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Abstract Lipids are the most abundant but poorly explored components of the human brain. Here, we present a lipidome map of the human brain comprising 75 regions, including 52 neocortical ones. The lipidome composition varies greatly among the brain regions, affecting 93% of the 419 analyzed lipids. These differences reflect the brain’s structural characteristics, such as myelin content (345 lipids) and cell type composition (353 lipids), but also functional traits: functional connectivity (76 lipids) and information processing hierarchy (60 lipids). Combining lipid composition and mRNA expression data further enhances functional connectivity association. Biochemically, lipids linked with structural and functional brain features display distinct lipid class distribution, unsaturation extent, and prevalence of omega-3 and omega-6 fatty acid residues. We verified our conclusions by parallel analysis of three adult macaque brains, targeted analysis of 216 lipids, mass spectrometry imaging, and lipidome assessment of sorted murine neurons.

Abstract Background Alzheimer’s disease (AD) and vascular dementia (VaD) are two of the commonest causes of dementia in the elderly. Of the myriad biomolecules implicated in dementia pathogenesis, sphingolipids have attracted relatively scant research attention despite their known involvement in multiple pathophysiological processes. The potential utility of peripheral sphingolipids as biomarkers in dementia cohorts with high concomitance of cerebrovascular diseases is also unclear. Methods Using a lipidomics platform, we performed a case–control study of plasma sphingolipids in a prospectively assessed cohort of 526 participants (non-cognitively impaired, NCI = 93, cognitively impaired = 217, AD = 166, VaD = 50) using a lipidomics platform. Results Distinct patterns of sphingolipid alterations were found in AD and VaD, namely an upregulation of d18:1 species in AD compared to downregulation of d16:1 species in VaD. In particular, GM3 d18:1/16:0 and GM3 d18:1/24:1 showed the strongest positive associations with AD. Furthermore, evaluation of sphingolipids panels showed specific combinations with higher sensitivity and specificity for classification of AD (Cer d16:1/24:0. Cer d18:1/16:0, GM3 d16:1/22:0, GM3 d18:1/16:0, SM d16:1/22:0, HexCer d18:1/18:0) and VAD (Cer d16:1/24:0, Cer d18:1/16:0, Hex2Cer d16:1/16:0, HexCer d18:1/18:0, SM d16:1/16:0, SM d16:1/20:0, SM d18:2/22:0) compared to NCI. Conclusions AD and VaD are associated with distinct changes of plasma sphingolipids, warranting further studies into underlying pathophysiological mechanisms and assessments of their potential utility as dementia biomarkers and therapeutic targets.

Abstract Untargeted lipidomics has been increasingly adopted for hypothesis generation in a biological context or discovery of disease biomarkers. Most of the current liquid chromatography mass spectrometry (LC–MS) based untargeted methodologies utilize a data dependent acquisition (DDA) approach in pooled samples for identification and MS-only acquisition for semi-quantification in individual samples. In this study, we present for the first time an untargeted lipidomic workflow that makes use of the newly implemented Quadrupole Resolved All-Ions (Q-RAI) acquisition function on the Agilent 6546 quadrupole time-of-flight (Q-TOF) mass spectrometer to acquire MS2 spectra in data independent acquisition (DIA) mode. This is followed by data processing and analysis on MetaboKit, a software enabling DDA-based spectral library construction and extraction of MS1 and MS2 peak areas, for reproducible identification and quantification of lipids in DIA analysis. This workflow was tested on lipid extracts from human plasma and showed quantification at MS1 and MS2 levels comparable to multiple reaction monitoring (MRM) targeted analysis of the same samples. Analysis of serum from Ceramide Synthase 2 (CerS2) null mice using the Q-RAI DIA workflow identified 88 lipid species significantly different between CerS2 null and wild type mice, including well-characterized changes previously associated with this phenotype. Our results show the Q-RAI DIA as a reliable option to perform simultaneous identification and reproducible relative quantification of lipids in exploratory biological studies.

BackgroundLong-term wolfberry intake as part of a healthy dietary pattern was recognized to have beneficial vascular outcomes. Characterization of the plasma lipidome may further provide comprehensive insights into pathways underlying these cardiovascular protective effects.ObjectiveWe analyzed the plasma lipidome of subjects who adhered to a healthy dietary pattern either with or without wolfberry and investigated the associations between the plasma lipidomic profile and cardiovascular health-related indicators.MethodsIn this 16-week, parallel design, randomized controlled trial, middle-aged and older adults (n = 41) were provided dietary counseling and assigned to either consume or not consume 15 g of wolfberry daily. At baseline and post-intervention, plasma lipidomics was assayed, and its relationships with classical CVD risk factors, vascular health, oxidant burden, carotenoids status, body composition, and anthropometry were examined.ResultsFrom the plasma lipidome, 427 lipid species from 26 sub-classes were quantified. In the wolfberry and control groups, significant changes were prominent for 27 and 42 lipid species, respectively (P < 0.05 with > 0.2-fold change). Fold changes for seven lipid species were also markedly different between the two groups. Examining the relationships between the plasma lipidome and CVD-related risk factors, total cholesterol revealed a marked positive correlation with 13 ceramide species, while HDL-cholesterol which was notably increased with wolfberry consumption showed a positive correlation with 10 phosphatidylcholine species. Oxidant burden, as represented by plasma 8-isoprostanes, was also inversely associated with lipidomic triglycerides and ether-triglycerides (41 species) and directly associated with hexosylceramides (eight species) and sphingomyelins (six species). There were no differential associations with CVD risk detected between groups.ConclusionCharacteristic alterations to the plasma lipidome were observed with healthy dietary pattern adherence and wolfberry consumption. An examination of these fluctuations suggests potential biochemical mechanisms that may mediate the antioxidant and cardiovascular protective effects of healthy dietary pattern adherence and wolfberry intake. This study was registered at clinicaltrials.gov as NCT0353584.

Abstract Defects in insulin processing and granule maturation are linked to pancreatic beta-cell failure during type 2 diabetes (T2D). Phosphatidylinositol transfer protein alpha (PITPNA) stimulates activity of phosphatidylinositol (PtdIns) 4-OH kinase to produce sufficient PtdIns-4-phosphate (PtdIns-4-P) in the trans-Golgi network to promote insulin granule maturation. PITPNA in beta-cells of T2D human subjects is markedly reduced suggesting its depletion accompanies beta-cell dysfunction. Conditional deletion of Pitpna in the beta-cells of Ins-Cre, Pitpna flox/flox mice leads to hyperglycemia resulting from decreasing glucose-stimulated insulin secretion (GSIS) and reducing pancreatic beta-cell mass. Furthermore, PITPNA silencing in human islets confirms its role in PtdIns-4-P synthesis and leads to impaired insulin granule maturation and docking, GSIS, and proinsulin processing with evidence of ER stress. Restoration of PITPNA in islets of T2D human subjects reverses these beta-cell defects and identify PITPNA as a critical target linked to beta-cell failure in T2D.

Lysosomal phospholipase A2 (LPLA2) is the PG to LPG converting enzyme in the BMP biosynthetic pathway, and changes in LPLA2 levels in cells affect cholesterol levels and late endosome/lysosome morphology.

