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Fatty acid (FA) metabolism is a series of processes that provide structural substances, signalling molecules and energy. Ample evidence has shown that FA uptake is mediated by plasma membrane transporters including FA transport proteins (FATPs), caveolin-1, fatty-acid translocase (FAT)/CD36, and fatty-acid binding proteins. Unlike other FA transporters, the functions of FATPs have been controversial because they contain both motifs of FA transport and fatty acyl-CoA synthetase (ACS). The widely distributed FATP4 is not a direct FA transporter but plays a predominant function as an ACS. FATP4 deficiency causes ichthyosis premature syndrome in mice and humans associated with suppression of polar lipids but an increase in neutral lipids including triglycerides (TGs). Such a shift has been extensively characterized in enterocyte-, hepatocyte-, and adipocyte-specific Fatp4-deficient mice. The mutants under obese and non-obese fatty livers induced by different diets persistently show an increase in blood non-esterified free fatty acids and glycerol indicating the lipolysis of TGs. This review also focuses on FATP4 role on regulatory networks and factors that modulate FATP4 expression in metabolic tissues including intestine, liver, muscle, and adipose tissues. Metabolic disorders especially regarding blood lipids by FATP4 deficiency in different cell types are herein discussed. Our results may be applicable to not only patients with FATP4 mutations but also represent a model of dysregulated lipid homeostasis, thus providing mechanistic insights into obesity and development of fatty liver disease., (© 2022 The Author(s).)

Total fatty acid analysis is a routine method in many areas, including lipotyping of individuals in personalized medicine, analysis of foodstuffs, and optimization of oil production in biotechnology. This analysis is commonly done by converting fatty acyl (FA) chains of intact lipids into FA methyl esters (FAMEs) and monitoring these by gas-chromatography (GC)-based methods, typically requiring at least 15 min of analysis per sample. Here, we describe a novel method that supports fast, precise and accurate absolute quantification of total FA levels in human plasma and serum samples. The method uses acid-catalyzed transesterification with 18 O-enriched H₂O (i.e., H₂ 18 O) to convert FA chains into 18 O-labeled free fatty acids. The resulting "mass-tagged" FA analytes can be specifically monitored with improved signal-to-background by 1 min of high resolution Fourier transform mass spectrometry (FTMS) on an Orbitrap-based mass spectrometer. By benchmarking to National Institute of Standards and Technology (NIST) certified standard reference materials we show that the performance of our method is comparable, and at times superior, to that of gold-standard GC-based methods. In addition, we demonstrate that the method supports the accurate quantification of FA differences in samples obtained in dietary intervention studies and also affords specific monitoring of ingested stable isotope-labeled fatty acids ( 13 C 16 -palmitate) in normoinsulinemic and hyperinsulinemic human subjects. Overall, our novel high-throughput method is generic and suitable for many application areas, spanning basic research to personalized medicine, and is particularly useful for laboratories equipped with high resolution mass spectrometers, but lacking access to GC-based instrumentation.

Interactions between the gut microbial ecosystem and host lipid homeostasis are highly relevant to host physiology and metabolic diseases. We present a comprehensive multi-omics view of the effect of intestinal microbial colonization on hepatic lipid metabolism, integrating transcriptomic, proteomic, phosphoproteomic, and lipidomic analyses of liver and plasma samples from germfree and specific pathogen-free mice. Microbes induce monounsaturated fatty acid generation by stearoyl-CoA desaturase 1 and polyunsaturated fatty acid elongation by fatty acid elongase 5, leading to significant alterations in glycerophospholipid acyl-chain profiles. A composite classification score calculated from the observed alterations in fatty acid profiles in germfree mice clearly differentiates antibiotic-treated mice from untreated controls with high sensitivity. Mechanistic investigations reveal that acetate originating from gut microbial degradation of dietary fiber serves as precursor for hepatic synthesis of C16 and C18 fatty acids and their related glycerophospholipid species that are also released into the circulation.

NOD-like receptor (NLR)X1 (NLRX1) is an ubiquitously expressed inflammasome-independent NLR that is uniquely localized in mitochondria with as yet unknown effects on metabolic diseases. Here, we report that NLRX1 is essential in regulating cellular metabolism in non-immune parenchymal hepatocytes by decreasing mitochondrial fatty acid-dependent oxidative phosphorylation (OXPHOS) and promoting glycolysis. NLRX1 loss in mice has a profound impact on the prevention of diet-induced metabolic syndrome parameters, non-alcoholic fatty liver disease (NAFLD) progression, and renal dysfunction. Despite enhanced caloric intake, NLRX1 deletion in mice fed a western diet (WD) results in protection from liver steatosis, hepatic fibrosis, obesity, insulin resistance, glycosuria and kidney dysfunction parameters independent from inflammation. While mitochondrial content was equal, NLRX1 loss in hepatocytes leads to increased fatty acid oxidation and decreased steatosis. In contrast, glycolysis was decreased in NLRX1-deficient cells versus controls. Thus, although first implicated in immune regulation, we show that NLRX1 function extends to the control of hepatocyte energy metabolism via the restriction of mitochondrial fatty acid-dependent OXPHOS and enhancement of glycolysis. As such NLRX1 may be an attractive novel therapeutic target for NAFLD and metabolic syndrome., (Copyright © 2018 Elsevier B.V. All rights reserved.)

