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Adaptations to infectious and dietary pressures shape mammalian physiology and disease risk. How such adaptations affect sex-biased diseases remains insufficiently studied. In this study, we show that sex-dependent hepatic gene programs confer a robust (~300%) survival advantage for male mice during lethal bacterial infection. The transcription factor B cell lymphoma 6 (BCL6), which masculinizes hepatic gene expression at puberty, is essential for this advantage. However, protection by BCL6 protein comes at a cost during conditions of dietary excess, which result in overt fatty liver and glucose intolerance in males. Deleting hepatic BCL6 reverses these phenotypes but markedly lowers male survival during infection, thus establishing a sex-dependent trade-off between host defense and metabolic systems. Our findings offer strong evidence that some current sex-biased diseases are rooted in ancient evolutionary trade-offs between immunity and metabolism.

S1. Effect of ASA on cell viability. S2. Effect of Celecoxib on cell viability and migration. S3. Mice are tolerant of chronic dailly gavage. S4. ASA delays tumor growth of cell implants. S5. ASA does not delay tumor growth of some cell implants. S6. Effect of chronic ASA exposure on cytokines.

There are conflicting epidemiologic data on whether chronic aspirin (ASA) use may reduce melanoma risk in humans. Potential anticancer effects of ASA may be mediated by its ability to suppress prostaglandin E2 (PGE2) production and activate 5′-adenosine monophosphate–activated protein kinase (AMPK). We investigated the inhibitory effects of ASA in a panel of melanoma and transformed melanocyte cell lines, and on tumor growth in a preclinical model. ASA and the COX-2 inhibitor celecoxib did not affect melanoma cell viability, but significantly reduced colony formation, cell motility, and pigmentation (melanin production) in vitro at concentrations of 1 mmol/L and 20 μmol/L, respectively. ASA-mediated inhibition of cell migration and pigmentation was rescued by exogenous PGE2 or Compound C, which inhibits AMPK activation. Levels of tyrosinase, MITF, and p-ERK were unaffected by ASA exposure. Following a single oral dose of 0.4 mg ASA to NOD/SCID mice, salicylate was detected in plasma and skin at 4 hours and PGE2 levels were reduced up to 24 hours. Some human melanoma tumors xenografted into NOD/SCID mice were sensitive to chronic daily ASA administration, exhibiting reduced growth and proliferation. ASA-treated mice bearing sensitive and resistant tumors exhibited both decreased PGE2 in plasma and tumors and increased phosphorylated AMPK in tumors. We conclude that ASA inhibits colony formation, cell motility, and pigmentation through suppression of PGE2 and activation of AMPK and reduces growth of some melanoma tumors in vivo. This preclinical model could be used for further tumor and biomarker studies to support future melanoma chemoprevention trials in humans. Cancer Prev Res; 11(10); 629–42. ©2018 AACR.

SUMMARY Metabolic reprogramming in response to infection plays a critical role for septic survival. During a septic episode, the heart heavily relies on hepatic lipid particles to prevent heart damage and failure. Inositol- Requiring Enzyme 1 (IRE1) is the most conserved unfolded protein response (UPR) regulator that governs homeostasis of the endoplasmic reticulum (ER), the major site for lipid synthesis and processing. Here we show that hepatocyte IRE1 is indispensable for protecting against septic mortality in two different rodent models of experimental sepsis. The protective effect of hepatic IRE1 was not attributed to the inflammatory response since hepatic IRE1 deletion did not alter hepatic or systemic cytokine response. However, loss of IRE1 in the liver significantly augmented septic cardiac dysfunction in part due to a skewed immune-metabolic balance. Lipidomic and metabolomic analyses further revealed that loss of IRE1 in the liver compromised adaptive intrahepatic and circulating lipid reprogramming, including VLDL, in response to septic challenge. Furthermore, we identified that the protective effects against septic mortality are mediated by a non-canonical IRE1-dependent mechanism. Together, our study provides the first insight into how a disruption of hepatic ER-mediated lipid metabolic regulation promotes sepsis-associated cardiac immuno-metabolic imbalance.