Abstract Background Adaptations in lipid metabolism are essential to meet the physiological demands of pregnancy and any aberration may result in adverse outcomes for both mother and offspring. However, there is a lack of population-level studies to define the longitudinal changes of maternal circulating lipids from preconception to postpartum in relation to cardiometabolic risk factors. Methods LC-MS/MS-based quantification of 689 lipid species was performed on 1595 plasma samples collected at three time points in a preconception and longitudinal cohort, Singapore PREconception Study of long-Term maternal and child Outcomes (S-PRESTO). We mapped maternal plasma lipidomic profiles at preconception (N = 976), 26–28 weeks’ pregnancy (N = 337) and 3 months postpartum (N = 282) to study longitudinal lipid changes and their associations with cardiometabolic risk factors including pre-pregnancy body mass index, body weight changes and glycaemic traits. Results Around 56% of the lipids increased and 24% decreased in concentration in pregnancy before returning to the preconception concentration at postpartum, whereas around 11% of the lipids went through significant changes in pregnancy and their concentrations did not revert to the preconception concentrations. We observed a significant association of body weight changes with lipid changes across different physiological states, and lower circulating concentrations of phospholipids and sphingomyelins in pregnant mothers with higher pre-pregnancy BMI. Fasting plasma glucose and glycated haemoglobin (HbA1c) concentrations were lower whereas the homeostatic model assessment of insulin resistance (HOMA-IR), 2-h post-load glucose and fasting insulin concentrations were higher in pregnancy as compared to both preconception and postpartum. Association studies of lipidomic profiles with these glycaemic traits revealed their respective lipid signatures at three physiological states. Assessment of glycaemic traits in relation to the circulating lipids at preconception with a large sample size (n = 936) provided an integrated view of the effects of hyperglycaemia on plasma lipidomic profiles. We observed a distinct relationship of lipidomic profiles with different measures, with the highest percentage of significant lipids associated with HOMA-IR (58.9%), followed by fasting insulin concentration (56.9%), 2-h post-load glucose concentration (41.8%), HbA1c (36.7%), impaired glucose tolerance status (31.6%) and fasting glucose concentration (30.8%). Conclusions We describe the longitudinal landscape of maternal circulating lipids from preconception to postpartum, and a comprehensive view of trends and magnitude of pregnancy-induced changes in lipidomic profiles. We identified lipid signatures linked with cardiometabolic risk traits with potential implications both in pregnancy and postpartum life. Our findings provide insights into the metabolic adaptations and potential biomarkers of modifiable risk factors in childbearing women that may help in better assessment of cardiometabolic health, and early intervention at the preconception period. Trial registration ClinicalTrials.gov, NCT03531658.

Dysregulated transplacental lipid transfer and fetal–placental lipid metabolism affect birthweight, as does maternal hyperglycemia. As the mechanisms are unclear, we aimed to identify the lipids in umbilical cord plasma that were most associated with birthweight. Seventy-five Chinese women with singleton pregnancies recruited into the GUSTO mother–offspring cohort were selected from across the glycemic range based on a mid-gestation 75 g oral glucose tolerance test, excluding pre-existing diabetes. Cord plasma samples collected at term delivery were analyzed using targeted liquid-chromatography tandem mass-spectrometry to determine the concentrations of 404 lipid species across 17 lipid classes. The birthweights were standardized for sex and gestational age by local references, and regression analyses were adjusted for the maternal age, BMI, parity, mode of delivery, insulin treatment, and fasting/2 h glucose, with a false discovery-corrected p < 0.05 considered significant. Ten lysophosphatidylcholines (LPCs) and two lysophosphatidylethanolamines were positively associated with the birthweight percentiles, while twenty-four triacylglycerols were negatively associated with the birthweight percentiles. The topmost associated lipid was LPC 20:2 [21.28 (95%CI 12.70, 29.87) percentile increase in the standardized birthweight with each SD-unit increase in log10-transformed concentration]. Within these same regression models, maternal glycemia did not significantly associate with the birthweight percentiles. Specific fetal circulating lysophospholipids and triacylglycerols associate with birthweight independently of maternal glycemia, but a causal relationship remains to be established.

The liver has a high demand for phosphatidylcholine (PC), particularly in overnutrition, where reduced phospholipid levels have been implicated in the development of nonalcoholic fatty liver disease (NAFLD). Whether other pathways exist in addition to de novo PC synthesis that contribute to hepatic PC pools remains unknown. Here, we identified the lysophosphatidylcholine (LPC) transporter major facilitator superfamily domain containing 2A (Mfsd2a) as critical for maintaining hepatic phospholipid pools. Hepatic Mfsd2a expression was induced in patients having NAFLD and in mice in response to dietary fat via glucocorticoid receptor action. Mfsd2a liver-specific deficiency in mice (L2aKO) led to a robust nonalcoholic steatohepatitis–like (NASH-like) phenotype within just 2 weeks of dietary fat challenge associated with reduced hepatic phospholipids containing linoleic acid. Reducing dietary choline intake in L2aKO mice exacerbated liver pathology and deficiency of liver phospholipids containing polyunsaturated fatty acids (PUFAs). Treating hepatocytes with LPCs containing oleate and linoleate, two abundant blood-derived LPCs, specifically induced lipid droplet biogenesis and contributed to phospholipid pools, while LPC containing the omega-3 fatty acid docosahexaenoic acid (DHA) promoted lipid droplet formation and suppressed lipogenesis. This study revealed that PUFA-containing LPCs drive hepatic lipid droplet formation, suppress lipogenesis, and sustain hepatic phospholipid pools — processes that are critical for protecting the liver from excess dietary fat.

Acute kidney injury (AKI) is a global public health concern with high mortality and morbidity. In ischemic–reperfusion injury (IRI), a main cause of AKI, the brush border membrane of S3 proximal tubules (PT) is lost to the tubular lumen. How injured tubules reconstitute lost membrane lipids during renal recovery is not known. Here, we identified Mfsd2a, a sodium-dependent lysophosphatidylcholine (LPC) transporter, to be expressed specifically in the basolateral membrane of S3 PT. Using an in vivo activity probe for Mfsd2a, transport activity was found to be specific to the S3 PT. Mice with haploinsufficiency of Mfsd2a exhibited delayed recovery of renal function after acute IRI, with depressed urine osmolality and elevated levels of histological markers of damage, fibrosis, and inflammation, findings corroborated by transcriptomic analysis. Lipidomics revealed a deficiency in docosahexaenoic acid (DHA) containing phospholipids in Mfsd2a haploinsufficiency. Treatment of Mfsd2a haploinsufficient mice with LPC-DHA improved renal function and reduced markers of injury, fibrosis, and inflammation. Additionally, LPC-DHA treatment restored S3 brush border membrane architecture and normalized DHA-containing phospholipid content. These findings indicate that Mfsd2a-mediated transport of LPC-DHA is limiting for renal recovery after AKI and suggest that LPC-DHA could be a promising dietary supplement for improving recovery following AKI.

Abstract Transplacental docosahexaenoic-acid (DHA) supply for fetal development is regulated by placental DHA-lipid metabolism. Both maternal diabetes and obesity are linked to possible decreased fetal circulating DHA and increased placental DHA-lipids. Since myo-inositol is a promising intervention for gestational diabetes (GDM), we aimed to determine whether myo-inositol could rectify perturbations in placental DHA metabolism associated with maternal increasing glycemia and obesity and examine links with birthweight. Term placental villous explants from 17 women representing a range of BMIs and mid-gestational glycemia, were incubated with 13C-labeled-DHA for 48 h, in 0.3 µmol/L (control) or 60 µmol/L myo-inositol. Individual newly synthesized 13C-DHA-labeled lipid species were quantified by liquid-chromatography-mass-spectrometry. Compared with controls, incubation with myo-inositol decreased most 13C-DHA-lipids in placental explants from women with higher BMI or higher glycemia, but increased 13C-DHA-lipids with normal BMI or lower glycemia. Myo-inositol also increased 13C-DHA-labeled lipids in cases of lower birthweight centile, but induced decreases at higher centiles. Myo-inositol therefore lowered DHA-lipids in placenta with high basal placental DHA-lipid production (higher BMI and glycemia) but increased DHA-lipids where basal processing capacity is low. Myo-inositol thus moderates placental DHA metabolism towards a physiological mean which may in turn moderate birthweight.