PLA2G6 or GVIA calcium-independent PLA2 (iPLA2β) is identified as one of the NAFLD modifier genes in humans, and thought to be a target for NAFLD therapy. iPLA2β is known to play a house-keeping role in phospholipid metabolism and remodeling. However, its role in NAFLD pathogenesis has not been supported by results obtained from high-fat feeding of iPLA2β-null (PKO) mice. Unlike livers of human NAFLD and genetically obese rodents, fatty liver induced by high-fat diet is not associated with depletion of hepatic phospholipids. We therefore tested whether iPLA2β could regulate obesity and hepatic steatosis in leptin-deficient mice by cross-breeding PKO with ob/ob mice to generate ob/ob-PKO mice. Here we observed an improvement in ob/ob-PKO mice with significant reduction in serum enzymes, lipids, glucose, insulin as well as improved glucose tolerance, and reduction in islet hyperplasia. The improvement in hepatic steatosis measured by liver triglycerides, fatty acids and cholesterol esters was associated with decreased expression of PPARγ and de novo lipogenesis genes, and the reversal of β-oxidation gene expression. Notably, ob/ob livers contained depleted levels of lysophospholipids and phospholipids, and iPLA2β deficiency in ob/ob-PKO livers lowers the former, but replenished the latter particularly phosphatidylethanolamine (PE) and phosphatidylcholine (PC) that contained arachidonic (AA) and docosahexaenoic (DHA) acids. Compared with WT livers, PKO livers also contained increased PE and PC containing AA and DHA. Thus, iPLA2β deficiency protected against obesity and ob/ob fatty liver which was associated with hepatic fatty-acyl phospholipid remodeling. Our results support the deleterious role of iPLA2β in severe obesity associated NAFLD., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Nature provides an enormous diversity of lipid molecules that originate from various pathways. To gain insight into the metabolism and dynamics of lipid species, the application of stable isotope-labeled tracers combined with mass spectrometric analysis represents a perfect tool. This review provides an overview of strategies to track fatty acid, glycerophospholipid, and sphingolipid metabolism. In particular, the selection of stable isotope-labeled precursors and their mass spectrometric analysis is discussed. Furthermore, examples of metabolic studies that were performed in cell culture, animal and clinical experiments are presented., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Ursodeoxycholyl lysophosphatidylethanolamide (UDCA-LPE) is a hepatoprotectant in inhibiting apoptosis, inflammation, and hyperlipidemia in mouse models of nonalcoholic steatohepatitis (NASH). We studied the ability of UDCA-LPE to inhibit palmitate (Pal)-induced apoptosis in primary hepatocytes and delineate cytoprotective mechanisms. We showed that lipoprotection by UDCA-LPE was mediated by cAMP and was associated with increases in triglycerides (TGs) and phospholipids (PLs). An inhibitor of cAMP-effector protein kinase A partially reversed the protective effects of UDCA-LPE. Lipidomic analyses of fatty acids and PL composition revealed a shift of lipid metabolism from saturated Pal to monounsaturated and polyunsaturated fatty acids, mainly, oleate, docosapentaenoate, and docosahexaenoate. The latter two ω-3 fatty acids were particularly found in phosphatidylcholine and phosphatidylserine pools. The catalysis of Pal by stearoyl-CoA desaturase-1 (SCD-1) is a known mechanism for the channeling of Pal away from apoptosis. SCD-1 protein was upregulated during UDCA-LPE lipoprotection. SCD-1 knockdown of Pal-treated cells showed further increased apoptosis, and the extent of UDCA-LPE protection was reduced. Thus, the major mechanism of UDCA-LPE lipoprotection involved a metabolic shift from toxic saturated toward cytoprotective unsaturated fatty acids in part via SCD-1. UDCA-LPE may thus be a therapeutic agent for treatment of NASH by altering distinct pools of fatty acids for storage into TGs and PLs, and the latter may protect lipotoxicity at the membrane levels.

The synthetic fatty acid 2-hydroxyoleic acid (2OHOA) is a potent antitumor drug that we rationally designed to regulate the membrane lipid composition and structure. The lipid modifications caused by 2OHOA treatments induce important signaling changes that end up with cell death (Terés et al., 2012 [1]). One of these regulatory effects is restoration of sphingomyelin levels, which are markedly lower in cancer cells compared to normal cells (Barceló-Coblijn et al., 2011 [2]). In this study, we report another important regulatory effect of 2OHOA on cancer cell membrane composition: a large increase in 2OHOA levels, accounting for ~15% of the fatty acids present in membrane phospholipids, in human glioma (SF767 and U118) and lung cancer (A549) cells. Concomitantly, we observed marked reductions in oleic acid levels and inhibition of stearoyl-CoA desaturase. The impact of these changes on the biophysical properties of the lipid bilayer was evaluated in liposomes reconstituted from cancer cell membrane lipid extracts. Thus, 2OHOA increased the packing of ordered domains and decreased the global order of the membrane. The present results further support and extend the knowledge about the mechanism of action for 2OHOA, based on the regulation of the membrane lipid composition and structure and subsequent modulation of membrane protein-associated signaling., (Copyright © 2013 Elsevier B.V. All rights reserved.)

