Fibroblast growth factor induced Ucp1 expression in preadipocytes requires PGE2 biosynthesis and glycolytic flux.

High uncoupling protein 1 (Ucp1) expression is a characteristic of differentiated brown adipocytes and is linked to adipogenic differentiation. Paracrine fibroblast growth factor 8b (FGF8b) strongly induces Ucp1 transcription in white adipocytes independent of adipogenesis. Here, we report that FGF8b and other paracrine FGFs act on brown and white preadipocytes to upregulate Ucp1 expression via a FGFR1-MEK1/2-ERK1/2 axis, independent of adipogenesis. Transcriptomic analysis revealed an upregulation of prostaglandin biosynthesis and glycolysis upon Fgf8b treatment of preadipocytes. Oxylipin measurement by LC-MS/MS in FGF8b conditioned media identified prostaglandin E2 as a putative mediator of FGF8b induced Ucp1 transcription. RNA interference and pharmacological inhibition of the prostaglandin E2 biosynthetic pathway confirmed that PGE2 is causally involved in the control over Ucp1 transcription. Importantly, impairment of or failure to induce glycolytic flux blunted the induction of Ucp1, even in the presence of PGE2 . Lastly, a screening of transcription factors identified Nrf1 and Hes1 as required regulators of FGF8b induced Ucp1 expression. Thus, we conclude that paracrine FGFs co-regulate prostaglandin and glucose metabolism to induce Ucp1 expression in a Nrf1/Hes1-dependent manner in preadipocytes, revealing a novel regulatory network in control of Ucp1 expression in a formerly unrecognized cell type.

Inflammatory macrophage memory in nonsteroidal anti-inflammatory drug-exacerbated respiratory disease.

BACKGROUND: Nonsteroidal anti-inflammatory drug-exacerbated respiratory disease (N-ERD) is a chronic inflammatory condition, which is driven by an aberrant arachidonic acid metabolism. Macrophages are major producers of arachidonic acid metabolites and subject to metabolic reprogramming, but they have been neglected in N-ERD. OBJECTIVE: This study sought to elucidate a potential metabolic and epigenetic macrophage reprogramming in N-ERD. METHODS: Transcriptional, metabolic, and lipid mediator profiles in macrophages from patients with N-ERD and healthy controls were assessed by RNA sequencing, Seahorse assays, and LC-MS/MS. Metabolites in nasal lining fluid, sputum, and plasma from patients with N-ERD (n = 15) and healthy individuals (n = 10) were quantified by targeted metabolomics analyses. Genome-wide methylomics were deployed to define epigenetic mechanisms of macrophage reprogramming in N-ERD. RESULTS: This study shows that N-ERD monocytes/macrophages exhibit an overall reduction in DNA methylation, aberrant metabolic profiles, and an increased expression of chemokines, indicative of a persistent proinflammatory activation. Differentially methylated regions in N-ERD macrophages included genes involved in chemokine signaling and acylcarnitine metabolism. Acylcarnitines were increased in macrophages, sputum, nasal lining fluid, and plasma of patients with N-ERD. On inflammatory challenge, N-ERD macrophages produced increased levels of acylcarnitines, proinflammatory arachidonic acid metabolites, cytokines, and chemokines as compared to healthy macrophages. CONCLUSIONS: Together, these findings decipher a proinflammatory metabolic and epigenetic reprogramming of macrophages in N-ERD.

Cross-Laboratory Standardization of Preclinical Lipidomics Using Differential Mobility Spectrometry and Multiple Reaction Monitoring.

Modern biomarker and translational research as well as personalized health care studies rely heavily on powerful omics' technologies, including metabolomics and lipidomics. However, to translate metabolomics and lipidomics discoveries into a high-throughput clinical setting, standardization is of utmost importance. Here, we compared and benchmarked a quantitative lipidomics platform. The employed Lipidyzer platform is based on lipid class separation by means of differential mobility spectrometry with subsequent multiple reaction monitoring. Quantitation is achieved by the use of 54 deuterated internal standards and an automated informatics approach. We investigated the platform performance across nine laboratories using NIST SRM 1950-Metabolites in Frozen Human Plasma, and three NIST Candidate Reference Materials 8231-Frozen Human Plasma Suite for Metabolomics (high triglyceride, diabetic, and African-American plasma). In addition, we comparatively analyzed 59 plasma samples from individuals with familial hypercholesterolemia from a clinical cohort study. We provide evidence that the more practical methyl-tert-butyl ether extraction outperforms the classic Bligh and Dyer approach and compare our results with two previously published ring trials. In summary, we present standardized lipidomics protocols, allowing for the highly reproducible analysis of several hundred human plasma lipids, and present detailed molecular information for potentially disease relevant and ethnicity-related materials.