Abstract Background Lipids play a vital role in health and disease, but changes to their circulating levels and the link with obesity remain poorly characterized in expecting mothers and their offspring in early childhood. Methods LC-MS/MS-based quantitation of 480 lipid species was performed on 2491 plasma samples collected at 4 time points in the mother-offspring Asian cohort GUSTO (Growing Up in Singapore Towards healthy Outcomes). These 4 time points constituted samples collected from mothers at 26–28 weeks of gestation (n=752) and 4–5 years postpartum (n=650), and their offspring at birth (n=751) and 6 years of age (n=338). Linear regression models were used to identify the pregnancy and developmental age-specific variations in the plasma lipidomic profiles, and their association with obesity risk. An independent birth cohort (n=1935), the Barwon Infant Study (BIS), comprising mother-offspring dyads of Caucasian origin was used for validation. Results Levels of 36% of the profiled lipids were significantly higher (absolute fold change > 1.5 and Padj < 0.05) in antenatal maternal circulation as compared to the postnatal phase, with phosphatidylethanolamine levels changing the most. Compared to antenatal maternal lipids, cord blood showed lower concentrations of most lipid species (79%) except lysophospholipids and acylcarnitines. Changes in lipid concentrations from birth to 6 years of age were much higher in magnitude (log2FC=−2.10 to 6.25) than the changes observed between a 6-year-old child and an adult (postnatal mother) (log2FC=−0.68 to 1.18). Associations of cord blood lipidomic profiles with birth weight displayed distinct trends compared to the lipidomic profiles associated with child BMI at 6 years. Comparison of the results between the child and adult BMI identified similarities in association with consistent trends (R 2=0.75). However, large number of lipids were associated with BMI in adults (67%) compared to the children (29%). Pre-pregnancy BMI was specifically associated with decrease in the levels of phospholipids, sphingomyelin, and several triacylglycerol species in pregnancy. Conclusions In summary, our study provides a detailed landscape of the in utero lipid environment provided by the gestating mother to the growing fetus, and the magnitude of changes in plasma lipidomic profiles from birth to early childhood. We identified the effects of adiposity on the circulating lipid levels in pregnant and non-pregnant women as well as offspring at birth and at 6 years of age. Additionally, the pediatric vs maternal overlap of the circulating lipid phenotype of obesity risk provides intergenerational insights and early opportunities to track and intervene the onset of metabolic adversities. Clinical trial registration This birth cohort is a prospective observational study, which was registered on 1 July 2010 under the identifier NCT01174875 .

Background: Metabolic disruption commonly follows Anterior Cruciate Ligament Reconstruction (ACLR) surgery. Brief exposure to low amplitude and frequency pulsed electromagnetic fields (PEMFs) has been shown to promote in vitro and in vivo murine myogeneses via the activation of a calcium–mitochondrial axis conferring systemic metabolic adaptations. This randomized-controlled pilot trial sought to detect local changes in muscle structure and function using MRI, and systemic changes in metabolism using plasma biomarker analyses resulting from ACLR, with or without accompanying PEMF therapy. Methods: 20 patients requiring ACLR were randomized into two groups either undergoing PEMF or sham exposure for 16 weeks following surgery. The operated thighs of 10 patients were exposed weekly to PEMFs (1 ​mT for 10 ​min) for 4 months following surgery. Another 10 patients were subjected to sham exposure and served as controls to allow assessment of the metabolic repercussions of ACLR and PEMF therapy. Blood samples were collected prior to surgery and at 16 weeks for plasma analyses. Magnetic resonance data were acquired at 1 and 16 weeks post-surgery using a Siemens 3T Tim Trio system. Phosphorus (31P) Magnetic Resonance Spectroscopy (MRS) was utilized to monitor changes in high-energy phosphate metabolism (inorganic phosphate (Pi), adenosine triphosphate (ATP) and phosphocreatine (PCr)) as well as markers of membrane synthesis and breakdown (phosphomonoesters (PME) and phosphodiester (PDE)). Quantitative Magnetization Transfer (qMT) imaging was used to elucidate changes in the underlying tissue structure, with T1-weighted and 2-point Dixon imaging used to calculate muscle volumes and muscle fat content. Results: Improvements in markers of high-energy phosphate metabolism including reductions in ΔPi/ATP, Pi/PCr and (Pi ​+ ​PCr)/ATP, and membrane kinetics, including reductions in PDE/ATP were detected in the PEMF-treated cohort relative to the control cohort at study termination. These were associated with reductions in the plasma levels of certain ceramides and lysophosphatidylcholine species. The plasma levels of biomarkers predictive of muscle regeneration and degeneration, including osteopontin and TNNT1, respectively, were improved, whilst changes in follistatin failed to achieve statistical significance. Liquid chromatography with tandem mass spectrometry revealed reductions in small molecule biomarkers of metabolic disruption, including cysteine, homocysteine, and methionine in the PEMF-treated cohort relative to the control cohort at study termination. Differences in measurements of force, muscle and fat volumes did not achieve statistical significance between the cohorts after 16 weeks post-ACLR. Conclusion: The detected changes suggest improvements in systemic metabolism in the post-surgical PEMF-treated cohort that accords with previous preclinical murine studies. PEMF-based therapies may potentially serve as a manner to ameliorate post-surgery metabolic disruptions and warrant future examination in more adequately powered clinical trials. The Translational Potential of this Article: Some degree of physical immobilisation must inevitably follow orthopaedic surgical intervention. The clinical paradox of such a scenario is that the regenerative potential of the muscle mitochondrial pool is silenced. The unmet need was hence a manner to maintain mitochondrial activation when movement is restricted and without producing potentially damaging mechanical stress. PEMF-based therapies may satisfy the requirement of non-invasively activating the requisite mitochondrial respiration when mobility is restricted for improved metabolic and regenerative recovery.

Patients with autosomal recessive microcephaly 15 caused by deficiency in the sodium-dependent lysophosphatidylcholine (LPC) transporter major facilitator superfamily domain–containing 2a (Mfsd2a) present with both microcephaly and hypomyelination, suggesting an important role for LPC uptake by oligodendrocytes in the process of myelination. Here we demonstrate that Mfsd2a is specifically expressed in oligodendrocyte precursor cells (OPCs) and is critical for oligodendrocyte development. Single-cell sequencing of the oligodendrocyte lineage revealed that OPCs from OPC-specific Mfsd2a-KO mice (2aOKO mice) underwent precocious differentiation into immature oligodendrocytes and impaired maturation into myelinating oligodendrocytes, correlating with postnatal brain hypomyelination. 2aOKO mice did not exhibit microcephaly, a finding consistent with the notion that microcephaly is the consequence of an absence of LPC uptake at the blood-brain barrier rather than a deficiency in OPCs. Lipidomic analysis showed that OPCs and iOLs from 2aOKO mice had significantly decreased levels of phospholipids containing omega-3 fatty acids, with a corresponding increase in unsaturated fatty acids, the latter being products of de novo synthesis governed by Srebp-1. RNA-Seq indicated activation of the Srebp-1 pathway and defective expression of regulators of oligodendrocyte development. Taken together, these findings indicate that the transport of LPCs by Mfsd2a in OPCs is important for maintaining OPC state to regulate postnatal brain myelination.