So far the most frequently used method for fatty acid (FA) analysis is GC coupled to flame ionization detector (FID). However, GC-FID does not allow profiling of FA synthesis and metabolism using stable isotopes. Here we present a rapid and sensitive GC-MS method for determination of fatty acid methyl esters (FAMEs). Fatty acid methylation was carried out by transesterification with acetyl-chloride and methanol. FAME separation applies a short and polar cyano-column resulting in an analysis time of 17.2min. Separation was achieved for positional and geometrical (cis/trans) isomers with chain lengths between C8 and C28. Partial overlap of FAMEs (e.g. for C20:2 (n-6) and C21:0) could be resolved using selected ion monitoring (SIM). The precisions for human plasma samples were better than 10% coefficient of variation (CV) except for very low abundant FAs and LODs were in the low femtomol range on column. The developed GC-MS method also allows quantification of conjugated FAs such as conjugated linoleic acid (CLA) isomers because lowering the derivatization temperature from 95 °C to room temperature prevented cis to trans double bond isomerization. Finally, profiling of fatty acid synthesis and metabolism was exemplified with stable isotope labeling of macrophages using fatty acid precursors or deuterated fatty acids. In summary, we present a fast and robust GC-MS method for fatty acid profiling of positional and geometrical isomers including CLAs as well as very long chain fatty acids (VLCFAs). The method is suitable for both clinical studies and basic research including application of stable isotope compounds., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The degree of fatty acid unsaturation, that is, the ratio of unsaturated versus saturated fatty acyl chains, determines membrane fluidity. Regulation of expression of the fatty acid desaturase Ole1p was hitherto the only known mechanism governing the degree of fatty acid unsaturation in Saccharomyces cerevisiae. We report a novel mechanism for the regulation of fatty acid desaturation that is based on competition between Ole1p and the glycerol-3-phosphate acyltransferase Sct1p/Gat2p for the common substrate C16:0-CoA. Deletion of SCT1 decreases the content of saturated fatty acids, whereas overexpression of SCT1 dramatically decreases the desaturation of fatty acids and affects phospholipid composition. Whereas overexpression of Ole1p increases desaturation, co-overexpression of Ole1p and Sct1p results in a fatty acid composition intermediate between those obtained upon overexpression of the enzymes separately. On the basis of these results, we propose that Sct1p sequesters C16:0-CoA into lipids, thereby shielding it from desaturation by Ole1p. Ta-king advantage of the growth defect conferred by overexpressing SCT1, we identified the acyltransferase Cst26p/Psi1p as a regulator of Sct1p activity by affecting the phosphorylation state and overexpression level of Sct1p. The level of Sct1p phosphorylation is increased when cells are supplemented with saturated fatty acids, demonstrating the physiological relevance of our findings.

Nuclear hormone receptors are transcription factors that can be activated by nutrition-derived ligands and alter the expression of various specific target genes. Stearoyl-Coenzyme A desaturase (SCD1) converts palmitic acid (16:0) to palmitoleic acid (16:1n-7) as well as stearic acid (18:0) to oleic acid (18:1n-9). At the same time, elongase 6 (ELOVL6) elongates 16:1n-7 and 18:1n-9 to vaccenic acid (18:1n-7) and eicosenoic acid (20:1n-9). We examined how synthetic selective ligands of nuclear hormone receptors alter the gene expression of hepatic enzymes in mice. In addition, we examined how the regulation of these two enzymes influences fatty acid composition of phospholipids in liver and plasma. Mice were gavaged daily for 1 week with synthetic ligands of peroxisome proliferator-activated receptor (PPAR) α, β/δ, γ, liver X receptor (LXR), retinoic acid receptor (RAR) and retinoid-X receptor (RXR) for 1 week. Phospholipids from liver and plasma were analysed using ESI-MS/MS and GC after saponification. Hepatic gene expression of SCD1 and ELOVL6 was measured using QRT-PCR. SCD1 and ELOVL6 expression increased after the gavage of LXR and RXR ligands. The analysis of fatty acid composition of total phospholipids in plasma and liver showed increased percentage contributions of the SCD1 and ELOVL6 products 18:1n-9, 18:1n-7 and 20:1n-9 after LXR and RXR ligand application. Analysis of total phospholipids from plasma and liver revealed a significant increase in monounsaturated fatty acids bound in phosphatidylcholine (PtdCho) and lysophosphatidylcholine (PtdEtn) after LXR and RXR ligand administration. Increased hepatic gene expression of SCD1 and ELOVL6 after gavage of selective RXR or LXR ligands to mice resulted in increased concentrations of their metabolic products in phospholipids of liver and plasma.