Thermogenesis by uncoupling protein 1 (UCP1) is one of the primary mechanisms by which brown adipose tissue (BAT) increases energy expenditure. UCP1 resides in the inner mitochondrial membrane (IMM), where it dissipates membrane potential independent of adenosine triphosphate (ATP) synthase. Here, we provide evidence that phosphatidylethanolamine (PE) modulates UCP1-dependent proton conductance across the IMM to modulate thermogenesis. Mitochondrial lipidomic analyses revealed PE as a signature molecule whose abundance bidirectionally responds to changes in thermogenic burden. Reduction in mitochondrial PE by deletion of phosphatidylserine decarboxylase (PSD) made mice cold intolerant and insensitive to β3 adrenergic receptor agonist–induced increase in whole-body oxygen consumption. High-resolution respirometry and fluorometry of BAT mitochondria showed that loss of mitochondrial PE specifically lowers UCP1-dependent respiration without compromising electron transfer efficiency or ATP synthesis. These findings were confirmed by a reduction in UCP1 proton current in PE-deficient mitoplasts. Thus, PE performs a previously unknown role as a temperature-responsive rheostat that regulates UCP1-dependent thermogenesis.

Cells utilize multiple mechanisms to maintain mitochondrial homeostasis in response to stress. We recently identified a cellular structure, called the mitochondria-derived compartment (MDC), that is generated from mitochondria in response to amino acid overabundance. MDCs selectively sequester proteins from mitochondria for subsequent degradation, and loss of MDCs sensitizes cells to amino acid stress. Here, we conducted a microscopy-based screen in budding yeast to identify factors that regulate MDC formation. We found that levels of two mitochondrial phospholipids, cardiolipin (CL) and phosphatidylethanolamine (PE), regulate MDC biogenesis in opposing directions. CL depletion impairs MDC biogenesis, whereas PE reduction leads to constitutive MDC formation. Additionally, in response to MDC-inducing agents, cellular and mitochondrial PE declines in an amino acid-dependent manner. Overexpressing mitochondrial PE synthesis pathway components suppresses MDC biogenesis during amino acid stress. Altogether, our data indicate a requirement for CL in MDC biogenesis, and suggest that PE depletion may serve as a regulatory signal for MDC formation downstream of MDC-inducing metabolic stress.SUMMARYXiao et al. identify distinct roles for the phospholipids cardiolipin and phosphatidylethanolamine in mitochondria-derived compartment (MDC) biogenesis. They show that cardiolipin is required for MDC formation, whereas a decline in phosphatidylethanolamine in response to amino acid stress triggers MDC biogenesis.

Weight loss is associated with a disproportionate decrease in whole-body energy expenditure that may contribute to the heightened risk for weight-regain. Evidence suggests that this energetic mismatch originates from lean tissue. Although this phenomenon is well documented, the mechanisms have remained elusive. We hypothesized that increased mitochondrial energy efficiency in skeletal muscle is associated with reduced expenditure under weight loss. Wildtype male C57BL6/N mice were fed with high-fat diet for 10 wks, followed by a subset of mice that were maintained on the obesogenic diet (OB) or switched to standard chow to promote weight loss (WL) for additional 6 wks. Mitochondrial energy efficiency was evaluated using high-resolution respirometry and fluorometry. Mass spectrometric analyses were employed to describe the mitochondrial proteome and lipidome. Weight loss promoted ~50% increase in the efficiency of oxidative phosphorylation (ATP produced per O2consumed, or P/O) in skeletal muscle. However, weight loss did not appear to induce significant changes in mitochondrial proteome, nor any changes in respiratory supercomplex formation. Instead, it accelerated the remodeling of mitochondrial cardiolipin (CL) acyl-chains to increase tetralinoleoyl CL (TLCL) content, a species of lipids thought to be functionally critical for the respiratory enzymes. We further show that lowering TLCL by deleting the CL transacylase tafazzin was sufficient to reduce skeletal muscle P/O and protect mice from diet-induced weight gain. These findings implicate skeletal muscle mitochondrial efficiency as a novel mechanism by which weight loss reduces energy expenditure in obesity.