Predicting and elucidating the etiology of fatty liver disease: A machine learning modeling and validation study in the IMI DIRECT cohorts.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is highly prevalent and causes serious health complications in individuals with and without type 2 diabetes (T2D). Early diagnosis of NAFLD is important, as this can help prevent irreversible damage to the liver and, ultimately, hepatocellular carcinomas. We sought to expand etiological understanding and develop a diagnostic tool for NAFLD using machine learning. METHODS AND FINDINGS: We utilized the baseline data from IMI DIRECT, a multicenter prospective cohort study of 3,029 European-ancestry adults recently diagnosed with T2D (n = 795) or at high risk of developing the disease (n = 2,234). Multi-omics (genetic, transcriptomic, proteomic, and metabolomic) and clinical (liver enzymes and other serological biomarkers, anthropometry, measures of beta-cell function, insulin sensitivity, and lifestyle) data comprised the key input variables. The models were trained on MRI-image-derived liver fat content (<5% or ≥5%) available for 1,514 participants. We applied LASSO (least absolute shrinkage and selection operator) to select features from the different layers of omics data and random forest analysis to develop the models. The prediction models included clinical and omics variables separately or in combination. A model including all omics and clinical variables yielded a cross-validated receiver operating characteristic area under the curve (ROCAUC) of 0.84 (95% CI 0.82, 0.86; p < 0.001), which compared with a ROCAUC of 0.82 (95% CI 0.81, 0.83; p < 0.001) for a model including 9 clinically accessible variables. The IMI DIRECT prediction models outperformed existing noninvasive NAFLD prediction tools. One limitation is that these analyses were performed in adults of European ancestry residing in northern Europe, and it is unknown how well these findings will translate to people of other ancestries and exposed to environmental risk factors that differ from those of the present cohort. Another key limitation of this study is that the prediction was done on a binary outcome of liver fat quantity (<5% or ≥5%) rather than a continuous one. CONCLUSIONS: In this study, we developed several models with different combinations of clinical and omics data and identified biological features that appear to be associated with liver fat accumulation. In general, the clinical variables showed better prediction ability than the complex omics variables. However, the combination of omics and clinical variables yielded the highest accuracy. We have incorporated the developed clinical models into a web interface (see: https://www.predictliverfat.org/) and made it available to the community. TRIAL REGISTRATION: ClinicalTrials.gov NCT03814915.

House dust mite drives proinflammatory eicosanoid reprogramming and macrophage effector functions.

BACKGROUND: Eicosanoid lipid mediators play key roles in type 2 immune responses, for example in allergy and asthma. Macrophages represent major producers of eicosanoids and they are key effector cells of type 2 immunity. We aimed to comprehensively track eicosanoid profiles during type 2 immune responses to house dust mite (HDM) or helminth infection and to identify mechanisms and functions of eicosanoid reprogramming in human macrophages. METHODS: We established an LC-MS/MS workflow for the quantification of 52 oxylipins to analyze mediator profiles in human monocyte-derived macrophages (MDM) stimulated with HDM and during allergic airway inflammation (AAI) or nematode infection in mice. Expression of eicosanoid enzymes was studied by qPCR and western blot and cytokine production was assessed by multiplex assays. RESULTS: Short (24 h) exposure of alveolar-like MDM (aMDM) to HDM suppressed 5-LOX expression and product formation, while triggering prostanoid (thromboxane and prostaglandin D2 and E2 ) production. This eicosanoid reprogramming was p38-dependent, but dectin-2-independent. HDM also induced proinflammatory cytokine production, but reduced granulocyte recruitment by aMDM. In contrast, high levels of cysteinyl leukotrienes (cysLTs) and 12-/15-LOX metabolites were produced in the airways during AAI or nematode infection in mice. CONCLUSION: Our findings show that a short exposure to allergens as well as ongoing type 2 immune responses are characterized by a fundamental reprogramming of the lipid mediator metabolism with macrophages representing particularly plastic responder cells. Targeting mediator reprogramming in airway macrophages may represent a viable approach to prevent pathogenic lipid mediator profiles in allergy or asthma.