Fatp4 exhibits acyl-CoA synthetase activity and is thereby able to catalyze the activation of fatty acids for further metabolism. However, its actual function in most tissues remains unresolved, and its role in cellular fatty acid uptake is still controversial. To characterize Fatp4 functions in adipocytes in vivo, we generated a mouse line with adipocyte-specific inactivation of the Fatp4 gene (Fatp4(A-/-)). Under standard conditions mutant mice showed no phenotypical aberrance. Uptake of radiolabeled palmitic and lignoceric acid into adipose tissue of Fatp4(A-/-) mice was unchanged. When exposed to a diet enriched in long chain fatty acids, Fatp4(A-/-) mice gained more body weight compared with control mice, although they were not consuming more food. Pronounced obesity was accompanied by a thicker layer of subcutaneous fat and greater adipocyte circumference, although expression of genes involved in de novo lipogenesis was not changed. However, the increase in total fat mass was contrasted by a significant decrease in various phospholipids, sphingomyelin, and cholesteryl esters in adipocytes. Livers of Fatp4-deficient animals under a high fat diet exhibited a higher degree of fatty degeneration. Nonetheless, no evidence for changes in insulin sensitivity and adipose inflammation was found. In summary, the results of this study confirm that Fatp4 is not crucial for fatty acid uptake into adipocytes. Instead, under the condition of a diet enriched in long chain fatty acids, adipocyte-specific Fatp4 deficiency results in adipose hypertrophy and profound alterations in the metabolism of complex lipids.

Monocytes are precursors of macrophages. Here we demonstrate that macrophage colony-stimulating factor (M-CSF)-dependent differentiation of primary human monocytes from healthy volunteers induces transcription of SREBP-1c target genes required for fatty acid (FA) biosynthesis and impairs transcription of SREBP-2 target genes required for cholesterol synthesis. Detailed lipid metabolic profiling showed that this transcriptional regulation leads to a dramatically increased fatty acid synthesis as driving force for enhanced phospholipid synthesis. During cell differentiation the major lipid class switches from cholesterol in monocytes to phosphatidylcholine in macrophages. Ultrastructural analysis revealed that this transcriptional and metabolic regulation is essential for development of macrophage filopodia and cellular organelles including primary lysosomes, endoplasmic reticulum, and Golgi network. Additional functional studies showed that suppression of fatty acid synthesis prevents phagocytosis representing a central macrophage function. Therefore induction of fatty acid synthesis is a key requirement for phagocyte development and function.

The fatty acid transport protein family is a group of evolutionarily conserved proteins that are involved in the cellular uptake and metabolism of long and very long chain fatty acids. However, little is known about their respective physiological roles. To analyze the functional significance of fatty acid transport protein 4 (Fatp4, Slc27a4), we generated mice with a targeted disruption of the Fatp4 gene. Fatp4-null mice displayed features of a neonatally lethal restrictive dermopathy. Their skin was characterized by hyperproliferative hyperkeratosis with a disturbed epidermal barrier, a flat dermal-epidermal junction, a reduced number of pilo-sebaceous structures, and a compact dermis. The rigid skin consistency resulted in an altered body shape with facial dysmorphia, generalized joint flexion contractures, and impaired movement including suckling and breathing deficiencies. Lipid analysis demonstrated a disturbed fatty acid composition of epidermal ceramides, in particular a decrease in the C26:0 and C26:0-OH fatty acid substitutes. These findings reveal a previously unknown, essential function of Fatp4 in the formation of the epidermal barrier.

Background/Objectives: Inflammatory bowel disease (IBD) is a chronic immune-mediated pathology associated with the dysregulation of lipid metabolism. The administration of nervonic acid, a very long-chain fatty acid, has been shown to improve colonic inflammation in a mouse model of colitis. Our study aimed to quantify fecal levels of nervonic acid, as well as the very long-chain fatty acids, lignoceric acid, and pentacosanoic acid, to identify associations with IBD activity. Methods: Stool samples were collected from 62 patients with IBD and 17 healthy controls. Nervonic acid, lignoceric acid, and pentacosanoic acid were quantified by gas chromatography coupled with mass spectrometry (GC-MS). Lipid levels, normalized to the dry weight of fecal homogenates, were used for calculations. Results: Patients with IBD exhibited elevated fecal nervonic acid levels compared to healthy controls, with no significant differences observed between ulcerative colitis and Crohn's disease. A fecal nervonic acid concentration of 0.49 µmol/g distinguished IBD patients from controls, achieving a sensitivity of 71% and a specificity of 82%. Fecal nervonic acid levels showed a positive correlation with both C-reactive protein and fecal calprotectin and increased proportionally with rising fecal calprotectin levels. IBD patients treated with corticosteroids or interleukin-12/23 antibodies had higher levels of fecal nervonic acid than those in other therapies, with no difference in serum C-reactive protein and calprotectin levels between these groups. Conclusions: In summary, this analysis indicates that fecal nervonic acid may emerge as a novel specific biomarker for IBD diagnosis and disease monitoring. [ABSTRACT FROM AUTHOR]

There is general consent that with decreasing bone mineral density the amount of marrow adipose tissue increases. While image-based techniques, claim an increase in saturated fatty acids responsible for this effect, this study shows an increase in both saturated and unsaturated fatty acids in the bone marrow. Using fatty acid methyl ester gas chromatography-mass spectrometry, characteristic fatty acid patterns for patients with normal BMD (N = 9), osteopenia (N = 12), and osteoporosis (N = 9) have been identified, which differ between plasma, red bone marrow and yellow bone marrow. Selected fatty acids, e.g. FA10:0, FA14:1, or FA16:1 n-7 in the bone marrow or FA18:0, FA18:1 n-9, FA18:1 n-7, FA20:0, FA20:1 n-9, or FA20:3 n-6 in the plasma, correlated with osteoclast activity, suggesting a possible mechanism how these fatty acids may interfere with BMD. Although several fatty acids correlated well with the osteoclast activity and BMD, there was not a single fatty acid contained in our fatty acid profile that can be claimed for controlling BMD, a fact that may be attributed to the genetic heterogeneity of the patients. [ABSTRACT FROM AUTHOR]