Disruption of sphingolipid homeostasis and signaling has been implicated in diabetes, cancer, cardiometabolic, and neurodegenerative disorders. Yet, mechanisms governing cellular sensing and regulation of sphingolipid homeostasis remain largely unknown. In yeast, serine palmitoyltransferase, catalyzing the first and rate-limiting step of sphingolipid de novo biosynthesis, is negatively regulated by Orm1 and 2. Lowering sphingolipids triggers Orms phosphorylation, upregulation of serine palmitoyltransferase activity and sphingolipid de novo biosynthesis. However, mammalian orthologs ORMDLs lack the N-terminus hosting the phosphosites. Thus, which sphingolipid(s) are sensed by the cells, and mechanisms of homeostasis remain largely unknown. Here, we identify sphingosine-1-phosphate (S1P) as key sphingolipid sensed by cells via S1PRs to maintain homeostasis. The increase in S1P-S1PR signaling stabilizes ORMDLs, restraining SPT activity. Mechanistically, the hydroxylation of ORMDLs at Pro137 allows a constitutive degradation of ORMDLs via ubiquitin-proteasome pathway, preserving SPT activity. Disrupting S1PR/ORMDL axis results in ceramide accrual, mitochondrial dysfunction, impaired signal transduction, all underlying endothelial dysfunction, early event in the onset of cardio- and cerebrovascular diseases. Our discovery may provide the molecular basis for therapeutic intervention restoring sphingolipid homeostasis.

Carbohydrates and lipids provide the majority of substrates to fuel mitochondrial oxidative phosphorylation (OXPHOS). Metabolic inflexibility, defined as an impaired ability to switch between these fuels, is implicated in a number of metabolic diseases. Here we explore the mechanism by which physical inactivity promotes metabolic inflexibility in skeletal muscle. We developed a mouse model of sedentariness by small mouse cage (SMC) that, unlike other classic models of disuse in mice, faithfully recapitulates metabolic responses that occur in humans. Bioenergetic phenotyping of mitochondria displayed metabolic inflexibility induced by physical inactivity, demonstrated by a reduction in pyruvate-stimulated respiration (JO2) in absence of a change in palmitate-stimulatedJO2. Pyruvate resistance in these mitochondria was likely driven by a decrease in phosphatidylethanolamine (PE) abundance in the mitochondrial membrane. Reduction in mitochondrial PE by deletion of phosphatidylserine decarboxylase (PSD) was sufficient to induce metabolic inflexibility measured at the whole-body level, as well as at the level of skeletal muscle mitochondria. Low mitochondrial PE was sufficient to increase glucose flux towards lactate. We further implicate that resistance to pyruvate metabolism is due to attenuated mitochondrial entry via mitochondrial pyruvate carrier (MPC). These findings suggest a novel mechanism by which mitochondrial PE directly regulates MPC activity to modulate metabolic flexibility.

SUMMARYThermogenesis by uncoupling protein 1 (UCP1) is one of the primary mechanisms by which brown adipose tissue (BAT) increases energy expenditure. UCP1 resides in the inner mitochondrial membrane (IMM), where it dissipates membrane potential independent of ATP synthase. Here we provide evidence that mitochondrial phosphatidylethanolamine (PE) directly regulates UCP1-dependent proton conductance across IMM to modulate thermogenesis. Mitochondrial lipidomic analyses revealed PE as a signature molecule whose abundance bidirectionally responds to changes in thermogenic burden. Reduction in mitochondrial PE by deletion of phosphatidylserine decarboxylase (PSD) made mice cold intolerant and insensitive to β3 adrenergic receptor agonist-induced increase in whole-body oxygen consumption. High-resolution respirometry and fluorometry of BAT mitochondria showed that loss of mitochondrial PE specifically lowers UCP1-dependent respiration without compromising electron transfer efficiency or ATP synthesis. These findings were confirmed by a reduction in UCP1 proton current in PSD-deficient mitoplasts. Thus, PE performs a previously unknown role as a temperature-responsive rheostat that regulates UCP1-dependent thermogenesis.