Long-Term Stability of Human Plasma Metabolites during Storage at -80 °C.

Prolonged storage of biospecimen can lead to artificially altered metabolite concentrations and thus bias data analysis in metabolomics experiments. To elucidate the potential impact of long-term storage on the metabolite profile, a pooled human plasma sample was aliquoted and stored at -80 °C. During a time period of five years, 1012 of the aliquots were measured with the Biocrates AbsoluteIDQ p180 targeted-metabolomics assay at 193 time points. Modeling the concentration courses over time revealed that 55 out of 111 metabolites remained stable. The statistically significantly changed metabolites showed on average an increase or decrease of +13.7% or -14.5%, respectively. In detail, increased concentration levels were observed for amino acids (mean: + 15.4%), the sum of hexoses (+7.9%), butyrylcarnitine (+9.4%), and some phospholipids mostly with chain lengths exceeding 40 carbon atoms (mean: +18.0%). Lipids tended to exhibit decreased concentration levels with the following mean concentration changes: acylcarnitines, -12.1%; lysophosphatidylcholines, -15.1%; diacyl-phosphatidylcholines, -17.0%; acyl-alkyl-phosphatidylcholines, -13.3%; sphingomyelins, -14.8%. We conclude that storage of plasma samples at -80 °C for up to five years can lead to altered concentration levels of amino acids, acylcarnitines, glycerophospholipids, sphingomyelins, and the sum of hexoses. These alterations must be considered when analyzing metabolomics data from long-term epidemiological studies.

Metabolite ratios as potential biomarkers for type 2 diabetes: a DIRECT study.

AIMS/HYPOTHESIS: Circulating metabolites have been shown to reflect metabolic changes during the development of type 2 diabetes. In this study we examined the association of metabolite levels and pairwise metabolite ratios with insulin responses after glucose, glucagon-like peptide-1 (GLP-1) and arginine stimulation. We then investigated if the identified metabolite ratios were associated with measures of OGTT-derived beta cell function and with prevalent and incident type 2 diabetes. METHODS: We measured the levels of 188 metabolites in plasma samples from 130 healthy members of twin families (from the Netherlands Twin Register) at five time points during a modified 3 h hyperglycaemic clamp with glucose, GLP-1 and arginine stimulation. We validated our results in cohorts with OGTT data (n = 340) and epidemiological case-control studies of prevalent (n = 4925) and incident (n = 4277) diabetes. The data were analysed using regression models with adjustment for potential confounders. RESULTS: There were dynamic changes in metabolite levels in response to the different secretagogues. Furthermore, several fasting pairwise metabolite ratios were associated with one or multiple clamp-derived measures of insulin secretion (all p < 9.2 × 10-7). These associations were significantly stronger compared with the individual metabolite components. One of the ratios, valine to phosphatidylcholine acyl-alkyl C32:2 (PC ae C32:2), in addition showed a directionally consistent positive association with OGTT-derived measures of insulin secretion and resistance (p ≤ 5.4 × 10-3) and prevalent type 2 diabetes (ORVal_PC ae C32:2 2.64 [ß 0.97 ± 0.09], p = 1.0 × 10-27). Furthermore, Val_PC ae C32:2 predicted incident diabetes independent of established risk factors in two epidemiological cohort studies (HRVal_PC ae C32:2 1.57 [ß 0.45 ± 0.06]; p = 1.3 × 10-15), leading to modest improvements in the receiver operating characteristics when added to a model containing a set of established risk factors in both cohorts (increases from 0.780 to 0.801 and from 0.862 to 0.865 respectively, when added to the model containing traditional risk factors + glucose). CONCLUSIONS/INTERPRETATION: In this study we have shown that the Val_PC ae C32:2 metabolite ratio is associated with an increased risk of type 2 diabetes and measures of insulin secretion and resistance. The observed effects were stronger than that of the individual metabolites and independent of known risk factors.

Multi-omics analysis of diabetic pig lungs reveals molecular derangements underlying pulmonary complications of diabetes mellitus.