Background and Purpose: The analysis of human faecal metabolites can provide an insight into metabolic interactions between gut microbiota and the host organism. The creation of metabolic profiles in faeces has received little attention until now, and reference values, especially in the context of dietary and therapeutic interventions, are missing. Exposure to xenobiotics significantly affects the physiology of the microbiome, and microbiota manipulation and short‐chain fatty acid administration have been proposed as treatment targets for several diseases. The aim of the present study is to give concomitant concentration ranges of faecal sterol species, bile acids and short‐chain fatty acids, based on a large cohort. Experimental Approach Sterol species, bile acids and short‐chain fatty acids in human faeces from 165 study participants were quantified by LC–MS/MS. For standardization, we refer all values to dry weight of faeces. Based on the individual intestinal sterol conversion, we classified participants into low and high converters according to their coprostanol/cholesterol ratio. Key Results: Low converters excrete more straight‐chain fatty acids and bile acids than high converters; 5th and 95th percentile and median of bile acids and short‐chain fatty acids were calculated for both groups. Conclusion and Implications: We give concentration ranges for 16 faecal metabolites that can serve as reference values. Patient stratification into high or low sterol converter groups is associated with significant differences in faecal metabolites with biological activities. Such stratification should then allow better assessment of faecal metabolites before therapeutic interventions. LINKED ARTICLES: This article is part of a themed issue on Oxysterols, Lifelong Health and Therapeutics. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.16/issuetoc [ABSTRACT FROM AUTHOR]

Background: Plasmalogens are either phosphatidylcholine (PC P) or phosphatidylethanolamine (PE P) glycerophospholipids containing a vinyl ether moiety in sn-1-position and an esterified fatty acid in sn-2 position. Multiple functions have been proposed, including reservoir of precursors for inflammatory mediators, modulation of membrane fluidity, and anti-oxidative properties. They could therefore play a role under conditions of metabolic stress. Especially enzymatically modified LDL (eLDL) and oxidatively modified LDL (oxLDL) represent modifications of LDL that are taken up by macrophages in atherosclerotic plaques. The aim of this study was to analyze plasmalogen related effects of eLDL and oxLDL in human monocyte derived macrophages, as well as the effects of HDL3 mediated deloading. Methods: Elutriated monocytes from nine healthy donors were differentiated in vitro for four days. Macrophages were then loaded with native LDL, eLDL and oxLDL for 24h and subsequently deloaded with HDL3 for another 24h. Lipidomic and transcriptomic profiles were obtained. Results: Loading of macrophages with eLDL and oxLDL led to a transient but strong elevation of lysophosphatidylcholine (LPC) most likely through direct uptake. Only eLDL induced increased levels of total PC, presumably through an induction of PC synthesis. On the other hand treatment with oxLDL led to a significant increase in PC P. Analysis of individual lipid species showed lipoprotein and saturation specific effects for LPC, PC P and PE P species. Membrane fluidity was decreased by the large amount of FC contained in the lipoproteins, as indicated by a lower PC to FC ratio after lipoprotein loading. In contrast the observed changes in the saturated to mono-unsaturated fatty acid (SFA to MUFA) and saturated to poly-unsaturated fatty acid (SFA to PUFA) ratios in PE P could represent a cellular reaction to counteract this effect by producing more fluid membranes. Transcriptomic analysis showed considerable differences between eLDL and oxLDL treated macrophages. As a common feature of both lipoproteins we detected a strong downregulation of pathways for endogenous lipid synthesis as well as for exogenous lipid uptake. Deloading with HDL3 had only minor effects on total lipid class as well as on individual lipid species levels, most of the time not reaching significance. Interestingly treatment with HDL3 had no effect on membrane fluidity under these conditions, although incubation with HDL3 was partially able to counteract the oxLDL induced transcriptomic effects. To investigate the functional effect of lipoprotein treatment on macrophage polarization we performed surface marker flow cytometry. Under our experimental conditions oxLDL was able to partially shift the surface marker pattern towards a pro-inflammatory M1-like phenotype. This is consistent with the consumption of arachidonic acid containing PE P species in oxLDL treated cells, presumably for the synthesis of inflammatory mediators. Summary: Our findings provide novel data on the lipoprotein induced, lipidomic and transcriptomic changes in macrophages. This can help us better understand the development of metabolic, inflammatory diseases as well as improve our background knowledge on lipid biomarkers in serum. [ABSTRACT FROM AUTHOR]

