Elevated lipid class concentrations in females with anorexia nervosa before and after intensive weight restoration treatment-A lipidomics study.

OBJECTIVE: To study the plasma lipidome of patients with anorexia nervosa (AN) before and after weight restoration treatment and report associations with AN subtypes and oral contraceptive pill (OCP) usage. METHODS: Quantitative shotgun lipidomics analysis was used to study plasma lipids of 50 female patients with AN before and after weight restoration treatment and 50 healthy female controls (HC). The AN group was assessed with blood samples and questionnaires before and after weight restoration. RESULTS: In total we quantified 260 lipid species representing 26 lipid classes of which 13 lipid class concentrations were elevated in patients with AN at admission compared with HC. Lipid classes remained elevated after weight restoration treatment of 84 days (median; interquartile range 28), and only the concentration of the ceramide lipid class increased between pre- and post-treatment (p = .03), whereas lysophosphatidylcholine (LPC, p = .02), ether-linked Phosphatidylcholine (LPCO, p = .02), and lysophosphatidylethanolamine (LPE, p = .009) decreased. CONCLUSION: In AN, 13 out of 26 lipid class concentrations were elevated at admission and remained elevated post-treatment. Ceramides increased further between pre- and post-weight restoration treatment, which could be related to the rapid weight gain during re-nutrition. Further research is needed to elucidate the effects of weight restoration treatment on short- and long-term lipid profiles in individuals with AN. PUBLIC SIGNIFICANCE STATEMENT: Lipidomics research can increase the understanding of AN, a complex and potentially life-threatening eating disorder. By analyzing lipids, or fats, in the body, we can identify biological markers that may inform diagnosis and develop more effective treatments. This research can also shed light on the underlying mechanisms of the disorder, leading to a better understanding of the processes involved in eating behavior.

Cationic amphiphilic drugs induce elevation in lysoglycerophospholipid levels and cell death in leukemia cells.

INTRODUCTION: Repurposing of cationic amphiphilic drugs (CADs) emerges as an attractive therapeutic solution against various cancers, including leukemia. CADs target lysosomal lipid metabolism and preferentially kill cancer cells via induction of lysosomal membrane permeabilization, but the exact effects of CADs on the lysosomal lipid metabolism remain poorly illuminated. OBJECTIVES: We aimed to systematically monitor CAD-induced alterations in the quantitative lipid profiles of leukemia cell lines in order to chart effects of CADs on the metabolism of various lipid classes present in these cells. METHODS: We conducted this study on eight cultured cell lines representing two leukemia types, acute lymphoblastic leukemia and acute myeloid leukemia. Mass spectrometry-based quantitative shotgun lipidomics was employed to quantify the levels of around 400 lipid species of 26 lipid classes in the leukemia cell lines treated or untreated with a CAD, siramesine. RESULTS: The two leukemia types displayed high, but variable sensitivities to CADs and distinct profiles of cellular lipids. Treatment with siramesine rapidly altered the levels of diverse lipid classes in both leukemia types. These included sphingolipid classes previously reported to play key roles in CAD-induced cell death, but also lipids of other categories. We demonstrated that the treatment with siramesine additionally elevated the levels of numerous cytolytic lysoglycerophospholipids in positive correlation with the sensitivity of individual leukemia cell lines to siramesine. CONCLUSIONS: Our study shows that CAD treatment alters balance in the metabolism of glycerophospholipids, and proposes elevation in the levels of lysoglycerophospholipids as part of the mechanism leading to CAD-induced cell death of leukemia cells.

Fenofibrate increases very-long-chain sphingolipids and improves blood glucose homeostasis in NOD mice.

