity of the acyl-ACP thioesterases, the major odd straight chain FFA components of ML103 (pXZM12, pBAD-prpE) were undecanoic acid and tridecanoic acid, while the ML103 (pXZ18, pBAD-prpE) preferred pentadecanoic acid.     

Metabolic engineering of Escherichia coli for efficient free fatty acid production from glycerol

Crude glycerol, generated as waste by-product in biodiesel production process, has been considered as an important carbon source for converting to value-added bioproducts recently. Free fatty acids (FFAs) can be used as precursors for the production of biofuels or biochemicals. Microbial biosynthesis of FFAs can be achieved by introducing an acyl–acyl carrier protein thioesterase into Escherichia coli. In this study, the effect of metabolic manipulation of FFAs synthesis cycle, host genetic background and cofactor engineering on FFAs production using glycerol as feed stocks was investigated. The highest concentration of FFAs produced by the engineered stain reached 4.82 g/L with the yield of 29.55% (g FFAs/g glycerol), about 83% of the maximum theoretical pathway value by the type II fatty acid synthesis pathway. In addition, crude glycerol from biodiesel plant was also used as feedstock in this study. The FFA production was 3.53 g/L with a yield of 24.13%. The yield dropped slightly when crude glycerol was used as a carbon source instead of pure glycerol, while it still can reach about 68% of the maximum theoretical pathway yield.     

Improvement of free fatty acid production in Escherichia coli using codon-optimized Streptococcus pyogenes acyl-ACP thioesterase

Fatty acyl–acyl carrier protein (ACP) thioesterase (acyl-ACP TE) from Streptococcus pyogenes (strain MGAS10270) was codon-optimized and expressed in Escherichia coli K-12 W3110 and Escherichia coli K-12 MG1655. By employing codon-optimized S. pyogenes acyl-ACP TE to improve the total free fatty acids (FFAs) and to tailor the composition of FFAs, high-specificity production of saturated fatty acids (C12, C14) and unsaturated fatty acids (C18:1 C18:2) was achieved in recombinants. E. coli SGJS41 and SGJS46 (codon-optimized acyl-ACP TE of S. pyogenes) demonstrated the highest intracellular total FFA content (339 mg/l vs 342 mg/l); in particular, the content of C12 and C14 FFAs was about 3–5 fold, and the content of C18:1 and C18:2 FFAs was about 8–42 fold higher than that in the control E. coli and E. coli JES1017 (original acyl-ACP TE of S. pyogenes).     

Effects of human serum albumin complexed with free fatty acids on cell viability and insulin secretion in the hamster pancreatic β-cell line HIT-T15

AimsThe effects of human serum albumin (HSA) complexed with various free fatty acids (FFAs) on ß-cells have not been studied in detail. In this study, we examined the effects of HSA and its mutants on FFA-induced cell viability changes and insulin secretion from the hamster pancreatic insulinoma cell line, HIT-TI5.Main methodsCells were exposed to different FFAs in the presence of HSA or its mutants and/or bovine serum albumin (BSA) for 24 h. Cell viability, apoptosis, insulin secretion, and unbound FFA (FFA_u) levels were determined.Key findingsIn the presence of 0.1 mM HSA, palmitate and stearate induced significant cell death at 0.1 mM or higher, whereas myristate, palmitoleate, oleate, elaidate, linoleate, linoelaidate, and conjugated linoleate showed minimal changes on cell viability. Furthermore, oleate and linoleate were clearly cytoprotective against palmitate-induced cell death. The apoptosis inhibitors, cyclosporin A (csA) and the caspase inhibitor ZVAD-FMK, did not completely prevent FFA-induced cell death, although ZVAD-FMK blocked apoptosis with no differences in the presence of either HSA or BSA. In addition, insulin secretion from the cells was significantly reduced in the presence of HSA/oleate complexes. We also found differential effects of HSA mutants complexed with FFAs on cell viability.SignificanceIn summary, our results showed that saturated FFAs induced more cell death than unsaturated FFAs. Furthermore, modified HSA/FFA interactions caused by mutations of key amino acids involved in the binding of FFA to HSA resulted in changes in cell viability, suggesting a possible role of HSA polymorphism on FFA-induced changes in cellular functions.     