Context: Besides their role in intestinal resorption of lipids, bile acids are regarded as endocrine and metabolic signaling molecules. The detailed profile of bile acid species in peripheral blood after an oral lipid tolerance test (OLTT) is unknown. Objective: We quantified the regulation of 18 bile acids after OLTT in healthy individuals. Material and methods: 100 volunteers were characterized by anthropometric and laboratory parameters and underwent OLTT. Venous blood was drawn in the fasted state (0 h) and at 2h, 4h, and 6 h after OLTT. Serum concentrations of 18 bile acids were measured by LC-MS/MS. Results: All of the 6 taurine-conjugated bile acids (TUDCA, THDCA, TCA, TCDCA, TDCA, TLCA) and all of the 6 glycine-conjugated bile acids (GUDCA, GHDCA, GCA, GCDCA, GDCA, GLCA) rose significantly at 2h and remained elevated during OLTT. Of the primary bile acids, CA remained unchanged, whereas CDCA significantly decreased at 4h. Of the secondary bile acids, DCA, UDCA and HDCA were not altered, whereas LCA decreased. There was a significant positive correlation between the intestinal feed-back regulator of bile acid synthesis FGF-19 and bile acids. This correlation seems to depend on all of the six taurine-conjugated bile acids and on GCA, GDCA, and GCDCA. Females and users of hormonal contraception displayed higher levels of taurine-conjugated bile acids. Conclusions: The novelty of the study is based on the identification of single bile acids during OLTT. LC-MS/MS-based quantification of bile acids in serum provides a reliable tool for future investigation of endocrine and metabolic effects of bile acids. [ABSTRACT FROM AUTHOR]

Background Hepatic fat accumulation with disturbed lipid homoeostasis is a hallmark of nonalcoholic fatty liver disease ( NAFLD). The bile acid phospholipid conjugate Ursodeoxycholyl lysophosphatidylethanolamide ( UDCA- LPE) is a novel anti-inflammatory agent with hepatoprotective effects in murine high-fat-diet ( HFD)-induced NAFLD. The aim of this work was to study changes in the hepatic lipidome due to UDCA- LPE. Materials and methods High fat diet mouse model, mass spectometry, RT-PCR. Results Hepatic lipid extracts of HFD mice were analysed by mass spectrometry. The results determined higher levels of total, saturated, mono- and diunsaturated fatty acids ( FA) in HFD mice, which were decreased by UDCA- LPE predominantly by the reducing the most abundant FA species palmitic acid and oleic acid. Unlike other FA species, levels of long-chain polyunsaturated fatty acids ( LCPUFA), which are composed of arachidonic acid ( ARA), eicosapentaenoic acid ( EPA) and docosahexaenoic acid ( DHA), were increased in HFD mice upon UDCA- LPE treatment, mainly due to elevated hepatic ARA pools. Analysis of hepatic phospholipids species showed a decrease in total phosphatidylcholine ( PC), especially monounsaturated PC ( PUFA- PC) levels in HFD mice. Loss of total PC was reversed due to UDCA- LPE by increasing hepatic PUFA- PC pools. Gene expression analysis showed that UDCA- LPE upregulated PPARα, a key transcriptional regulator of fatty acid oxidation, as well as downstream target genes CPT1α and AOX, which are crucially involved in mitochondrial and peroxisomal fatty acid oxidation. Conclusion UDCA- LPE modulates defective fatty acid metabolism during experimental NAFLD thereby restoring altered lipid profiles in addition to its pronounced anti-inflammatory effects. Thus, UDCA- LPE may be a promising drug candidate for the management of NAFLD. [ABSTRACT FROM AUTHOR]

Vascular and metabolic diseases cause half of total mortality in Europe. New prognostic markers would provide a valuable tool to improve outcome. First evidence supports the usefulness of plasma lipid species as easily accessible markers for certain diseases. Here we analyzed association of plasma lipid species with mortality in the Ludwigshafen Risk and Cardiovascular Health (LURIC) study. Plasma lipid species were quantified by electrospray ionization tandem mass spectrometry and Cox proportional hazards regression was applied to assess their association with total and cardiovascular mortality. Overall no differences were detected between total and cardiovascular mortality. Highly polyunsaturated phosphatidylcholine species together with lysophosphatidylcholine species and long chain saturated sphingomyelin and ceramide species seem to be associated with a protective effect. The predominantly circulating phosphatidylcholine-based as well as phosphatidylethanolamine-based ether species and phosphatidylethanolamine species were positively associated with total and cardiovascular mortality. Saturated and monounsaturated phosphatidylcholine species, especially phosphatidylcholine 32∶0 (most probably dipalmitoyl-phosphatidylcholine) and palmitate containing sphingomyelin and ceramide species showed together with 24∶1 containing sphingomyelin and ceramide species strongest positive association with mortality. A quotient of the sums of the six most protective species and the six species with the strongest positive mortality association indicated an almost 3-fold increased risk of mortality, which was higher than the hazard ratio for known risk factors in our cohort. Plasma lipid species levels and especially ratios of certain species may be valuable prognostic marker for cardiovascular and total mortality. [ABSTRACT FROM AUTHOR]