Older pregnant women have increased risks of complications including gestational diabetes and stillbirth. Carnitine palmitoyl transferase (CPT) expression declines with age in several tissues and is linked with poorer metabolic health. Mitochondrial CPTs catalyze acylcarnitine synthesis, which facilitates fatty acid oxidization as fuel. We hypothesized that the placenta, containing maternally-inherited mitochondria, shows an age-related CPT decline that lowers placental acylcarnitine synthesis, increasing vulnerability to pregnancy complications. We assessed CPT1A, CPT1B, CPT1C and CPT2 mRNA expression by qPCR in 77 placentas and quantified 10 medium and long-chain acylcarnitines by LC-MS/MS in a subset of 50 placentas. Older maternal age associated with lower expression of placental CPT1B, but not CPT1A, CPT1C or CPT2. CPT1B expression positively associated with eight acylcarnitines and CPT1C with three acylcarnitines, CPT1A negatively associated with nine acylcarnitines, while CPT2 did not associate with any acylcarnitine. Older maternal age associated with reductions in five acylcarnitines, only in those with BMI≥ 25 kg/m2, and not after adjusting for CPT1B expression. Our findings suggest that CPT1B is the main transferase for placental long-chain acylcarnitine synthesis, and age-related CPT1B decline may underlie decreased placental metabolic flexibility, potentially contributing to pregnancy complications in older women, particularly if they are overweight.

BackgroundPatients suffering from acute myocardial infarction (AMI) are at risk of secondary outcomes including major adverse cardiovascular events (MACE) and heart failure (HF). Comprehensive molecular phenotyping and cardiac imaging during the post-discharge time window may provide cues for risk stratification for the outcomes.Materials and methodsIn a prospective AMI cohort in New Zealand (N = 464), we measured plasma proteins and lipids 30 days after hospital discharge and inferred a unified partial correlation network with echocardiographic variables and established clinical biomarkers (creatinine, c-reactive protein, cardiac troponin I and natriuretic peptides). Using a network-based data integration approach (iOmicsPASS+), we identified predictive signatures of long-term secondary outcomes based on plasma protein, lipid, imaging markers and clinical biomarkers and assessed the prognostic potential in an independent cohort from Singapore (N = 190).ResultsThe post-discharge levels of plasma proteins and lipids showed strong correlations within each molecular type, reflecting concerted homeostatic regulation after primary MI events. However, the two molecular types were largely independent with distinct correlation structures with established prognostic imaging parameters and clinical biomarkers. To deal with massively correlated predictive features, we used iOmicsPASS + to identify subnetwork signatures of 211 and 189 data features (nodes) predictive of MACE and HF events, respectively (160 overlapping). The predictive features were primarily imaging parameters, including left ventricular and atrial parameters, tissue Doppler parameters, and proteins involved in extracellular matrix (ECM) organization, cell differentiation, chemotaxis, and inflammation. The network signatures contained plasma protein pairs with area-under-the-curve (AUC) values up to 0.74 for HF prediction in the validation cohort, but the pair of NT-proBNP and fibulin-3 (EFEMP1) was the best predictor (AUC = 0.80). This suggests that there were a handful of plasma proteins with mechanistic and functional roles in predisposing patients to the secondary outcomes, although they may be weaker prognostic markers than natriuretic peptides individually. Among those, the diastolic function parameter (E/e' - an indicator of left ventricular filling pressure) and two ECM proteins, EFEMP1 and follistatin-like 3 (FSTL3) showed comparable performance to NT-proBNP and outperformed left ventricular measures as benchmark prognostic factors for post-MI HF.ConclusionPost-discharge levels of E/e', EFEMP1 and FSTL3 are promising complementary markers of secondary adverse outcomes in AMI patients.

ABSTRACT The bacterial cell membrane is an interface for cell envelope synthesis, protein secretion, virulence factor assembly, and a target for host cationic antimicrobial peptides (CAMPs). To resist CAMP killing, several Gram-positive pathogens encode the multiple peptide resistance factor (MprF) enzyme that covalently attaches cationic amino acids to anionic phospholipids in the cell membrane. While E. faecalis encodes two mprF paralogs, MprF2 plays a dominant role in conferring resistance to killing by the CAMP human β-defensin 2 (hBD-2) in E. faecalis strain OG1RF. The goal of the current study is to understand the broader lipidomic and functional roles of E. faecalis mprF. We analyzed the lipid profiles of parental wild-type and mprF mutant strains and show that while ΔmprF2 and ΔmprF1 ΔmprF2 mutants completely lacked cationic lysyl-phosphatidylglycerol (L-PG), the ΔmprF1 mutant synthesized ~70% of L-PG compared to the parent. Unexpectedly, we also observed a significant reduction of PG in ΔmprF2 and ΔmprF1 ΔmprF2. In the mprF mutants, particularly ΔmprF1 ΔmprF2, the decrease in L-PG and phosphatidylglycerol (PG) is compensated by an increase in a phosphorus-containing lipid, glycerophospho-diglucosyl-diacylglycerol (GPDGDAG), and D-ala-GPDGDAG. These changes were accompanied by a downregulation of de novo fatty acid biosynthesis and an accumulation of long-chain acyl-acyl carrier proteins (long-chain acyl-ACPs), suggesting that the suppression of fatty acid biosynthesis was mediated by the transcriptional repressor FabT. Growth in chemically defined media lacking fatty acids revealed severe growth defects in the ΔmprF1 ΔmprF2 mutant strain, but not the single mutants, which was partially rescued through supplementation with palmitic and stearic acids. Changes in lipid homeostasis correlated with lower membrane fluidity, impaired protein secretion, and increased biofilm formation in both ΔmprF2 and ΔmprF1 ΔmprF2, compared to the wild type and ΔmprF1. Collectively, our findings reveal a previously unappreciated role for mprF in global lipid regulation and cellular physiology, which could facilitate the development of novel therapeutics targeting MprF. IMPORTANCE The cell membrane plays a pivotal role in protecting bacteria against external threats, such as antibiotics. Cationic phospholipids such as lysyl-phosphatidyglycerol (L-PG) resist the action of cationic antimicrobial peptides through electrostatic repulsion. Here we demonstrate that L-PG depletion has several unexpected consequences in Enterococcus faecalis, including a reduction of phosphatidylglycerol (PG), enrichment of a phosphorus-containing lipid, reduced fatty acid synthesis accompanied by an accumulation of long-chain acyl-acyl carrier proteins (long chain acyl-ACPs), lower membrane fluidity, and impaired secretion. These changes are not deleterious to the organism as long as exogenous fatty acids are available for uptake from the culture medium. Our findings suggest an adaptive mechanism involving compensatory changes across the entire lipidome upon removal of a single phospholipid modification. Such adaptations must be considered when devising antimicrobial strategies that target membrane lipids.

Patients with pancreatic cancer have a poor prognosis, more research is required to identify the disease at an earlier stage. Here, the authors use lipid profiles of blood samples and show that they can distinguish patients with pancreatic cancer from healthy controls.

Abstract Background Fetal docosahexaenoic acid (DHA) supply relies on preferential transplacental transfer, which is regulated by placental DHA lipid metabolism. Maternal hyperglycemia and obesity associate with higher birthweight and fetal DHA insufficiency but the role of placental DHA metabolism is unclear. Methods Explants from 17 term placenta were incubated with 13C-labeled DHA for 48 h, at 5 or 10 mmol/L glucose treatment, and the production of 17 individual newly synthesized 13C-DHA labeled lipids quantified by liquid chromatography mass spectrometry. Results Maternal BMI positively associated with 13C-DHA-labeled diacylglycerols, triacylglycerols, lysophospholipids, phosphatidylcholine and phosphatidylethanolamine plasmalogens, while maternal fasting glycemia positively associated with five 13C-DHA triacylglycerols. In turn, 13C-DHA-labeled phospholipids and triacylglycerols positively associated with birthweight centile. In-vitro glucose treatment increased most 13C-DHA-lipids, but decreased 13C-DHA phosphatidylethanolamine plasmalogens. However, with increasing maternal BMI, the magnitude of the glucose treatment induced increase in 13C-DHA phosphatidylcholine and 13C-DHA lysophospholipids was curtailed, with further decline in 13C-DHA phosphatidylethanolamine plasmalogens. Conversely, with increasing birthweight centile glucose treatment induced increases in 13C-DHA triacylglycerols were exaggerated, while glucose treatment induced decreases in 13C-DHA phosphatidylethanolamine plasmalogens were diminished. Conclusions Maternal BMI and glycemia increased the production of different placental DHA lipids implying impact on different metabolic pathways. Glucose-induced elevation in placental DHA metabolism is moderated with higher maternal BMI. In turn, findings of associations between many DHA lipids with birthweight suggest that BMI and glycemia promote fetal growth partly through changes in placental DHA metabolism.