AIMS/HYPOTHESIS: Sphingolipid metabolism regulates beta cell biology and inflammation and is abnormal at the onset of type 1 diabetes. Fenofibrate, a regulator of sphingolipid metabolism, is known to prevent diabetes in NOD mice. Here, we aimed to investigate the effects of fenofibrate on the pancreatic lipidome, pancreas morphology, pancreatic sympathetic nerves and blood glucose homeostasis in NOD mice. METHODS: We treated female NOD mice with fenofibrate from 3 weeks of age. The pancreatic lipidome was analysed using MS. Analysis of pancreas and islet volume was performed by stereology. Islet sympathetic nerve fibre volume was evaluated using tyrosine hydroxylase staining. The effect on blood glucose homeostasis was assessed by measuring non-fasting blood glucose from age 12 to 30 weeks. Furthermore, we measured glucose tolerance, fasting insulin and glucagon levels, and insulin tolerance. RESULTS: We found that fenofibrate selectively increases the amount of very-long-chain sphingolipids in the pancreas of NOD mice. In addition, we found that fenofibrate causes a remodelling of the pancreatic lipidome with an increased amount of lysoglycerophospholipids. Fenofibrate did not affect islet or pancreas volume, but led to a higher volume of islet sympathetic nerve fibres and tyrosine hydroxylase-positive cells. Fenofibrate-treated NOD mice had a more stable blood glucose, which was associated with reduced non-fasting and increased fasting blood glucose. Furthermore, fenofibrate improved glucose tolerance, reduced fasting glucagon levels and prevented fasting hyperinsulinaemia. CONCLUSIONS/INTERPRETATION: These data indicate that fenofibrate alters the pancreatic lipidome to a more anti-inflammatory and anti-apoptotic state. The beneficial effects on islet sympathetic nerve fibres and blood glucose homeostasis indicate that fenofibrate could be used as a therapeutic approach to improve blood glucose homeostasis and prevent diabetes-associated pathologies.

Endocannabinoids are conserved inhibitors of the Hedgehog pathway.

Hedgehog ligands control tissue development and homeostasis by alleviating repression of Smoothened, a seven-pass transmembrane protein. The Hedgehog receptor, Patched, is thought to regulate the availability of small lipophilic Smoothened repressors whose identity is unknown. Lipoproteins contain lipids required to repress Smoothened signaling in vivo. Here, using biochemical fractionation and lipid mass spectrometry, we identify these repressors as endocannabinoids. Endocannabinoids circulate in human and Drosophila lipoproteins and act directly on Smoothened at physiological concentrations to repress signaling in Drosophila and mammalian assays. Phytocannabinoids are also potent Smo inhibitors. These findings link organismal metabolism to local Hedgehog signaling and suggest previously unsuspected mechanisms for the physiological activities of cannabinoids.

Quantitative profiling of endocannabinoids in lipoproteins by LC-MS/MS.

Endocannabinoids belong to a diverse family of endogenous lipid bioregulators acting as physiological ligands of cannabinoid receptor type 1 and cannabinoid receptor type 2 in the central and peripheral nervous system. They are also present in nmol L(-1) concentrations in human blood plasma; however, their association with possible molecular carriers remains poorly characterized. Here we report on the quantification of 46 endogenous molecular species from five major classes of endocannabinoids and endocannabinoid-related compounds in three lipoprotein fractions of human blood plasma: VLDL, LDL, HDL, and in the plasma lipoprotein-free fraction. Although sizable quantities of endocannabinoid-related molecules are associated with lipoproteins, we identified the lipoprotein-free fraction as a major carrier of endocannabinoids in blood circulation with the exception of 2-acylglycerols, which are markedly abundant in VLDL.

Gem1 and ERMES do not directly affect phosphatidylserine transport from ER to mitochondria or mitochondrial inheritance.