Growing evidence shows that the lung is an organ prone to injury by diabetes mellitus. However, the molecular mechanisms of these pulmonary complications have not yet been characterized comprehensively. To systematically study the effects of insulin deficiency and hyperglycaemia on the lung, we combined proteomics and lipidomics with quantitative histomorphological analyses to compare lung tissue samples from a clinically relevant pig model for mutant INS gene induced diabetes of youth (MIDY) with samples from wild-type (WT) littermate controls. Among others, the level of pulmonary surfactant-associated protein A (SFTPA1), a biomarker of lung injury, was moderately elevated. Furthermore, key proteins related to humoral immune response and extracellular matrix (ECM) organization were significantly altered in abundance. Importantly, a lipoxygenase pathway was dysregulated as indicated by a 2.5-fold reduction of polyunsaturated fatty acid lipoxygenase ALOX15 levels, associated with corresponding changes in the levels of lipids influenced by this enzyme. Our multi-omics study points to an involvement of reduced ALOX15 levels and an associated lack of eicosanoid switching as mechanisms contributing to a proinflammatory milieu in the lungs of subjects suffering from diabetes mellitus.

The association of cardiometabolic, diet and lifestyle parameters with plasma glucagon-like peptide-1: An IMI DIRECT study.

CONTEXT: The role of glucagon-like peptide-1(GLP-1) in Type 2 diabetes (T2D) and obesity is not fully understood. OBJECTIVE: We investigate the association of cardiometabolic, diet and lifestyle parameters on fasting and postprandial GLP-1 in people at risk of, or living with, T2D. METHOD: We analysed cross-sectional data from the two Innovative Medicines Initiative (IMI) Diabetes Research on Patient Stratification (DIRECT) cohorts, cohort 1(n=2127) individuals at risk of diabetes; cohort 2 (n=789) individuals with new-onset of T2D. RESULTS: Our multiple regression analysis reveals that fasting total GLP-1 is associated with an insulin resistant phenotype and observe a strong independent relationship with male sex, increased adiposity and liver fat particularly in the prediabetes population. In contrast, we showed that incremental GLP-1 decreases with worsening glycaemia, higher adiposity, liver fat, male sex and reduced insulin sensitivity in the prediabetes cohort. Higher fasting total GLP-1 was associated with a low intake of wholegrain, fruit and vegetables inpeople with prediabetes, and with a high intake of red meat and alcohol in people with diabetes. CONCLUSION: These studies provide novel insights into the association between fasting and incremental GLP-1, metabolic traits of diabetes and obesity, and dietary intake and raise intriguing questions regarding the relevance of fasting GLP-1 in the pathophysiology T2D.

Maternal hyperglycemia induces alterations in hepatic amino acid, glucose and lipid metabolism of neonatal offspring: Multi-omics insights from a diabetic pig model.

OBJECTIVE: To gain mechanistic insights into adverse effects of maternal hyperglycemia on the liver of neonates, we performed a multi-omics analysis of liver tissue from piglets developed in genetically diabetic (mutant INS gene induced diabetes of youth; MIDY) or wild-type (WT) pigs. METHODS: Proteome, metabolome and lipidome profiles of liver and clinical parameters of serum samples from 3-day-old WT piglets (n = 9) born to MIDY mothers (PHG) were compared with those of WT piglets (n = 10) born to normoglycemic mothers (PNG). Furthermore, protein-protein interaction network analysis was used to reveal highly interacting proteins that participate in the same molecular mechanisms and to relate these mechanisms with human pathology. RESULTS: Hepatocytes of PHG displayed pronounced lipid droplet accumulation, although the abundances of central lipogenic enzymes such as fatty acid-synthase (FASN) were decreased. Additionally, circulating triglyceride (TG) levels were reduced as a trend. Serum levels of non-esterified free fatty acids (NEFA) were elevated in PHG, potentially stimulating hepatic gluconeogenesis. This is supported by elevated hepatic phosphoenolpyruvate carboxykinase (PCK1) and circulating alanine transaminase (ALT) levels. Even though targeted metabolomics showed strongly elevated phosphatidylcholine (PC) levels, the abundances of multiple key enzymes involved in major PC synthesis pathways - most prominently those from the Kennedy pathway - were paradoxically reduced in PHG liver. Conversely, enzymes involved in PC excretion and breakdown such as PC-specific translocase ATP-binding cassette 4 (ABCB4) and phospholipase A2 were increased in abundance. CONCLUSIONS: Our study indicates that maternal hyperglycemia without confounding obesity induces profound molecular changes in the liver of neonatal offspring. In particular, we found evidence for stimulated gluconeogenesis and hepatic lipid accumulation independent of de novo lipogenesis. Reduced levels of PC biosynthesis enzymes and increased levels of proteins involved in PC translocation or breakdown may represent counter-regulatory mechanisms to maternally elevated PC levels. Our comprehensive multi-omics dataset provides a valuable resource for future meta-analysis studies focusing on liver metabolism in newborns from diabetic mothers.