The mechanisms by which platelets release sphingosine-1-phosphate (S1P) is not well characterized. Here the authors show that Mfsd2b is required for S1P release from both resting and activated platelets and that deletion of Mfsd2b impairs thrombotic functions of platelets.

Toll-like receptor 4 (TLR4)-mediated changes in macrophages reshape intracellular lipid pools to coordinate an effective innate immune response. Although this has been previously well-studied in different model systems, it remains incompletely understood in primary human macrophages. Here we report time-dependent lipidomic and transcriptomic responses to lipopolysaccharide (LPS) in primary human macrophages from healthy donors. We grouped the variation of ~200 individual lipid species measured by LC-MS/MS into eight temporal clusters. Among all other lipids, glycosphingolipids (glycoSP) and cholesteryl esters (CE) showed a sharp increase during the resolution phase (between 8h or 16h post LPS). GlycoSP, belonging to the globoside family (Gb3 and Gb4), showed the greatest inter-individual variability among all lipids quantified. Integrative network analysis between GlycoSP/CE levels and genome-wide transcripts, identified Gb4 d18:1/16:0 and CE 20:4 association with subnetworks enriched for T cell receptor signaling (PDCD1, CD86, PTPRC, CD247, IFNG) and DC-SIGN signaling (RAF1, CD209), respectively. Our findings reveal Gb3 and Gb4 globosides as sphingolipids associated with the resolution phase of inflammatory response in human macrophages.

Complexity is a fundamental feature of biological systems. Omics techniques like lipidomics can simultaneously quantify many thousands of molecules, thereby directly capturing the underlying biological complexity. However, this approach transfers the original biological complexity to the resulting datasets, posing challenges in data reduction and analysis. Aging is a prime example of a process that exhibits complex behaviour across multiple scales of biological organisation. The aging process is characterised by slow, cumulative and detrimental changes that are driven by intrinsic biological stochasticity and mediated through non-linear interactions and feedback within and between these levels of organization (ranging from metabolites, macromolecules, organelles and cells to tissue and organs). Only collectively and over long timeframes do these changes manifest as the exponential increases in morbidity and mortality that define biological aging, making aging a problem more difficult to study than the aetiologies of specific diseases. But aging’s time dependence can also be exploited to extract key insights into its underlying biology. Here we explore this idea by using data on changes in lipid composition across the lifespan of an organism to construct and test a LipidClock to predict biological age in the nematode Caenorhabdits elegans. The LipidClock consist of a feature transformation via Principal Component Analysis followed by Elastic Net regression and yields and Mean Absolute Error of 1.45 days for wild type animals and 4.13 days when applied to mutant strains with lifespans that are substantially different from that of wild type. Gompertz aging rates predicted by the LipidClock can be used to simulate survival curves that are in agreement with those from lifespan experiments.

Abstract Background There has been ongoing research impetus to uncover novel blood-based diagnostic and prognostic biomarkers for Alzheimer’s disease (AD), vascular dementia (VaD), and related cerebrovascular disease (CEVD)-associated conditions within the spectrum of vascular cognitive impairment (VCI). Sphingosine-1-phosphates (S1Ps) are signaling lipids which act on the S1PR family of cognate G-protein-coupled receptors and have been shown to modulate neuroinflammation, a process known to be involved in both neurodegenerative and cerebrovascular diseases. However, the status of peripheral S1P in AD and VCI is at present unclear. Methods We obtained baseline bloods from individuals recruited into an ongoing longitudinal cohort study who had normal cognition (N = 80); cognitive impairment, no dementia (N = 160); AD (N = 113); or VaD (N = 31), along with neuroimaging assessments of cerebrovascular diseases. Plasma samples were processed for the measurements of major S1P species: d16:1, d17:1, d18:0, and d18:1, along with pro-inflammatory cytokines interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF). Furthermore, in vitro effects of S1Ps on cytokine expression were also studied in an astrocytoma cell line and in rodent primary astrocytes. Results Of the S1Ps species measured, only d16:1 S1P was significantly reduced in the plasma of VaD, but not AD, patients, while the d18:1 to d16:1 ratios were increased in all cognitive subgroups (CIND, AD, and VaD). Furthermore, d18:1 to d16:1 ratios correlated with levels of IL-6, IL-8, and TNF. In both primary astrocytes and an astroglial cell line, treatment with d16:1 or d18:1 S1P resulted in the upregulation of mRNA transcripts of pro-inflammatory cytokines, with d18:1 showing a stronger effect than d16:1. Interestingly, co-treatment assays showed that the addition of d16:1 reduced the extent of d18:1-mediated gene expression, indicating that d16:1 may function to “fine-tune” the pro-inflammatory effects of d18:1. Conclusion Taken together, our data suggest that plasma d16:1 S1P may be useful as a diagnostic marker for VCI, while the d18:1 to d16:1 S1P ratio is an index of dysregulated S1P-mediated immunomodulation leading to chronic inflammation-associated neurodegeneration and cerebrovascular damage.

Targeted mass spectrometry enables reproducible and accurate lipid quantification but dedicated software tools to develop targeted lipidomics assays are lacking. Here, the authors develop a targeted lipidomics workbench and lipid knowledgebase for the streamlined generation of targeted assays.

Quantitative MS of human plasma lipids is a promising technology for translation into clinical applications. Current MS-based lipidomic methods rely on either direct infusion (DI) or chromatographic lipid separation methods (including reversed phase and hydrophilic interaction LC). However, the use of lipid markers in laboratory medicine is limited by the lack of reference values, largely because of considerable differences in the concentrations measured by different laboratories worldwide. These inconsistencies can be explained by the use of different sample preparation protocols, method-specific calibration procedures, and other experimental and data-reporting parameters, even when using identical starting materials. Here, we systematically investigated the roles of some of these variables in multiple approaches to lipid analysis of plasma samples from healthy adults by considering: 1) different sample introduction methods (separation vs. DI methods); 2) different MS instruments; and 3) between-laboratory differences in comparable analytical platforms. Each of these experimental variables resulted in different quantitative results, even with the inclusion of isotope-labeled internal standards for individual lipid classes. We demonstrated that appropriate normalization to commonly available reference samples (i.e., “shared references”) can largely correct for these systematic method-specific quantitative biases. Thus, to harmonize data in the field of lipidomics, in-house long-term references should be complemented by a commonly available shared reference sample, such as NIST SRM 1950, in the case of human plasma.

Maternal hyperglycemia is associated with disrupted transplacental arachidonic acid (AA) supply and eicosanoid synthesis, which contribute to adverse pregnancy outcomes. Since placental inositol is lowered with increasing glycemia, and since myo-inositol appears a promising intervention for gestational diabetes, we hypothesized that myo-inositol might rectify glucose-induced perturbations in placental AA metabolism. Term placental explants (n = 19) from women who underwent a mid-gestation oral glucose-tolerance-test were cultured with 13C-AA for 48 h in media containing glucose (5, 10 or 17 mM) and myo-inositol (0.3 or 60 µM). Newly synthesized 13C-AA-lipids were quantified by liquid-chromatography-mass-spectrometry. Increasing maternal fasting glycemia was associated with decreased proportions of 13C-AA-phosphatidyl-ethanolamines (PE, PE-P), but increased proportions of 13C-AA-triacylglycerides (TGs) relative to total placental 13C-AA lipids. This suggests altered placental AA compartmentalization towards storage and away from pools utilized for eicosanoid production and fetal AA supply. Compared to controls (5 mM glucose), 10 mM glucose treatment decreased the amount of four 13C-AA-phospholipids and eleven 13C-AA-TGs, whilst 17 mM glucose increased 13C-AA-PC-40:8 and 13C-AA-LPC. Glucose-induced alterations in all 13C-AA lipids (except PE-P-38:4) were attenuated by concurrent 60 µM myo-inositol treatment. Myo-inositol therefore rectifies some glucose-induced effects, but further studies are required to determine if maternal myo-inositol supplementation could reduce AA-associated pregnancy complications.