In yeast, a protein complex termed the ER-Mitochondria Encounter Structure (ERMES) tethers mitochondria to the endoplasmic reticulum. ERMES proteins are implicated in a variety of cellular functions including phospholipid synthesis, mitochondrial protein import, mitochondrial attachment to actin, polarized mitochondrial movement into daughter cells during division, and maintenance of mitochondrial DNA (mtDNA). The mitochondrial-anchored Gem1 GTPase has been proposed to regulate ERMES functions. Here, we show that ERMES and Gem1 have no direct role in the transport of phosphatidylserine (PS) from the ER to mitochondria during the synthesis of phosphatidylethanolamine (PE), as PS to PE conversion is not affected in ERMES or gem1 mutants. In addition, we report that mitochondrial inheritance defects in ERMES mutants are a secondary consequence of mitochondrial morphology defects, arguing against a primary role for ERMES in mitochondrial association with actin and mitochondrial movement. Finally, we show that ERMES complexes are long-lived, and do not depend on the presence of Gem1. Our findings suggest that the ERMES complex may have primarily a structural role in maintaining mitochondrial morphology.

Dietary intake of a MFGM/EV-rich concentrate promotes accretion of very long odd-chain sphingolipids and increases lipid metabolic turnover at the whole-body level.

Lipids from cow milk fat globule membranes (MFGMs) and extracellular vesicles (EVs) are considered beneficial for neurodevelopment, cognitive maintenance and human health in general. Nevertheless, it is largely unknown whether intake of infant formulas and medical nutrition products rich in these particles promote accretion of specific lipids and whether this affects metabolic homeostasis. To address this, we carried out a 16-week dietary intervention study where mice were supplemented with a MFGM/EV-rich concentrate, a control diet supplemented with a whey protein concentrate and devoid of milk lipids, or regular chow. Assessment of commonly used markers of metabolic health, including body weight, glucose intolerance and liver microanatomy, demonstrated no differences across the dietary regimes. In contrast, in-depth lipidomic analysis revealed accretion of milk-derived very long odd-chain sphingomyelins and ceramides in blood plasma and multiple tissues of mice fed the MFGM/EV diet. Furthermore, lipidomic flux analysis uncovered that mice fed the MFGM/EV diet have increased lipid metabolic turnover at the whole-body level. These findings help fill a long-lasting knowledge gap between the intake of MFGM/EV-containing foods and the health-promoting effects of their lipid constituents. In addition, the findings suggest that dietary sphingomyelins or ceramide-breakdown products with very long-chains can be used as structural components of cellular membranes, lipoprotein particles and signaling molecules that modulate metabolic homeostasis and health.

Cationic amphiphilic drugs induce accumulation of cytolytic lysoglycerophospholipids in the lysosomes of cancer cells and block their recycling into common membrane glycerophospholipids.

Lysosomes are acidic organelles responsible for lipid catabolism, and their functions can be disrupted by cationic amphiphilic drugs that neutralize lumenal pH and thereby inhibit most lysosomal hydrolases. These drugs can also induce lysosomal membrane permeabilization and cancer cell death, but the underlying mechanism remains elusive. Here, we uncover that the cationic amphiphilic drugs induce a substantial accumulation of cytolytic lysoglycerophospholipids within the lysosomes of cancer cells, and thereby prevent the recycling of lysoglycerophospholipids to produce common membrane glycerophospholipids. Using quantitative mass spectrometry-based shotgun lipidomics, we demonstrate that structurally diverse cationic amphiphilic drugs, along with other types of lysosomal pH-neutralizing reagents, elevate the amounts of lysoglycerophospholipids in MCF7 breast carcinoma cells. Lysoglycerophospholipids constitute ∼11 mol% of total glycerophospholipids in lysosomes purified from MCF7 cells, compared with ∼1 mol% in the cell lysates. Treatment with cationic amphiphilic drug siramesine further elevates the lysosomal lysoglycerophospholipid content to ∼24 mol% of total glycerophospholipids. Exogenously added traceable lysophosphatidylcholine is rapidly acylated to form diacylphosphatidylcholine, but siramesine treatment sequesters the lysophosphatidylcholine in the lysosomes and prevents it from undergoing acylation. These findings shed light on the unexplored role of lysosomes in the recycling of lysoglycerophospholipids and uncover the mechanism of action of promising anticancer agents.