Syndromic ciliopathies and retinal degenerations are large heterogeneous groups of genetic diseases. CFAP418 is a causative gene of both disorders, and its protein sequence is evolutionarily conserved. However, the pathogenic mechanism caused by CFAP418 mutations is largely unknown. Here, we employed affinity purification coupled with mass spectrometry and quantitative lipidomic, proteomic, and phosphoproteomic approaches to address the molecular function of CFAP418 in mouse retinas. We showed that CFAP418 bound to lipid metabolism precursor phosphatidic acid (PA) and mitochondrion-specific lipid cardiolipin but did not form a tight and static complex with proteins. Loss of Cfap418 led to membrane lipid imbalance and protein-membrane association alteration, which subsequently caused mitochondrial defects and membrane remodeling abnormalities in multiple vesicular trafficking pathways. Loss of Cfap418 also increased the activity of PA-binding protein kinase Cα. Our results indicate that membrane lipid imbalance is a new pathological mechanism underlying syndromic ciliopathies and retinal degenerations, which is associated with other known causative genes for these diseases, such as RAB28 and BBS genes.

Background and aims Iron is essential yet also highly chemically reactive and potentially toxic. The mechanisms that allow cells to use iron safely are not clear; defects in iron management are a causative factor in the cell-death pathway known as ferroptosis. Poly rC binding protein 1 (PCBP1) is a multifunctional protein that serves as a cytosolic iron chaperone, binding and transferring iron to recipient proteins in mammalian cells. Although PCBP1 distributes iron in cells, its role in managing iron in mammalian tissues remains open for study. The liver is highly specialized for iron uptake, utilization, storage, and secretion. Approach and results Mice lacking PCBP1 in hepatocytes exhibited defects in liver iron homeostasis with low levels of liver iron, reduced activity of iron enzymes, and misregulation of the cell-autonomous iron regulatory system. These mice spontaneously developed liver disease with hepatic steatosis, inflammation, and degeneration. Transcriptome analysis indicated activation of lipid biosynthetic and oxidative-stress response pathways, including the antiferroptotic mediator, glutathione peroxidase type 4. Although PCBP1-deleted livers were iron deficient, dietary iron supplementation did not prevent steatosis; instead, dietary iron restriction and antioxidant therapy with vitamin E prevented liver disease. PCBP1-deleted hepatocytes exhibited increased labile iron and production of reactive oxygen species (ROS), were hypersensitive to iron and pro-oxidants, and accumulated oxidatively damaged lipids because of the reactivity of unchaperoned iron. Conclusions Unchaperoned iron in PCBP1-deleted mouse hepatocytes leads to production of ROS, resulting in lipid peroxidation (LPO) and steatosis in the absence of iron overload. The iron chaperone activity of PCBP1 is therefore critical for limiting the toxicity of cytosolic iron and may be a key factor in preventing the LPO that triggers the ferroptotic cell-death pathway.