Abstract: Adiponectin protects from hepatic fat storage but adiponectin deficient mice (APN−/−) fed a standard chow do not develop liver steatosis. This indicates that other pathways might be activated to compensate for adiponectin deficiency. An unbiased and comprehensive screen was performed to identify hepatic alterations of lipid classes in these mice. APN−/− mice had decreased hepatic cholesteryl esters while active SREBP2 and systemic total cholesterol were not altered. Upregulation of cytochromes for bile acid synthesis suggests enhanced biliary cholesterol excretion. Analysis of 37 individual fatty acid species showed reduced stearate whereas total fatty acids were not altered. Total amount of triglycerides and phospholipids were equally abundant. A selective increase of monounsaturated phosphatidylcholine and phosphatidylethanolamine which positively correlate with hepatic and systemic triglycerides with the latter being elevated in APN−/− mice, was identified. Stearoyl-CoA desaturase 1 (SCD1) is involved in the synthesis of monounsaturated fatty acids and despite higher mRNA expression enzyme activity was not enhanced. Glucosylceramide postulated to contribute to liver damage was decreased. This study demonstrates that adiponectin deficiency is associated with hepatic changes in lipid classes in mice fed a standard chow which may protect from liver steatosis. [Copyright &y& Elsevier]

Abstract: Conjugated linoleic acid (CLA) isomers are dietary fatty acids that modulate gene expression in many cell types. We have previously reported that specifically trans-9,trans-11 (t9,t11)-CLA induces expression of genes involved in lipid metabolism of human macrophages. To elucidate the molecular mechanism underlying this transcriptional activation, we asked whether t9,t11-CLA affects activity of liver X receptor (LXR) α, a major regulator of macrophage lipid metabolism. Here we show that t9,t11-CLA is a regulator of LXRα. We further demonstrate that the CLA isomer induces expression of direct LXRα target genes in human primary macrophages. Knockdown of LXRα with RNA interference in THP-1 cells inhibited t9,t11-CLA mediated activation of LXRα including its target genes. To evaluate the effective concentration range of t9,t11-CLA, human primary macrophages were treated with various doses of CLA and well known natural and synthetic LXR agonists and mRNA expression of ABCA1 and ABCG1 was analyzed. Incubation of human macrophages with 10μM t9,t11-CLA led to a significant modulation of ABCA1 and ABCG1 transcription and caused enhanced cholesterol efflux to high density lipoproteins and apolipoprotein AI. In summary, these data show that t9,t11-CLA is an agonist of LXRα in human macrophages and that its effects on macrophage lipid metabolism can be attributed to transcriptional regulations associated with this nuclear receptor. [Copyright &y& Elsevier]

Abstract: The majority of the human ATP-binding cassette (ABC)-transporters function in cellular lipid trafficking and in the regulation of membrane lipid composition associating their dysfunction with human disease phenotypes related to sterol, phospholipid and fatty acid homeostasis. Based on findings from monogenetic disorders, animal models, and in vitro systems, major clues on the expression, function and cellular localization of human ABC-transporters have been gained. Here we review novel lipidomic technologies including quantitative mRNA expression monitoring by realtime RT-PCR and DNA-microarrays, lipid mass spectrometry, cellular fluorescence imaging and flow cytometry as promising tools to further define regulatory networks, lipid species patterns and subcellular domains important for ABC-transporter-mediated lipid trafficking. [Copyright &y& Elsevier]

Even though colorectal cancer (CRC) is one of the most preventable cancers, it is currently one of the deadliest. Worryingly, incidence in people <50 years has increased unexpectedly, and for unknown causes, despite the successful implementation of screening programs in the population aged >50 years. Thus, there is a need to improve early diagnosis detection strategies by identifying more precise biomarkers. In this scenario, the analysis of exosomes is given considerable attention. Previously, we demonstrated the exosome lipidome was able to classify CRC cell lines according to their malignancy. Herein, we investigated the use of the lipidome of plasma extracellular vesicles as a potential source of non-invasive biomarkers for CRC. A plasma exosome-enriched fraction was analyzed from patients undergoing colonoscopic procedure. Patients were divided into a healthy group and four pathological groups (patients with hyperplastic polyps; adenomatous polyps; invasive neoplasia (CRC patients); or hereditary non-polyposis CRC. The results showed a shift from 34:1- to 38:4-containing species in the pathological groups. We demonstrate that the ratio Σ34:1-containing species/Σ38:4-containing species has the potential to discriminate between healthy and pathological patients. Altogether, the results reinforce the utility of plasma exosome lipid fingerprint to provide new non-invasive biomarkers in a clinical context. [ABSTRACT FROM AUTHOR]

Short chain fatty acids (SCFAs) are generated by the degradation and fermentation of complex carbohydrates, (i.e., dietary fiber) by the gut microbiota relevant for microbe–host communication. Here, we present a method for the quantification of SCFAs in fecal samples by liquid chromatography tandem mass spectrometry (LC-MS/MS) upon derivatization to 3-nitrophenylhydrazones (3NPH). The method includes acetate, propionate, butyrate, and isobutyrate with a run time of 4 min. The reproducible (coefficients of variation (CV) below 10%) quantification of SCFAs in human fecal samples was achieved by the application of stable isotope labelled internal standards. The specificity was demonstrated by the introduction of a quantifier and qualifier ions. The method was applied to investigate the pre-analytic stability of SCFAs in human feces. Concentrations of SCFA may change substantially within hours; the degree and kinetics of these changes revealed huge differences between the donors. The fecal SCFA level could be preserved by the addition of organic solvents like isopropanol. An analysis of the colon content of mice either treated with antibiotics or fed with a diet containing a non-degradable and -fermentable fiber source showed decreased SCFA concentrations. In summary, this fast and reproducible method for the quantification of SCFA in fecal samples provides a valuable tool for both basic research and large-scale studies. [ABSTRACT FROM AUTHOR]