Profiling the Mammalian Lipidome by Quantitative Shotgun Lipidomics.

The emerging field of lipidomics presents the systems biology approach to identify and quantify the full lipid repertoire of cells, tissues, and organisms. The importance of the lipidome is demonstrated by a number of biological studies on dysregulation of lipid metabolism in human diseases such as cancer, diabetes, and neurodegenerative diseases. Exploring changes and regulations in the huge networks of lipids and their metabolic pathways requires a lipidomics methodology: advanced mass spectrometry that resolves the complexity of the lipidome. Here, we report a comprehensive protocol of quantitative shotgun lipidomics that enables identification and quantification of hundreds of molecular lipid species, covering a wide range of lipid classes, extracted from cultured mammalian cells.

Hibernation and plasma lipids in free-ranging brown bears-implications for diabetes.

Brown bears (Ursus arctos) prepare for winter by overeating and increasing adipose stores, before hibernating for up to six months without eating, drinking, and with minimal movement. In spring, the bears exit the den without any damage to organs or physiology. Recent clinical research has shown that specific lipids and lipid profiles are of special interest for diseases such as diabetes type 1 and 2. Furthermore, rodent experiments show that lipids such as sulfatide protects rodents against diabetes. As free-ranging bears experience fat accumulation and month-long physical inactivity without developing diabetes, they could possibly be affected by similar protective measures. In this study, we investigated whether lipid profiles of brown bears are related to protection against hibernation-induced damage. We sampled plasma from 10 free-ranging Scandinavian brown bears during winter hibernation and repeated sampling during active state in the summer period. With quantitative shotgun lipidomics and liquid chromatography-mass spectrometry, we profiled 314 lipid species from 26 lipid classes. A principal component analysis revealed that active and hibernation samples could be distinguished from each other based on their lipid profiles. Six lipid classes were significantly altered when comparing plasma from active state and hibernation: Hexosylceramide, phosphatidylglycerol, and lysophosphatidylglycerol were higher during hibernation, while phosphatidylcholine ether, phosphatidylethanolamine ether, and phosphatidylinositol were lower. Additionally, sulfatide species with shorter chain lengths were lower, while longer chain length sulfatides were higher during hibernation. Lipids that are altered in bears are described by others as relevant for and associated with diabetes, which strengthens their position as potential effectors during hibernation. From this analysis, a range of lipids are suggested as potential protectors of bear physiology, and of potential importance in diabetes.

Identification of non-conventional small molecule degraders and stabilizers of squalene synthase.

Squalene synthase (SQS) is an essential enzyme in the mevalonate pathway, which controls cholesterol biosynthesis and homeostasis. Although catalytic inhibitors of SQS have been developed, none have been approved for therapeutic use so far. Herein we sought to develop SQS degraders using targeted protein degradation (TPD) to lower overall cellular cholesterol content. We found that KY02111, a small molecule ligand of SQS, selectively causes SQS to degrade in a proteasome-dependent manner. Unexpectedly, compounds based on the same scaffold linked to E3 ligase recruiting ligands led to SQS stabilization. Proteomic analysis found KY02111 to reduce only the levels of SQS, while lipidomic analysis determined that KY02111-induced degradation lowered cellular cholesteryl ester content. Stabilizers shielded SQS from its natural turnover without recruiting their matching E3 ligase or affecting enzymatic target activity. Our work shows that degradation of SQS is possible despite a challenging biological setting and provides the first chemical tools to degrade and stabilize SQS.

Reduced phosphatidylcholine level in the intestinal mucus layer of prediabetic NOD mice.