A Novel Antidiabetic Drug,Fasiglifam/TAK-875, Acts as an Ago-Allosteric Modulator of FFAR1

Selective free fatty acid receptor 1 (FFAR1)/GPR40 agonist fasiglifam (TAK-875), an antidiabetic drug under phase 3 development, potentiates insulin secretion in a glucose-dependent manner by activating FFAR1 expressed in pancreatic β cells. Although fasiglifam significantly improved glycemic control in type 2 diabetes patients with a minimum risk of hypoglycemia in a phase 2 study, the precise mechanisms of its potent pharmacological effects are not fully understood. Here we demonstrate that fasiglifam acts as an ago-allosteric modulator with a partial agonistic activity for FFAR1. In both Ca²⁺ influx and insulin secretion assays using cell lines and mouse islets, fasiglifam showed positive cooperativity with the FFAR1 ligand γ-linolenic acid (γ-LA). Augmentation of glucose-induced insulin secretion by fasiglifam, γ-LA, or their combination was completely abolished in pancreatic islets of FFAR1-knockout mice. In diabetic rats, the insulinotropic effect of fasiglifam was suppressed by pharmacological reduction of plasma free fatty acid (FFA) levels using a lipolysis inhibitor, suggesting that fasiglifam potentiates insulin release in conjunction with plasma FFAs in vivo. Point mutations of FFAR1 differentially affected Ca²⁺ influx activities of fasiglifam and γ-LA, further indicating that these agonists may bind to distinct binding sites. Our results strongly suggest that fasiglifam is an ago-allosteric modulator of FFAR1 that exerts its effects by acting cooperatively with endogenous plasma FFAs in human patients as well as diabetic animals. These findings contribute to our understanding of fasiglifam as an attractive antidiabetic drug with a novel mechanism of action.     

Interaction between altered insulin and lipid metabolism in CEACAM1-inactive transgenic mice

Inactivation of CEACAM1 in L-SACC1 mice by a dominant-negative transgene in liver impairs insulin clearance and increases serum free fatty acid (FFA) levels, resulting in insulin resistance. The contribution of elevated FFAs in the pathogenesis of insulin resistance is herein investigated. Treatment of L-SACC1 female mice with carnitine restored plasma FFA content. Concomitantly, it normalized insulin levels without directly regulating receptor-mediated insulin internalization and prevented glucose tolerance in these mice. Similarly, treatment with nicotinic acid, a lipolysis inhibitor, restored insulin-stimulated receptor uptake in L-SACC1 mice. Taken together, these data suggest that chronic elevation in plasma FFAs levels contributes to the regulation of insulin metabolism and action in L-SACC1 mice.     

Enzymatic purification of dihomo-γ-linolenic acid from Mortierella single-cell oil

Purification of dihomo-γ-linolenic acid (20:3n−6; DGLA) from a single-cell oil containing 39 wt.% DGLA was attempted. The process comprised: (i) non-selective hydrolysis of the oil to prepare a mixture of free fatty acids (FFAs); (ii) urea adduct fractionation of the FFA mixture to remove saturated fatty acids; and (iii) repeated selective esterification of the resulting mixture with two kinds of lipases. In the first step, Candida rugosa lipase (Lipase-OF from Meito Sangyo Co. Ltd., Aichi, Japan) was the most effective for preparation of the FFAs from the oil; 99% hydrolysis was achieved by the reaction at 40 °C for 72 h. Urea adduct fractionation of the FFA mixture removed almost completely behenic and lignoceric acids, and the content of DGLA increased from 39 to 55 wt.%. The FFAs were esterified with 2 mol equivalent of lauryl alcohol (LauOH) using C. rugosa lipase (Lipase-AY from Amano Enzyme Inc., Aichi, Japan). In consequent, DGLA was enriched to 86 wt.% in the unesterified FFA fraction. To further increase the content of DGLA, the esterification was repeated using the same lipase. Accordingly, the content of DGLA increased to 91 wt.%, but the preparation was contaminated with 3.3 wt.% γ-linolenic acid. This contaminant was removed finally by selective esterification of the FFAs with 2 mol equivalent of LauOH using Pseudomonas aeruginosa lipase. A series of procedures purified DGLA to 95 wt.% in a yield of 51% of the initial content in the single-cell oil.     