BACKGROUND: Ceramides are sphingolipids that play causative roles in diabetes and heart disease, with their serum levels measured clinically as biomarkers of cardiovascular disease (CVD). METHODS: We performed targeted lipidomics on serum samples from individuals with familial coronary artery disease (CAD) (n = 462) and population-based controls (n = 212) to explore the relationship between serum sphingolipids and CAD, using unbiased machine learning to identify sphingolipid species positively associated with CAD. RESULTS: Nearly every sphingolipid measured (n = 30 of 32) was significantly elevated in subjects with CAD compared with measurements in population controls. We generated a novel sphingolipid-inclusive CAD risk score, termed SIC, that demarcates patients with CAD independently and more effectively than conventional clinical CVD biomarkers including serum LDL cholesterol and triglycerides. This new metric comprises several minor lipids that likely serve as measures of flux through the ceramide biosynthesis pathway rather than the abundant deleterious ceramide species that are included in other ceramide-based scores. CONCLUSION: This study validates serum ceramides as candidate biomarkers of CVD and suggests that comprehensive sphingolipid panels should be considered as measures of CVD. FUNDING: The NIH (DK112826, DK108833, DK115824, DK116888, and DK116450); the Juvenile Diabetes Research Foundation (JDRF 3-SRA-2019-768-A-B); the American Diabetes Association; the American Heart Association; the Margolis Foundation; the National Cancer Institute, NIH (5R00CA218694-03); and the Huntsman Cancer Institute Cancer Center Support Grant (P30CA040214).

Current concepts in evolutionary medicine propose that trade-offs and mismatches with a shifting environment increase disease risk. While biological sex also impacts disease prevalence, contributions of environmental pressures to sex-biased diseases remain unexplored. Here, we show that sex-dependent hepatic programs confer a robust (~300%) survival advantage for male mice during lethal bacterial infection. The transcription factor BCL6, which masculinizes hepatic gene expression at puberty, is essential for this advantage. However, protection by BCL6 comes at a cost following dietary excess, resulting in overt fatty liver and glucose intolerance in males. Deleting hepatic BCL6 reverses these phenotypes but markedly lowers male fitness during infection, thus establishing a sex-dependent tradeoff between host defense and metabolic systems. We suggest that these tradeoffs, coupled with current environmental pressures, drive metabolic disease in males.

Ift88 gene mutations cause primary cilia loss and polycystic kidney disease (PKD) in mice. Nephron intraflagellar transport protein 88 (Ift88) knockout (KO) at 2 mo postnatal does not affect renal histology at 4 mo postnatal and causes PKD only in males by 11 mo postnatal. To identify factors associated with PKD development, kidneys from 4-mo-old male and female control and Ift88 KO mice underwent transcriptomic, proteomic, Western blot, metabolomic, and lipidomic analyses. mRNAs involved in extracellular matrix (ECM) synthesis and degradation were selectively upregulated in male KO mice. Proteomic analysis was insufficiently sensitive to detect most ECM components, while Western blot analysis paradoxically revealed reduced fibronectin and collagen type I in male KO mice. Only male KO mice had upregulated mRNAs encoding fibrinogen subunits and receptors for vascular endothelial growth factor and platelet-derived growth factor; period 2, period 3, and nuclear receptor subfamily 1 group D member 1 clock mRNAs were selectively decreased in male KO mice. Proteomic, metabolomic, and lipidomic analyses detected a relative (vs. the same-sex control) decrease in factors involved in fatty acid β-oxidation in female KO mice, while increased or unchanged levels in male KO mice, including medium-chain acyl-CoA dehydrogenase, 3-hydroxybutyrate, and acylcarnitine. Three putative mRNA biomarkers of cystogenesis in male Ift88 KO mice (similar control levels between sexes and uniquely altered by KO in males) were identified, including high levels (fibrinogen α-chain and stromal cell-derived factor 2-like 1) and low levels (BTG3-associated nuclear protein) in male KO mice. These findings suggest that relative alterations in renal ECM metabolism, fatty acid β-oxidation, and other pathways precede cystogenesis in Ift88 KO mice. In addition, potential novel biomarkers of cystogenesis in Ift88 KO mice have been identified. NEW & NOTEWORTHY Male, but not female, mice with nephron intraflagellar transport protein 88 (Ift88) gene knockout (KO) develop polycystic kidneys by ∼1 yr postnatal. We performed multiomic analysis of precystic male and female Ift88 KO and control kidneys. Precystic male Ift88 KO mice exhibited differential alterations (vs. females) in mRNA, proteins, metabolites, and/or lipids associated with renal extracellular matrix metabolism, fatty acid β-oxidation, circadian rhythm, and other pathways. These findings suggest targets for evaluation in the pathogenesis of Ift88 KO polycystic kidneys.