Corrigendum: The aryl hydrocarbon receptor regulates lipid mediator production in alveolar macrophages.

[This corrects the article DOI: 10.3389/fimmu.2023.1157373.].

The aryl hydrocarbon receptor regulates lipid mediator production in alveolar macrophages.

Allergic inflammation of the airways such as allergic asthma is a major health problem with growing incidence world-wide. One cardinal feature in severe type 2-dominated airway inflammation is the release of lipid mediators of the eicosanoid family that can either promote or dampen allergic inflammation. Macrophages are key producers of prostaglandins and leukotrienes which play diverse roles in allergic airway inflammation and thus require tight control. Using RNA- and ATAC-sequencing, liquid chromatography coupled to mass spectrometry (LC-MS/MS), enzyme immunoassays (EIA), gene expression analysis and in vivo models, we show that the aryl hydrocarbon receptor (AhR) contributes to this control via transcriptional regulation of lipid mediator synthesis enzymes in bone marrow-derived as well as in primary alveolar macrophages. In the absence or inhibition of AhR activity, multiple genes of both the prostaglandin and the leukotriene pathway were downregulated, resulting in lower synthesis of prostanoids, such as prostaglandin E2 (PGE2), and cysteinyl leukotrienes, e.g., Leukotriene C4 (LTC4). These AhR-dependent genes include PTGS1 encoding for the enzyme cyclooxygenase 1 (COX1) and ALOX5 encoding for the arachidonate 5-lipoxygenase (5-LO) both of which major upstream regulators of the prostanoid and leukotriene pathway, respectively. This regulation is independent of the activation stimulus and partially also detectable in unstimulated macrophages suggesting an important role of basal AhR activity for eicosanoid production in steady state macrophages. Lastly, we demonstrate that AhR deficiency in hematopoietic but not epithelial cells aggravates house dust mite induced allergic airway inflammation. These results suggest an essential role for AhR-dependent eicosanoid regulation in macrophages during homeostasis and inflammation.

Genetic analysis of blood molecular phenotypes reveals common properties in the regulatory networks affecting complex traits.

We evaluate the shared genetic regulation of mRNA molecules, proteins and metabolites derived from whole blood from 3029 human donors. We find abundant allelic heterogeneity, where multiple variants regulate a particular molecular phenotype, and pleiotropy, where a single variant associates with multiple molecular phenotypes over multiple genomic regions. The highest proportion of share genetic regulation is detected between gene expression and proteins (66.6%), with a further median shared genetic associations across 49 different tissues of 78.3% and 62.4% between plasma proteins and gene expression. We represent the genetic and molecular associations in networks including 2828 known GWAS variants, showing that GWAS variants are more often connected to gene expression in trans than other molecular phenotypes in the network. Our work provides a roadmap to understanding molecular networks and deriving the underlying mechanism of action of GWAS variants using different molecular phenotypes in an accessible tissue.

Discovery of drug-omics associations in type 2 diabetes with generative deep-learning models.

The application of multiple omics technologies in biomedical cohorts has the potential to reveal patient-level disease characteristics and individualized response to treatment. However, the scale and heterogeneous nature of multi-modal data makes integration and inference a non-trivial task. We developed a deep-learning-based framework, multi-omics variational autoencoders (MOVE), to integrate such data and applied it to a cohort of 789 people with newly diagnosed type 2 diabetes with deep multi-omics phenotyping from the DIRECT consortium. Using in silico perturbations, we identified drug-omics associations across the multi-modal datasets for the 20 most prevalent drugs given to people with type 2 diabetes with substantially higher sensitivity than univariate statistical tests. From these, we among others, identified novel associations between metformin and the gut microbiota as well as opposite molecular responses for the two statins, simvastatin and atorvastatin. We used the associations to quantify drug-drug similarities, assess the degree of polypharmacy and conclude that drug effects are distributed across the multi-omics modalities.