Free fatty acid and triacylglycerol forms of CLA isomers are not incorporated equally in the liver but do not lead to differences in bone density and biomarkers of bone metabolism

Few studies have compared differences between conjugated linoleic acid (CLA) in triacylglycerol (TG) and free fatty acid (FFA) form. This study assessed differences in liver incorporation, mineral mass balance, bone density, and biomarkers of bone metabolism between FFA and TG CLA diets. Rats (n=36) were fed a control (CTRL) or 1% CLA diet in FFA or TG form (1:1 mixture c9, t11: t10, c12). Liver content of c9, t11 CLA from FFA was greater than TG form and CTRL (FFA: 0.05±0.01 vs. TG: 0.02±0.01 vs. CTRL: 0.001±0.001% total fatty acids, P<0.0001). Liver t10, c12 CLA did not differ among groups (P=0.11). No diet differences among groups for growth, bone biomarkers or mass nor mineral balance were found. These findings suggest that c9, t11 CLA in FFA form is preferentially incorporated in the liver but fatty acid forms of CLA do not affect bone or mineral outcomes.     

Exploring the protective effects of Danqi Tongmai tablet on acute myocardial ischemia rats by comprehensive metabolomics profiling

BackgroundDanqi Tongmai tablet (DQTM), a combination of salvianolic acids (SA) and panax notoginsenosides (PNS), is now in phase II clinical trial developed for the treatment of cardiovascular diseases. However, the mechanisms of its protective effects through regulating endogenous metabolites remain unclear.PurposeThe purpose of this study was to explore the protective effects of DQTM on acute myocardial ischemia rats by comprehensive metabolomics profiling.Study designThe rats were divided into three groups: sham-operating, acute myocardial ischemia (AMI) and DQTM groups. The plasma and heart were collected and profiled by LC-MS based metabolomics and lipidomics. Based on the identified differential metabolites, the pathway analysis results were obtained and further validated using the network pharmacology approach.MethodsThe AMI model was induced by ligating the left anterior descending coronary artery. The metabolomics and lipidomics profiling were based on two established LC–QTOF/MS analysis methods. The raw data were processed using XCMS Online, then the differential metabolites with nonparametric t-test p value less than 0.05 were selected and identified using HMDB and METLIN. The pathway analysis was conducted using MetaboAnalyst and validated with the predicted network results obtained by BATMAN-TCM.ResultsThe metabolomics and lipidomics profiles of plasma and heart in response to AMI and DQTM were significantly different. The AMI operation had a serious influence on metabolites in heart ischemia region, while DQTM had a greater impact on lipids in heart non-ischemia region. A total of 151 differential metabolites were identified, including mainly amino acids and fatty acids. Multiple metabolic pathways were disturbed after AMI and could be restored by DQTM, of which arachidonic acid metabolism was further validated with the predicted results of network pharmacology.ConclusionThe protective effects of DQTM on acute myocardial ischemia rats could be achieved through the regulation of multiple metabolic pathways.     

Protective effect of Salvia miltiorrhiza aqueous extract on myocardium oxidative injury in ischemic–reperfusion rats

Salvia miltiorrhiza has strong antioxidative activity. They may have a strong potential as cardioprotective agents in ischemic–reperfusion injury. Experiments were carried out in Sprague–Dawley rats with myocardium ischemia reperfusion (IR). Myocardial injuries during IR were determined by changes in electrocardiogram analysis of arrhythmias, antioxidant enzyme activities, AST, CK-MB, lactate dehydrogenase (LDH) levels, and myocyte apoptosis. Results showed that S. miltiorrhiza aqueous extract (SAME) pre-treatment significantly decreased the ST-segment (ΣST₁₂₀) and myocardium MDA, AST, CK-MB, lactate dehydrogenase (LDH) levels, increased myocardium antioxidant enzyme activities, and inhibit myocardium cell apoptosis. Furthermore, the SAME pre-treatment significantly upregulated p-JAK2 and p-STAT3 protein expression, decreased myocardium TNF-α and IL-6 concentrations in IR rats. The levels of TNF-α and IL-6 were positively correlated with the changes in myocardium p-JAK2 and p-STAT3 protein expression levels in IR rats. It can be concluded that the SAME pre-treatment has anti-ischemic and anti-apoptosis activity in heart IR rats. SAME pre-treatment protects heart against IR injury, at least in part, through its stimulating effects on injury-induced deactivation of JAK2/STAT3 signaling pathway.     