Sphingolipids (SL) are both membrane building blocks and potent signaling molecules regulating a variety of cellular functions in both physiological and pathological conditions. Under normal physiology, sphingolipid levels are tightly regulated, whereas disruption of sphingolipid homeostasis and signaling has been implicated in diabetes, cancer, cardiovascular and autoimmune diseases. Yet, mechanisms governing cellular sensing of SL, and according regulation of their biosynthesis remain largely unknown.In yeast, serine palmitoyltransferase (SPT), catalyzing the first and rate limiting step of sphingolipid de novo biosynthesis, is negatively regulated by Orosomucoid 1 and 2 (Orm) proteins. Lowering sphingolipid levels triggers Orms phosphorylation, resulting in the removal of the inhibitory brake on SPT to enhance sphingolipid de novo biosynthesis. However, mammalian orthologs ORMDLs lack the N-terminus hosting the phosphosites. Thus, which sphingolipid(s) are sensed by the cells, and mechanisms of homeostasis remain largely unknown. This study is aimed at filling this knowledge gap.Here, we identify sphingosine-1-phosphate (S1P) as the key sphingolipid sensed by endothelial cells via S1PRs. The increase of S1P-S1PR signaling stabilizes ORMDLs, which downregulates SPT activity to maintain SL homeostasis. These findings reveal the S1PR/ORMDLs axis as the sensor-effector unit regulating SPT activity accordingly. Mechanistically, the hydroxylation of ORMDLs at Pro137 allows a constitutive degradation of ORMDLs via ubiquitin-proteasome pathway, therefore preserving SPT activity at steady state. The disruption of the S1PR/ORMDL axis results in ceramide accrual, mitochondrial dysfunction, and impaired signal transduction, all leading to endothelial dysfunction, which is an early event in the onset of cardio- and cerebrovascular diseases.The disruption of S1P-ORMDL-SPT signaling may be implicated in the pathogenesis of conditions such as diabetes, cancer, cardiometabolic disorders, and neurodegeneration, all characterized by deranged sphingolipid metabolism. Our discovery may provide the molecular basis for a therapeutic intervention to restore sphingolipid homeostasis.

Obesity alters skeletal muscle lipidome and promotes myopathy, but it is unknown whether aberrant muscle lipidome contributes to the reduction in skeletal muscle contractile force-generating capacity. Comprehensive lipidomic analyses of mouse skeletal muscle revealed a very strong positive correlation between the abundance of lysophosphatidylcholine (lyso-PC), a class of lipids that is known to be downregulated with obesity, with maximal tetanic force production. The level of lyso-PC is regulated primarily by lyso-PC acyltransferase 3 (LPCAT3), which acylates lyso-PC to form phosphatidylcholine. Tamoxifen-inducible skeletal muscle-specific overexpression of LPCAT3 (LPCAT3-MKI) was sufficient to reduce muscle lyso-PC content in both standard chow diet- and high-fat diet (HFD)-fed conditions. Strikingly, the assessment of skeletal muscle force-generating capacity ex vivo revealed that muscles from LPCAT3-MKI mice were weaker regardless of diet. Defects in force production were more apparent in HFD-fed condition, where tetanic force production was 40% lower in muscles from LPCAT3-MKI compared to that of control mice. These observations were partly explained by reductions in the cross-sectional area in type IIa and IIx fibers, and signs of muscle edema in the absence of fibrosis. Future studies will pursue the mechanism by which LPCAT3 may alter protein turnover to promote myopathy.