In the case of many bacteria, such as Escherichia coli, the composition of lipid molecules, termed the lipidome, temporally adapts to different environmental conditions and thus modifies membrane properties to permit growth and survival. Details of the relationship between the environment and lipidome composition are lacking, particularly for growing cultures under either favourable or under stress conditions. Here, we highlight compositional lipidome changes by describing the dynamics of molecular species throughout culture-growth phases. We show a steady cyclopropanation of fatty acyl chains, which acts as a driver for lipid diversity. There is a bias for the cyclopropanation of shorter fatty acyl chains (FA 16:1) over longer ones (FA 18:1), which likely reflects a thermodynamic phenomenon. Additionally, we observe a nearly two-fold increase in saturated fatty acyl chains in response to the presence of ampicillin and chloramphenicol, with consequences for membrane fluidity and elasticity, and ultimately bacterial stress tolerance. Our study provides the detailed quantitative lipidome composition of three E. coli strains across culture-growth phases and at the level of the fatty acyl chains and provides a general reference for phospholipid composition changes in response to perturbations. Thus, lipidome diversity is largely transient and the consequence of lipid synthesis and cyclopropanation.

Pregnancy complications such as maternal hyperglycemia increase perinatal mortality and morbidity, but risks are higher in males than in females. We hypothesized that fetal sex-dependent differences in placental palmitic-acid (PA) and oleic-acid (OA) metabolism influence such risks. Placental explants (n = 22) were incubated with isotope-labeled fatty acids (13C-PA or 13C-OA) for 24 or 48 h and the production of forty-seven 13C-PA lipids and thirty-seven 13C-OA lipids quantified by LCMS. Linear regression was used to investigate associations between maternal glycemia, BMI and fetal sex with 13C lipids, and between 13C lipids and birthweight centile. Placental explants from females showed greater incorporation of 13C-OA and 13C-PA into almost all lipids compared to males. Fetal sex also influenced relationships with maternal glycemia, with many 13C-OA and 13C-PA acylcarnitines, 13C-PA-diacylglycerols and 13C-PA phospholipids positively associated with glycemia in females but not in males. In contrast, several 13C-OA triacylglycerols and 13C-OA phospholipids were negatively associated with glycemia in males but not in females. Birthweight centile in females was positively associated with six 13C-PA and three 13C-OA lipids (mainly acylcarnitines) and was negatively associated with eight 13C-OA lipids, while males showed few associations. Fetal sex thus influences placental lipid metabolism and could be a key modulator of the impact of maternal metabolic health on perinatal outcomes, potentially contributing toward sex-specific adaptions in which females prioritize survival.

Quantitative analysis of bile acids in human feces can potentially help to better understand the influence of the gut microbiome and diet on human health. Feces is a highly heterogeneous sample matrix, mainly consisting of water and indigestible solid material (as plant fibers) that show high inter-individual variability. To compare bile acid concentrations among different individuals, a reliable normalization approach is needed. Here, we compared the impact of three normalization approaches, namely sample wet weight, dry weight, and protein concentration, on the absolute concentrations of fecal bile acids. Bile acid concentrations were determined in 70 feces samples from healthy humans. Our data show that bile acid concentrations normalized by the three different approaches are substantially different for each individual sample. Fecal bile acid concentrations normalized by wet weight show the narrowest distribution. Therefore, our analysis will provide the basis for the selection of a suitable normalization approach for the quantitative analysis of bile acids in feces.

Summary: Sphingolipids (SPs) have both a structural role in the cell membranes and a signaling function that regulates many cellular processes. The enormous structural diversity and low abundance of many SPs pose a challenge for their identification and quantification. Recent advances in lipidomics, in particular liquid chromatography (LC) coupled with mass spectrometry (MS), provide methods to detect and quantify many low-abundant SP species reliably. Here we use LC-MS to compile a “murine sphingolipid atlas,” containing the qualitative and quantitative distribution of 114 SPs in 21 tissues of a widely utilized wild-type laboratory mouse strain (C57BL/6). We report tissue-specific SP fingerprints, as well as sex-specific differences in the same tissue. This is a comprehensive, quantitative sphingolipidomic map of mammalian tissues collected in a systematic fashion. It will complement other tissue compendia for interrogation into the role of SP in mammalian health and disease.

Nonalcoholic fatty liver disease is currently the most common liver disease and is a leading cause of liver‐related morbidity and mortality. However, its pathogenesis remains largely unclear. We previously showed that mice deficient in mitogen‐activated protein kinase (MAPK) phosphatase 5 (MKP5) spontaneously developed insulin resistance and glucose intolerance, which are associated with visceral obesity and adipose tissue inflammation. In this study, we discovered that mice deficient in MKP5 developed more severe hepatic steatosis and steatohepatitis with age or with feeding on a high‐fat diet (HFD) compared to wild‐type (WT) mice, and this was associated with increased expression of proinflammatory cytokines and collagen genes. Increased p38 activation in MKP5 knockout (KO) liver compared to that in WT liver was detected, which contributed to increased expression of lipid droplet‐associated protein cell death‐inducing DFF45‐like effector A (CIDEA) and CIDEC/fat‐specific protein 27 but not CIDEB through activating transcription factor 2 (ATF2). In addition, MKP5 KO liver had higher peroxisome proliferator‐activated receptor gamma (PPARγ) expression compared with WT liver. On the other hand, overexpression of MKP5 or inhibition of p38 activation in hepatocytes resulted in reduced expression of PPARγ. Inhibition of p38 resulted in alleviation of hepatic steatosis in KO liver in response to HFD feeding, and this was associated with reduced expression of CIDEA, CIDEC, and proinflammatory cytokines. Conclusion: MKP5 prevents the development of nonalcoholic steatohepatitis by suppressing p38–ATF2 and p38–PPARγ to reduce hepatic lipid accumulation, inflammation, and fibrosis.

Increasingly, evidence is accumulating pointing at a protective role of a healthy diet at decreasing the risk of Alzheimer's disease. To test the effectiveness of nutritional components, the following food-derived compounds: curcumin alone (curcumin), curcumin combined with (−)epigallocatechin-3-gallate (EGCG), docosahexaenoic acid (DHA) and α-lipoic acid (ALA) (curcumin + EDA), or a combination of EGCG, DHA and ALA (EDA) were assessed in male Tg2576 transgenic mice on amyloid plaque load, amyloid levels (Aβ40/Aβ42, but not oligomers due to tissue limitations), microglial activation and memory using the contextual and cued fear conditioning test. The combination diet EDA, resulted in the strongest reduction of amyloid plaque load in both the cortical (p

Human blood is a self-regenerating lipid-rich biological fluid that is routinely collected in hospital settings. The inventory of lipid molecules found in blood plasma (plasma lipidome) offers insights into individual metabolism and physiology in health and disease. Disturbances in the plasma lipidome also occur in conditions that are not directly linked to lipid metabolism; therefore, plasma lipidomics based on MS is an emerging tool in an array of clinical diagnostics and disease management. However, challenges exist in the translation of such lipidomic data to clinical applications. These relate to the reproducibility, accuracy, and precision of lipid quantitation, study design, sample handling, and data sharing. This position paper emerged from a workshop that initiated a community-led process to elaborate and define a set of generally accepted guidelines for quantitative MS-based lipidomics of blood plasma or serum, with harmonization of data acquired on different instrumentation platforms across independent laboratories as an ultimate goal. We hope that other fields may benefit from and follow such a precedent.