Type 1 diabetes (T1D) is an autoimmune disease with rising incidence. Pre- and manifest T1D is associated with intestinal barrier dysfunction, skewed microbiota composition, and serum dyslipidemia. The intestinal mucus layer protects against pathogens and its structure and phosphatidylcholine (PC) lipid composition may be compromised in T1D, potentially contributing to barrier dysfunction. This study compared prediabetic Non-Obese Diabetic (NOD) mice to healthy C57BL/6 mice by analyzing the intestinal mucus PC profile by shotgun lipidomics, plasma metabolomics by mass spectrometry and nuclear magnetic resonance, intestinal mucus production by histology, and cecal microbiota composition by 16 S rRNA sequencing. Jejunal mucus PC class levels were decreased in early prediabetic NOD vs C57BL/6 mice. In colonic mucus of NOD mice, the level of several PC species was reduced throughout prediabetes. In plasma, similar reductions of PC species were observed in early prediabetic NOD mice, where also increased beta-oxidation was prominent. No histological alterations were found in jejunal nor colonic mucus between the mouse strains. However, the ß-diversity of the cecal microbiota composition differed between prediabetic NOD and C57BL/6 mice, and the bacterial species driving this difference were related to decreased short-chain fatty acid (SCFA)-production in the NOD mice. This study reports reduced levels of PCs in the intestinal mucus layer and plasma of prediabetic NOD mice as well as reduced proportions of SCFA-producing bacteria in cecal content at early prediabetes, possibly contributing to intestinal barrier dysfunction and T1D.

Activation of invasion by oncogenic reprogramming of cholesterol metabolism via increased NPC1 expression and macropinocytosis.

Cancer cells are dependent on cholesterol, and they possess strictly controlled cholesterol homeostasis mechanisms. These allow them to smoothly switch between cholesterol synthesis and uptake to fulfill their needs and to adapt environmental changes. Here we describe a mechanism of how cancer cells employ oncogenic growth factor signaling to promote uptake and utilization of extracellular cholesterol via Myeloid Zinc Finger 1 (MZF1)-mediated Niemann Pick C1 (NPC1) expression and upregulated macropinocytosis. Expression of p95ErbB2, highly oncogenic, standard-treatment resistant form of ErbB2 mobilizes lysosomes and activates EGFR, invasion and macropinocytosis. This is connected to a metabolic shift from cholesterol synthesis to uptake due to macropinocytosis-enabled flow of extracellular cholesterol. NPC1 increase facilitates extracellular cholesterol uptake and is necessary for the invasion of ErbB2 expressing breast cancer spheroids and ovarian cancer organoids, indicating a regulatory role for NPC1 in the process. The ability to obtain cholesterol as a byproduct of increased macropinocytosis allows cancer cells to direct the resources needed for the energy-consuming cholesterol synthesis towards other activities such as invasion. These results demonstrate that macropinocytosis is not only an alternative energy source for cancer cells but also an efficient way to provide building material, such as cholesterol, for its macromolecules and membranes.

Quantitative profiling of PE, MMPE, DMPE, and PC lipid species by multiple precursor ion scanning: a tool for monitoring PE metabolism.

We report a method for the simultaneous identification and quantification of phosphatidylethanolamine (PE), monomethyl-phosphatidylethanolamine (MMPE), dimethyl-phosphatidylethanolamine (DMPE), and phosphatidylcholine (PC) species in lipid extracts. The method employs a specific "mass-tag" strategy where DMPE, MMPE, and PE species are chemically methylated with deuterated methyliodide (CD(3)I) to produce PC molecules having class-specific mass offsets of 3, 6 and 9Da, respectively. The derivatized aminoglycerophospholipids release characteristic phosphorylcholine-like fragment ions having specific mass offsets that powers sensitive and quantitative analysis by multiple precursor ion scanning on a hybrid quadrupole time-of-flight mass spectrometer. Using the mass-tag strategy, we could for the first time determine the stoichiometric relationship between the biosynthetic intermediates MMPE and DMPE, and abundant PE and PC species in a single mass spectrometric analysis. We demonstrated the efficacy of the methodology by conducting a series of biochemical experiments using stable isotope labeled ethanolamine to survey the activities and substrate specificities of enzymes involved in PE metabolism in Saccharomyces cerevisiae. Finally, we benchmarked the mass-tag strategy by specific and sensitive profiling of intermediate MMPE and DMPE species in liver.