Abstract Group VIA calcium-independent phospholipase A2 (iPla2β) is among modifier genes of non-alcoholic fatty liver disease which leads to non-alcoholic steatohepatitis (NASH). Consistently, iPla2β deletion protects hepatic steatosis and obesity in genetic ob/ob and obese mice chronically fed with high-fat diet by replenishing the loss of hepatic phospholipids (PL). As mouse feeding with methionine- and choline-deficient (MCD) diet is a model of lean NASH, we tested whether iPla2β-null mice could still be protected since PL syntheses are disturbed. MCD-diet feeding of female wild-type for 5 weeks induced hepatic steatosis with a severe reduction of body and visceral fat weights concomitant with a decrease of hepatic phosphatidylcholine. These parameters were not altered in MCD-fed iPla2β-null mice. However, iPla2β deficiency attenuated MCD-induced elevation of serum transaminase activities and hepatic expression of fatty-acid translocase Cd36, fatty-acid binding protein-4, peroxisome-proliferator activated receptorγ, and HDL-uptake scavenger receptor B type 1. The reduction of lipid uptake genes was consistent with a decrease of hepatic esterified and unesterified fatty acids and cholesterol esters. On the contrary, iPla2β deficiency under MCD did not have any effects on inflammasomes and pro-inflammatory markers but exacerbated hepatic expression of myofibroblast α-smooth muscle actin and vimentin. Thus, without any rescue of PL loss, iPla2β inactivation attenuated hepatocellular injury in MCD-induced NASH with a novel mechanism of lipid uptake inhibition. Taken together, we have shown that iPla2β mediates hepatic steatosis and lipotoxicity in hepatocytes in both obese and lean NASH, but elicits exacerbated liver fibrosis in lean NASH likely by affecting other cell types. Highlights • iPLA2β inactivation elicits obesity and steatosis protection in obese NASH models. • iPLA2β inactivation did not alter hepatic phospholipids in MCD-induced lean NASH. • However, hepatocellular injury was attenuated due to suppression of Cd36 and Sr-b1. • A decrease of hepatic esterified and unesterified fatty acids and CE was observed. • iPLA2β inactivation under MCD on the contrary exacerbated hepatic fibrosis. [ABSTRACT FROM AUTHOR]

Fatty liver is closely associated with obesity and sensitizes the liver to further insults. The aims of the current study are 1) to identify lipid species changed in rodent fatty liver, 2) to analyze for possible associations of these lipids with triglycerides, cholesterol or CXCL8 which is elevated in the steatotic liver and 3) to find out whether systemic levels of these lipids are concordantly altered. Lipidomic analysis has confirmed an already reported reduction of phosphatidylcholine in the steatotic liver. Phosphatidylserine is lower and phosphatidylethanolamine tends to be diminished. Sphingomyelin levels are normal while monounsaturated ceramides and hexosylceramides are reduced. Sixteen of the 20 fatty acid species measured in the total lipid fraction are elevated while α-linolenic acid is diminished. Of note, medium chain saturated fatty acids are markedly decreased. Plasmalogen 18:0 and 18:1 species are strongly increased in the steatotic liver. None of the markedly changed individual lipid species strongly correlates with hepatic CXCL8 mRNA, triglycerides or cholesterol. About 60% of the lipids altered in fatty liver are congruently altered in serum. These data show that there are multiple changes in lipid composition in fatty liver and part of the lipids may be monitored by serum analysis. [ABSTRACT FROM AUTHOR]

Abstract: Conjugated fatty acids (CFAs) exhibit growth inhibitory effects on colon cancer in vitro and in vivo. To investigate whether the anticancerogenic potency depends on number or configuration of the conjugated double bonds, the effect of conjugated linoleic acid (CLA; C18:2) isomers and conjugated linolenic acid (CLnA; C18:3) isomers on viability and growth of HT-29 cells were compared. Low concentrations of CLnAs (<10μM) yielded a higher degree of inhibitory effects compared to CLAs (40μM). All trans-CFAs were more effective compared to cis/trans-CFAs as follows: t9,t11,t13-CLnA≥ c9,t11,t13-CLnA> t11,t13-CLA≥ t9,t11-CLA> c9,t11-CLA. The mRNA expression analysis of important genes associated with fatty acid metabolism showed an absence of ∆5-/∆6-desaturases and elongases in HT-29 cells, which was confirmed by fatty acid analysis. Using time- and dose-dependent stimulation experiments several metabolites were determined. Low concentrations of all trans-CFAs (5–20μM) led to dose-dependent increase of conjugated t/t-C16:2 formed by β-oxidation of C18 CFAs, ranging from 1–5% of total FAME. Importantly, it was found that CLnA is converted to CLA and that CLA is inter-converted (t11,t13-CLA is metabolized to c9,t11-CLA) by HT-29 cells. In summary, our study shows that growth inhibition of human cancer cells is associated with a specific cellular transcriptomic and metabolic profile of fatty acid metabolism, which might contribute to the diversified ability of CFAs as anti-cancer compounds. [Copyright &y& Elsevier]