Trip13 Depletion in Liver Cancer Induces a Lipogenic Response Contributing to Plin2-Dependent Mitotic Cell Death.

Aberrant energy metabolism and cell cycle regulation both critically contribute to malignant cell growth and both processes represent targets for anticancer therapy. It is shown here that depletion of the AAA+-ATPase thyroid hormone receptor interacting protein 13 (Trip13) results in mitotic cell death through a combined mechanism linking lipid metabolism to aberrant mitosis. Diminished Trip13 levels in hepatocellular carcinoma cells result in insulin-receptor-/Akt-pathway-dependent accumulation of lipid droplets, which act as functional acentriolar microtubule organizing centers disturbing mitotic spindle polarity. Specifically, the lipid-droplet-coating protein perilipin 2 (Plin2) is required for multipolar spindle formation, induction of DNA damage, and mitotic cell death. Plin2 expression in different tumor cells confers susceptibility to cell death induced by Trip13 depletion as well as treatment with paclitaxel, a spindle-interfering drug commonly used against different cancers. Thus, assessment of Plin2 levels enables the stratification of tumor responsiveness to mitosis-targeting drugs, including clinically approved paclitaxel and Trip13 inhibitors currently under development.

Four groups of type 2 diabetes contribute to the etiological and clinical heterogeneity in newly diagnosed individuals: An IMI DIRECT study.

The presentation and underlying pathophysiology of type 2 diabetes (T2D) is complex and heterogeneous. Recent studies attempted to stratify T2D into distinct subgroups using data-driven approaches, but their clinical utility may be limited if categorical representations of complex phenotypes are suboptimal. We apply a soft-clustering (archetype) method to characterize newly diagnosed T2D based on 32 clinical variables. We assign quantitative clustering scores for individuals and investigate the associations with glycemic deterioration, genetic risk scores, circulating omics biomarkers, and phenotypic stability over 36 months. Four archetype profiles represent dysfunction patterns across combinations of T2D etiological processes and correlate with multiple circulating biomarkers. One archetype associated with obesity, insulin resistance, dyslipidemia, and impaired ß cell glucose sensitivity corresponds with the fastest disease progression and highest demand for anti-diabetic treatment. We demonstrate that clinical heterogeneity in T2D can be mapped to heterogeneity in individual etiological processes, providing a potential route to personalized treatments.

Characterization of O-phosphohydroxyproline in rat {alpha}-crystallin A.

Post-translational modifications have major importance for the structure and function of a multiplicity of proteins. Phosphorylation is a widespread phenomenon among eukaryotic proteins. Whereas O-phosphorylation on the side chains of serine, threonine, and tyrosine in proteins is well known and has been studied extensively, to our knowledge the endogenous phosphorylation of hydroxyproline has not previously been reported. In the present work, we provide evidence for the first time that O-phosphohydroxyproline (Hyp(P)) is a proteinogenic amino acid. To detect Hyp(P) in proteins we generated a Hyp(P)-specific polyclonal antibody. We could identify Hyp(P) in various proteins by Western blot analysis, and we characterized the sequence position of Hyp(P) in the protein α-crystallin A by electrospray ionization-tandem mass spectrometry. Our experiments clearly demonstrate hydroxylation and subsequent phosphorylation of a proline residue in α-crystallin A in the eye as well as in heart tissue of rat.

Lipidomic Phenotyping Reveals Extensive Lipid Remodeling during Adipogenesis in Human Adipocytes.

Differentiation of preadipocytes into mature adipocytes is a highly complex cellular process. At lipidome level, the adipogenesis remains poorly characterized. To investigate the lipidomic changes during human adipogenesis, we used the LipidyzerTM assay, which quantified 743 lipid species from 11 classes. The undifferentiated human SGBS cell strain showed a heterogeneous lipid class composition with the most abundant classes, phosphatidylethanolamines (PE), phosphatidylcholines (PC), and sphingomyelins (SM). The differentiation process was accompanied by increased ceramide concentrations. After completion of differentiation around day 4, massive lipid remodeling occurred during maturation, characterized by substantial synthesis of diacylglycerols (DAG), lysophosphatidylethanolamines (LPE), PC, PE, SM, and triacylglycerols (TAG). Lipid species composition became more homogeneous during differentiation to highly concentrated saturated and monounsaturated long-chain fatty acids (LCFA), with the four most abundant being C16:0, C16:1, C18:0, and C18:1. Simultaneously, the amount of polyunsaturated and very long-chain fatty acids (VLCFA) markedly decreased. High negative correlation coefficients between PE and PC species containing VLCFA and TAG species as well as between ceramides and SM imply that PE, PC, and ceramides might have served as additional sources for TAG and SM synthesis, respectively. These results highlight the enormous remodeling at the lipid level over several lipid classes during adipogenesis.