In the last 2 decades, lipidomics has become one of the fastest expanding scientific disciplines in biomedical research. With an increasing number of new research groups to the field, it is even more important to design guidelines for assuring high standards of data quality. The Lipidomics Standards Initiative is a community-based endeavor for the coordination of development of these best practice guidelines in lipidomics and is embedded within the International Lipidomics Society. It is the intention of this review to highlight the most quality-relevant aspects of the lipidomics workflow, including preanalytics, sample preparation, MS, and lipid species identification and quantitation. Furthermore, this review just does not only highlights examples of best practice but also sheds light on strengths, drawbacks, and pitfalls in the lipidomic analysis workflow. While this review is neither designed to be a step-by-step protocol by itself nor dedicated to a specific application of lipidomics, it should nevertheless provide the interested reader with links and original publications to obtain a comprehensive overview concerning the state-of-the-art practices in the field.

Mycobacterium tuberculosis, the etiologic agent of human tuberculosis, is the world’s leading cause of death from an infectious disease. One of the main features of this pathogen is the complex and dynamic lipid composition of the cell envelope, which adapts to the variable host environment and defines the fate of infection by actively interacting with and modulating immune responses. However, while much has been learned about the enzymes of the numerous lipid pathways, little knowledge is available regarding the proteins and metabolic signals regulating lipid metabolism during M. tuberculosis infection. In this work, we constructed and characterized a FasR-deficient mutant in M. tuberculosis and demonstrated that FasR positively regulates fas and acpS expression. Lipidomic analysis of the wild type and mutant strains revealed complete rearrangement of most lipid components of the cell envelope, with phospholipids, mycolic acids, sulfolipids, and phthiocerol dimycocerosates relative abundance severely altered. As a consequence, replication of the mutant strain was impaired in macrophages leading to reduced virulence in a mouse model of infection. Moreover, we show that the fasR mutant resides in acidified cellular compartments, suggesting that the lipid perturbation caused by the mutation prevented M. tuberculosis inhibition of phagolysosome maturation. This study identified FasR as a novel factor involved in regulation of mycobacterial virulence and provides evidence for the essential role that modulation of lipid homeostasis plays in the outcome of M. tuberculosis infection.

Hyperlipidemia (hypertriglyceridemia, hypercholesterolemia) is a common finding in human and veterinary patients with endocrinopathies (e.g., hypothyroidism and hypercortisolism (Cushing’s syndrome; CS)). Despite emerging use of lipidomics technology in medicine, the lipid profiles of these endocrinopathies have not been evaluated and characterized in dogs. The aim of this study was to compare the serum lipidomes of dogs with naturally occurring CS or hypothyroidism with those of healthy dogs. Serum samples from 39 dogs with CS, 45 dogs with hypothyroidism, and 10 healthy beagle dogs were analyzed using a targeted lipidomics approach with liquid chromatography-mass spectrometry. There were significant differences between the lipidomes of dogs with CS, hypothyroidism, and the healthy dogs. The most significant changes were found in the lysophosphatidylcholines, lysophosphatidylethanolamines, lysophosphatidylinositols, phosphatidylcholines, phosphatidylethanolamines, phosphatidylglycerols, ceramides, and sphingosine 1-phosphates. Lipid alterations were especially pronounced in dogs with hypothyroidism. Several changes suggested a more atherogenic lipid profile in dogs with HT than in dogs with CS. In this study, we found so far unknown effects of naturally occurring hypothyroidism and CS on lipid metabolism in dogs. Our findings provide starting points to further examine differences in occurrence of atherosclerotic lesion formation between the two diseases.

Malassezia are common components of human skin, and as the dominant human skin eukaryotic microbe, they take part in complex microbe–host interactions. Other phylogenetically related fungi (including within Ustilagomycotina) communicate with their plant host through bioactive oxygenated polyunsaturated fatty acids, generally known as oxylipins, by regulating the plant immune system to increase their virulence. Oxylipins are similar in structure and function to human eicosanoids, which modulate the human immune system. This study reports the development of a highly sensitive mass-spectrometry-based method to capture and quantify bioactive oxygenated polyunsaturated fatty acids from the human skin surface and in vitro Malassezia cultures. It confirms that Malassezia are capable of synthesizing eicosanoid-like lipid mediators in vitro in a species dependent manner, many of which are found on human skin. This method enables sensitive identification and quantification of bioactive lipid mediators from human skin that may be derived from metabolic pathways shared between skin and its microbial residents. This enables better cross-disciplinary and detailed studies to dissect the interaction between Malassezia and human skin, and to identify potential intervention points to promote or abrogate inflammation and to improve human skin health.

As the lipidomics field continues to advance, self-evaluation within the community is critical. Here, we performed an interlaboratory comparison exercise for lipidomics using Standard Reference Material (SRM) 1950–Metabolites in Frozen Human Plasma, a commercially available reference material. The interlaboratory study comprised 31 diverse laboratories, with each laboratory using a different lipidomics workflow. A total of 1,527 unique lipids were measured across all laboratories and consensus location estimates and associated uncertainties were determined for 339 of these lipids measured at the sum composition level by five or more participating laboratories. These evaluated lipids detected in SRM 1950 serve as community-wide benchmarks for intra- and interlaboratory quality control and method validation. These analyses were performed using nonstandardized laboratory-independent workflows. The consensus locations were also compared with a previous examination of SRM 1950 by the LIPID MAPS consortium. While the central theme of the interlaboratory study was to provide values to help harmonize lipids, lipid mediators, and precursor measurements across the community, it was also initiated to stimulate a discussion regarding areas in need of improvement.

Cancer metabolism is associated with the enhanced lipogenesis required for rapid growth and proliferation. However, the magnitude of dysregulation of diverse lipid species still requires significant characterization, particularly in ovarian clear cell carcinoma (OCCC). Here, we have implemented a robust sample preparation workflow together with targeted LC-MS/MS to identify the lipidomic changes in formalin-fixed paraffin-embedded specimens from OCCC compared to tumor-free ovarian tissue. We quantitated 340 lipid species, representing 28 lipid classes. We observed differential regulation of diverse lipid species belonging to several glycerophospholipid classes and trihexosylceramide. A number of unsaturated lipid species were increased in OCCC, whereas saturated lipid species showed a decrease in OCCC compared to the controls. We also carried out total fatty acid analysis and observed an increase in the levels of several unsaturated fatty acids with a concomitant increase in the index of stearoyl-CoA desaturase (SCD) in OCCC. We confirmed the upregulation of SCD (the rate-limiting enzyme for the synthesis of monounsaturated fatty acids) by immunohistochemistry (IHC) assays. Hence, by carrying out a mass spectrometry analysis of archival tissue samples, we were able to provide insights into lipidomic alterations in OCCC.

Lipid mediators, small molecules involved in regulating inflammation and its resolution, are a class of lipids of wide interest as their levels in blood and tissues may be used to monitor health and disease states or the effect of new treatments. These molecules are present at low levels in biological samples, and an enrichment step is often needed for their detection. We describe a rapid and selective method that uses new low-cost molecularly imprinted (MIP) and non-imprinted (NIP) polymeric sorbents for the extraction of lipid mediators from plasma and tissue samples. The extraction process was carried out in solid-phase extraction (SPE) cartridges, manually packed with the sorbents. After extraction, lipid mediators were quantified by liquid chromatography–tandem mass spectrometry (LC–MSMS). Various parameters affecting the extraction efficiency were evaluated to achieve optimal recovery and to reduce non-specific interactions. Preliminary tests showed that MIPs, designed using the prostaglandin biosynthetic precursor arachidonic acid, could effectively enrich prostaglandins and structurally related molecules. However, for other lipid mediators, MIP and NIP displayed comparable recoveries. Under optimized conditions, the recoveries of synthetic standards ranged from 62% to 100%. This new extraction method was applied to the determination of the lipid mediators concentration in human plasma and mouse tissues and compared to other methods based on commercially available cartridges. In general, the methods showed comparable performances. In terms of structural specificity, our newly synthesized materials accomplished better retention of prostaglandins (PGs), hydroxydocosahexaenoic acid (HDoHE), HEPE, hydroxyeicosatetraenoic acids (HETE), hydroxyeicosatrienoic acid (HETrE), and polyunsaturated fatty acid (PUFA) compounds, while the commercially available Strata-X showed a higher recovery for dihydroxyeicosatetraenoic acid (diHETrEs). In summary, our results suggest that this new material can be successfully implemented for the extraction of lipid mediators from biological samples.