Stinging Nettle (Urtica dioica L.) Attenuates FFA Induced Ceramide Accumulation in 3T3-L1 Adipocytes in an Adiponectin Dependent Manner

Objective

Excess dietary lipids result in the accumulation of lipid metabolites including ceramides that can attenuate insulin signaling. There is evidence that a botanical extract of Urtica dioica L. (stinging nettle) improves insulin action, yet the precise mechanism(s) are not known. Hence, we examined the effects of Urtica dioica L. (UT) on adipocytes.

Research Design

We investigated the effects of an ethanolic extract of UT on free fatty acid (palmitic acid) induced inhibition of insulin-stimulated Akt serine phosphorylation and modulation of ceramidase expression in 3T3-L1 adipocytes. Adipocytes were exposed to excess FFAs in the presence or absence of UT. Effects on adiponectin expression, ceramidase expression, ceramidase activity, ceramide accumulation and insulin signaling were determined.

Results

As expected, FFAs reduced adiponectin expression and increased the expression of ceramidase enzymes but not their activity. FFA also induced the accumulation of ceramides and reduced insulin-stimulated phosphorylation of Akt in adipocytes. The effects of FFA were partially reversed by UT. UT enhanced adiponectin expression and ceramidase activity in the presence of excess FFAs. UT abated ceramide accumulation and increased insulin sensitivity via enhanced Akt phosphorylation. A siRNA knockdown of adiponectin expression prevented UT from exerting positive effects on ceramidase activity but not Akt phosphorylation.

Conclusions

In adipocytes, the ability of UT to antagonize the negative effects of FFA by modulating ceramidase activity and ceramide accumulation is dependent on the presence of adiponectin. However, the ability of UT to enhance Akt phosphorylation is independent of adiponectin expression. These studies demonstrate direct effects of UT on adipocytes and suggest this botanical extract is metabolically beneficial.     

Role of free fatty acid receptors in the regulation of energy metabolism

Free fatty acids (FFAs) are energy-generating nutrients that act as signaling molecules in various cellular processes. Several orphan G protein-coupled receptors (GPCRs) that act as FFA receptors (FFARs) have been identified and play important physiological roles in various diseases. FFA ligands are obtained from food sources and metabolites produced during digestion and lipase degradation of triglyceride stores. FFARs can be grouped according to ligand profiles, depending on the length of carbon chains of the FFAs. Medium- and long-chain FFAs activate FFA1/GPR40 and FFA4/GPR120. Short-chain FFAs activate FFA2/GPR43 and FFA3/GPR41. However, only medium-chain FFAs, and not long-chain FFAs, activate GPR84 receptor. A number of pharmacological and physiological studies have shown that these receptors are expressed in various tissues and are primarily involved in energy metabolism. Because an impairment of these processes is a part of the pathology of obesity and type 2 diabetes, FFARs are considered as key therapeutic targets. Here, we reviewed recently published studies on the physiological functions of these receptors, primarily focusing on energy homeostasis.     

Identification and bioactivity of the growth inhibitors in Tuber spp. methanolic extracts

The methanolic extracts of fresh fruiting bodies of Tuber melanosporum, T. aestivum, T. magnatum and T. borchii were evaluated for their free fatty acids (FFAs) content to test their contribution to truffle allelopathic activity. Linoleic acid was the major FFA detected in all truffle species. In addition to linoleic acid, oleic acid and palmitic acid were the other abundant FFAs in the Tuber spp. extracts. Looking at the total production, T. melanosporum and T. aestivum synthesized significantly higher amounts of all identified FFA than T. magnatum and T. borchii. The allelopathic activity, based on the specific activity (EC₅₀) of the identified compounds, was evaluated in the growth of Arabidopsis thaliana, Lotus corniculatus, Melica ciliata and Silene vulgaris. Linoleic acid had the lowest specific activity and palmitic acid had the highest. The black truffle extracts had a higher inhibitory activity than white truffles, and this could be explained by the high relative amount of FFAs in them.