High levels of modified ceramides are a defining feature of murine and human cancer cachexia.

BACKGROUND: Cancer cachexia (CCx) is a multifactorial energy-wasting syndrome reducing the efficiency of anti-cancer therapies, quality of life, and survival of cancer patients. In the past years, most studies focused on the identification of tumour and host-derived proteins contributing to CCx. However, there is still a lack of studies addressing the changes in bioactive lipids. The aim of this study was to identify specific lipid species as a hallmark of CCx by performing a broad range lipid analysis of plasma from well-established CCx mouse models as well as cachectic and weight stable cancer patients. METHODS: Plasma from non-cachectic (PBS-injected mice, NC26 tumour-bearing mice), pre-cachectic and cachectic mice (C26 and LLC tumour-bearing mice, ApcMin/+ mutant mice), and plasma from weight stable and cachectic patients with gastrointestinal cancer, were analysed using the Lipidyzer™ platform. In total, 13 lipid classes and more than 1100 lipid species, including sphingolipids, neutral and polar glycerolipids, were covered by the analysis. Correlation analysis between specific lipid species and readouts of CCx were performed. Lipidomics data were confirmed by gene expression analysis of metabolic organs to analyse enzymes involved in sphingolipid synthesis and degradation. RESULTS: A decrease in several lysophosphatidylcholine (LPC) species and an increase in numerous sphingolipids including sphingomyelins (SMs), ceramides (CERs), hexosyl-ceramides (HCERs) and lactosyl-ceramides (LCERs), were mutual features of CCx in both mice and cancer patients. Notably, sphingolipid levels gradually increased during cachexia development. Key enzymes involved in ceramide synthesis were elevated in liver but not in adipose, muscle, or tumour tissues, suggesting that ceramide turnover in the liver is a major contributor to elevated sphingolipid levels in CCx. LPC(16:1), LPC(20:3), SM(16:0), SM(24:1), CER(16:0), CER(24:1), HCER(16:0), and HCER(24:1) were the most consistently affected lipid species between mice and humans and correlated negatively (LPCs) or positively (SMs, CERs and HCERs) with the severity of body weight loss. CONCLUSIONS: High levels of sphingolipids, specifically ceramides and modified ceramides, are a defining feature of murine and human CCx and may contribute to tissue wasting and skeletal muscle atrophy through the inhibition of anabolic signals. The progressive increase in sphingolipids during cachexia development supports their potential as early biomarkers for CCx.

Metabolite Shifts Induced by Marathon Race Competition Differ between Athletes Based on Level of Fitness and Performance: A Substudy of the Enzy-MagIC Study.

This study compared metabolite shifts induced by training for, participation in, and recovery from a marathon race competition among athletes divided into three groups based on fitness (relative maximum oxygen uptake (VO2max)) and performance levels (net running time). Plasma samples from 76 male runners participating in the Munich Marathon were analyzed for metabolite shifts using a targeted metabolomics panel. For the entire cohort of runners, pronounced increases were measured immediately after the race for plasma concentrations of acylcarnitines (AC), the ratio (palmitoylcarnitine + stearoylcarnitine)/free carnitine that is used as a proxy for the activity of the mitochondrial enzyme carnitine palmitoyltransferase, and arginine-related metabolites, with decreases in most amino acids (AA) and phospholipids. Plasma levels of AA and phospholipids were strongly increased 24 and 72 h post-race. Post-race plasma concentrations of AC and arginine-related metabolites were higher in the low compared to top performers, indicating an accumulation of fatty acids and a reliance on protein catabolism to provide energy after the marathon event. This study showed that marathon race competition is associated with an extensive and prolonged perturbation in plasma metabolite concentrations with a strong AC signature that is greater in the slower, less aerobically fit runners. Furthermore, changes in the arginine-related metabolites were observed.