The human milk fat globule membrane (MFGM) contains important lipids for growing infants. Anthropometric measurements, milk samples, and infant milk intake were collected in a cohort of eleven healthy mother–infant dyads during exclusive breastfeeding from birth to six months. One hundred and sixty-six MFGM lipids were analysed using liquid chromatography-mass spectrometry, and the infant intake was calculated. The concentrations and intake were compared and associations between infant intake and growth characteristics explored. The lipid concentrations and infant intake varied widely between mother–infant dyads and between months one and three. The infant intake for many species displayed positive correlations with infant growth, particularly phospholipid species. The high variation in lipid intake is likely an important factor in infant growth, with strong correlations identified between the intake of many MFGM lipids and infant head circumference and weight. This study highlights the need for intake measurements and inclusion in cohort studies to elucidate the role of the human milk lipidome in infant growth and development.

The Singapore Genome Variation projects characterized the genetics of Singapore’s Chinese, Malay, and Indian populations. The Singapore Integrative Omics Study introduced here goes further in providing multi-omic measurements in individuals from these populations, including genetic, transcriptome, lipidome, and lifestyle data, and will facilitate the study of common diseases in Asian communities.

Sphingolipid concentrations have been associated with risk of type 2 diabetes and cardiovascular diseases. Because sphingolipids can be synthesized de novo from saturated fatty acids (SFA), dietary fatty acids may affect plasma sphingolipid concentrations. We aimed to evaluate dietary fat and protein intakes in relation to circulating sphingolipid levels. We used cross-sectional data from 2860 ethnic Chinese Singaporeans collected from 2004–2007. Nutrient intakes were estimated on the basis of a validated 159-item food frequency questionnaire. We quantified 79 molecularly distinct sphingolipids in a large-scale lipidomic evaluation from plasma samples. Higher saturated fat intake was associated with higher concentrations of 16:1;O2 sphingolipids including ceramides, monohexosylcermides, dihexosylceramides, sphingomyelins, and sphingosine 1-phosphates. Higher polyunsaturated fat intake was associated with lower plasma long-chain ceramides and long-chain monohexosylcermide concentrations. Protein intake was inversely associated with concentrations of most subclasses of sphingolipids, with the exception of sphingolipids containing a 16:1;O2 sphingoid base. Lower intake of saturated fat and higher intake of polyunsaturated fat and protein may decrease plasma concentrations of several sphingolipid classes. These findings may represent a novel biological mechanism for the impact of nutrient intakes on cardio-metabolic health.

Lipids in breastmilk play a critical role in infant growth and development. However, few studies have investigated sources of variability of both high- and low-abundant milk lipids. The objective of our study was to investigate individual and morning–evening differences in the human milk lipidome. In this study, a modified two-phase method (MTBE: Methanol 7:2) was validated for the extraction of lipids from human breastmilk. This method was then applied to samples from a group of 20 healthy women to measure inter- and intra-individual (morning versus evening) variability of the breastmilk lipidome. We report here the levels of 237 lipid species from 13 sub-classes using reversed-phase liquid chromatography mass spectrometry (RP-LCMS) and direct-infusion mass spectrometry (DI-MS). About 85% of lipid species showed stable inter-individual differences across time points. Half of lipid species showed higher concentrations in the evening compared with the morning, with phosphatidylethanolamines (PEs) and triacylglycerols (TAGs) exhibiting the largest changes. In morning and evening samples, the biological variation was greater for diacylglycerols (DAGs) and TAGs compared with phospholipids and sphingolipids, and the variation in DAGs and TAGs was greater in evening samples compared with morning samples. These results demonstrate that variation in the milk lipidome is strongly influenced by individual differences and time of day.

Lipid metabolism and receptor-mediated signaling are highly intertwined processes that cooperate to fulfill cellular functions and safeguard cellular homeostasis. Activation of Toll-like receptors (TLRs) leads to a complex cellular response, orchestrating a diverse range of inflammatory events that need to be tightly controlled. Here, we identified the GPI-anchored Sphingomyelin Phosphodiesterase, Acid-Like 3B (SMPDL3B) in a mass spectrometry screening campaign for membrane proteins co-purifying with TLRs. Deficiency of Smpdl3b in macrophages enhanced responsiveness to TLR stimulation and profoundly changed the cellular lipid composition and membrane fluidity. Increased cellular responses could be reverted by re-introducing affected ceramides, functionally linking membrane lipid composition and innate immune signaling. Finally, Smpdl3b-deficient mice displayed an intensified inflammatory response in TLR-dependent peritonitis models, establishing its negative regulatory role in vivo. Taken together, our results identify the membrane-modulating enzyme SMPDL3B as a negative regulator of TLR signaling that functions at the interface of membrane biology and innate immunity.

Meibomian gland dysfunction (MGD) is a leading cause of evaporative dry eye and ocular discomfort characterized by an unstable tear film principally attributed to afflicted delivery of lipids to the ocular surface. Herein, we elucidated longitudinal tear lipid alterations associated with disease alleviation and symptom improvement in a cohort of MGD patients undergoing eyelid-warming treatment for 12 weeks. Remarkably, eyelid-warming resulted in stark reductions in lysophospholipids (P < 0.001 for lyso-plasmalogen phosphatidylethanolamine, lysophosphatidylcholine, and lysophosphatidylinositol), as well as numerous PUFA-containing diacylglyceride species in tears, accompanied by significant increases in several PUFA-containing phospholipids. These changes in tear lipidomes suggest that eyelid-warming leads to diminished activity of tear phospholipases that preferentially target PUFA-containing phospholipids. In addition, treatment led to appreciable increases (P < 0.001) in O-acyl-ω-hydroxy-FAs (OAHFAs), which are lipid amphiphiles critical to the maintenance of tear film stability. Longitudinal changes in the tear lipids aforementioned also significantly (P < 0.05) correlated with reduced rate of ocular evaporation and improvement in ocular symptoms. The foregoing data thus indicate that excess ocular surface phospholipase activity detrimental to tear film stability could be alleviated by eyelid warming alone without application of steroids and identify tear OAHFAs as suitable markers to monitor treatment response in MGD.

Influenza virus acquires a host-derived lipid envelope during budding, yet a convergent view on the role of host lipid metabolism during infection is lacking. Using a mass spectrometry-based lipidomics approach, we provide a systems-scale perspective on membrane lipid dynamics of infected human lung epithelial cells and purified influenza virions. We reveal enrichment of the minor peroxisome-derived ether-linked phosphatidylcholines relative to bulk ester-linked phosphatidylcholines in virions as a unique pathogenicity-dependent signature for influenza not found in other enveloped viruses. Strikingly, pharmacological and genetic interference with peroxisomal and ether lipid metabolism impaired influenza virus production. Further integration of our lipidomics results with published genomics and proteomics data corroborated altered peroxisomal lipid metabolism as a hallmark of influenza virus infection in vitro and in vivo. Influenza virus may therefore tailor peroxisomal and particularly ether lipid metabolism for efficient replication.