Galactosyl- and glucosylsphingosine induce lysosomal membrane permeabilization and cell death in cancer cells.

Isomeric lysosphingolipids, galactosylsphingosine (GalSph) and glucosylsphingosine (GlcSph), are present in only minute levels in healthy cells. Due to defects in their lysosomal hydrolysis, they accumulate at high levels and cause cytotoxicity in patients with Krabbe and Gaucher diseases, respectively. Here, we show that GalSph and GlcSph induce lysosomal membrane permeabilization, a hallmark of lysosome-dependent cell death, in human breast cancer cells (MCF7) and primary fibroblasts. Supporting lysosomal leakage as a causative event in lysosphingolipid-induced cytotoxicity, treatment of MCF7 cells with lysosome-stabilizing cholesterol prevented GalSph- and GlcSph-induced cell death almost completely. In line with this, fibroblasts from a patient with Niemann-Pick type C disease, which is caused by defective lysosomal cholesterol efflux, were significantly less sensitive to lysosphingolipid-induced lysosomal leakage and cell death. Prompted by the data showing that MCF7 cells with acquired resistance to lysosome-destabilizing cationic amphiphilic drugs (CADs) were partially resistant to the cell death induced by GalSph and GlcSph, we compared these cell death pathways with each other. Like CADs, GalSph and GlcSph activated the cyclic AMP (cAMP) signalling pathway, and cAMP-inducing forskolin sensitized cells to cell death induced by low concentrations of lysosphingolipids. Contrary to CADs, lysosphingolipid-induced cell death was independent of lysosomal Ca2+ efflux through P2X purinerigic receptor 4. These data reveal GalSph and GlcSph as lysosome-destabilizing lipids, whose putative use in cancer therapy should be further investigated. Furthermore, the data supports the development of lysosome stabilizing drugs for the treatment of Krabbe and Gaucher diseases and possibly other sphingolipidoses.

PCSK9 deficiency alters brain lipid composition without affecting brain development and function.

PCSK9 induces lysosomal degradation of the low-density lipoprotein (LDL) receptor (LDLR) in the liver, hereby preventing removal of LDL cholesterol from the circulation. Accordingly, PCSK9 inhibitory antibodies and siRNA potently reduce LDL cholesterol to unprecedented low levels and are approved for treatment of hypercholesterolemia. In addition, PCSK9 inactivation alters the levels of several other circulating lipid classes and species. Brain function is critically influenced by cholesterol and lipid composition. However, it remains unclear how the brain is affected long-term by the reduction in circulating lipids as achieved with potent lipid lowering therapeutics such as PCSK9 inhibitors. Furthermore, it is unknown if locally expressed PCSK9 affects neuronal circuits through regulation of receptor levels. We have studied the effect of lifelong low peripheral cholesterol levels on brain lipid composition and behavior in adult PCSK9 KO mice. In addition, we studied the effect of PCSK9 on neurons in culture and in vivo in the developing cerebral cortex. We found that PCSK9 reduced LDLR and neurite complexity in cultured neurons, but neither PCSK9 KO nor overexpression affected cortical development in vivo. Interestingly, PCSK9 deficiency resulted in changes of several lipid classes in the adult cortex and cerebellum. Despite the observed changes, PCSK9 KO mice had unchanged behavior compared to WT controls. In conclusion, our findings demonstrate that altered PCSK9 levels do not compromise brain development or function in mice, and are in line with clinical trials showing that PCSK9 inhibitors have no adverse effects on cognitive function.

Acid ceramidase controls apoptosis and increases autophagy in human melanoma cells treated with doxorubicin.

Acid ceramidase (AC) is a lysosomal hydrolase encoded by the ASAH1 gene, which cleaves ceramides into sphingosine and fatty acid. AC is expressed at high levels in most human melanoma cell lines and may confer resistance against chemotherapeutic agents. One such agent, doxorubicin, was shown to increase ceramide levels in melanoma cells. Ceramides contribute to the regulation of autophagy and apoptosis. Here we investigated the impact of AC ablation via CRISPR-Cas9 gene editing on the response of A375 melanoma cells to doxorubicin. We found that doxorubicin activates the autophagic response in wild-type A375 cells, which effectively resist apoptotic cell death. In striking contrast, doxorubicin fails to stimulate autophagy in A375 AC-null cells, which rapidly undergo apoptosis when exposed to the drug. The present work highlights changes that affect melanoma cells during incubation with doxorubicin, in A375 melanoma cells lacking AC. We found that the remarkable reduction in recovery rate after doxorubicin treatment is strictly associated with the impairment of autophagy, that forces the AC-inhibited cells into apoptotic path.

Comprehensive Evaluation of a Quantitative Shotgun Lipidomics Platform for Mammalian Sample Analysis on a High-Resolution Mass Spectrometer.

Shotgun lipidomics is a powerful tool that enables simultaneous and fast quantification of diverse lipid classes through mass spectrometry based analyses of directly infused crude lipid extracts. We present here a shotgun lipidomics platform established to quantify 38 lipid classes belonging to four lipid categories present in mammalian samples and show the fine-tuning and comprehensive evaluation of its experimental parameters and performance. We first determined for all the targeted lipid classes the collision energy levels optimal for the recording of their lipid class- and species-specific fragment ions and fine-tuned the energy levels applied in the platform. We then performed a series of titrations to define the boundaries of linear signal response for the targeted lipid classes, and demonstrated that the dynamic quantification range spanned more than 3 orders of magnitude and reached sub picomole levels for 35 lipid classes. The platform identified 273, 261, and 287 lipid species in brain, plasma, and cultured fibroblast samples, respectively, at the respective optimal working sample amounts. The platform properly quantified the majority of these identified lipid species, while lipid species measured to be below the limit of quantification were efficiently removed from the data sets by the use of statistical analyses of data reproducibility or a cutoff threshold. Finally, we demonstrated that a series of parameters of cell culture conditions influence lipidomics outcomes, including confluency, medium supplements, and use of transfection reagents. The present study provides a guideline for setting up and using a simple and efficient platform for quantitatively exploring the mammalian lipidome.

L-serine supplementation lowers diabetes incidence and improves blood glucose homeostasis in NOD mice.

Sphingolipids are a diverse group of lipids with important roles in beta-cell biology regulating insulin folding and controlling apoptosis. Sphingolipid biosynthesis begins with the condensation of L-serine and palmitoyl-CoA. Here we tested the effect of L-serine supplementation on autoimmune diabetes development and blood glucose homeostasis in female NOD mice. We found that continuous supplementation of L-serine reduces diabetes incidence and insulitis score. In addition, L-serine treated mice had an improved glucose tolerance test, reduced HOMA-IR, and reduced blood glucose levels. L-serine led to a small reduction in body weight accompanied by reduced food and water intake. L-serine had no effect on pancreatic sphingolipids as measured by mass spectrometry. The data thus suggests that L-serine could be used as a therapeutic supplement in the treatment of Type 1 Diabetes and to improve blood glucose homeostasis.

Global Monitoring of the Mammalian Lipidome by Quantitative Shotgun Lipidomics.

The emerging field of lipidomics presents the systems biology approach to identify and quantify the full lipid repertoire of cells, tissues, and organisms. The importance of the lipidome is demonstrated by a number of biological studies on dysregulation of lipid metabolism in human diseases such as cancer, diabetes, and neurodegenerative diseases. Exploring changes and regulations in the huge networks of lipids and their metabolic pathways requires a lipidomics methodology: Advanced mass spectrometry that resolves the complexity of the lipidome. Here, we report a comprehensive protocol of quantitative shotgun lipidomics that enables identification and quantification of hundreds of molecular lipid species, covering a wide range of lipid classes, extracted from cultured mammalian cells.