Zellweger spectrum disorder (ZSD) is the most severe peroxisomal biogenesis disorder (PBD). Why ZSD patients not only loose functional peroxisomes but also present with severe mitochondrial dysfunction was a long‐standing mystery. In this issue, Nuebel et al (2021) identified that loss of peroxisomes leads to re‐routing of peroxisomal proteins to mitochondria, thereby impairing mitochondrial structure and function. The findings provide the first molecular understanding of the mitochondrial‐peroxisomal link in ZSD., A study in this issue provides the first molecular understanding of the mitochondrial‐peroxisomal link in peroxisomal disorders.

The biophysical environment of membrane phospholipids affects structure, function, and stability of membrane-bound proteins.1,2 Obesity can disrupt membrane lipids, and in particular, alter the activity of sarco/endoplasmic reticulum (ER/SR) Ca2+-ATPase (SERCA) to affect cellular metabolism.3-5 Recent evidence suggests that transport efficiency (Ca2+ uptake / ATP hydrolysis) of skeletal muscle SERCA can be uncoupled to increase energy expenditure and protect mice from diet-induced obesity.6,7 In isolated SR vesicles, membrane phospholipid composition is known to modulate SERCA efficiency.8-11 Here we show that skeletal muscle SR phospholipids can be altered to decrease SERCA efficiency and increase whole-body metabolic rate. The absence of skeletal muscle phosphatidylethanolamine (PE) methyltransferase (PEMT) promotes an increase in skeletal muscle and whole-body metabolic rate to protect mice from diet-induced obesity. The elevation in metabolic rate is caused by a decrease in SERCA Ca2+-transport efficiency, whereas mitochondrial uncoupling is unaffected. Our findings support the hypothesis that skeletal muscle energy efficiency can be reduced to promote protection from obesity.

Supplemental data for "Characterizing a common CERS2 polymorphism in a mouse model of metabolic disease and in subjects from the Utah CAD Study."

ContextGenome-wide association studies have identified associations between a common single nucleotide polymorphism (SNP; rs267738) in CERS2, a gene that encodes a (dihydro)ceramide synthase that is involved in the biosynthesis of very-long-chain sphingolipids (eg, C20-C26) and indices of metabolic dysfunction (eg, impaired glucose homeostasis). However, the biological consequences of this mutation on enzyme activity and its causal roles in metabolic disease are unresolved.ObjectiveThe studies described herein aimed to characterize the effects of rs267738 on CERS2 enzyme activity, sphingolipid profiles, and metabolic outcomes.DesignWe performed in-depth lipidomic and metabolic characterization of a novel CRISPR knock-in mouse modeling the rs267738 variant. In parallel, we conducted mass spectrometry-based, targeted lipidomics on 567 serum samples collected through the Utah Coronary Artery Disease study, which included 185 patients harboring 1 (n = 163) or both (n = 22) rs267738 alleles.ResultsIn-silico analysis of the amino acid substitution within CERS2 caused by the rs267738 mutation suggested that rs267738 is deleterious for enzyme function. Homozygous knock-in mice had reduced liver CERS2 activity and enhanced diet-induced glucose intolerance and hepatic steatosis. However, human serum sphingolipids and a ceramide-based cardiac event risk test 1 score of cardiovascular disease were not significantly affected by rs267738 allele count.ConclusionsThe rs267738 SNP leads to a partial loss-of-function of CERS2, which worsened metabolic parameters in knock-in mice. However, rs267738 was insufficient to effect changes in serum sphingolipid profiles in subjects from the Utah Coronary Artery Disease Study.

Made available in DSpace on 2019-10-06T17:15:07Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-07-26 American Diabetes Association American Heart Association Ben B. and Iris M. Margolis Foundation Diabetes Research Center, University of Washington Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Preeclampsia Research Laboratories U.S. Department of Agriculture Juvenile Diabetes Research Foundation United Kingdom Agricultural Research Service National Institutes of Health Norges Idrettshøgskole Ceramides contribute to the lipotoxicity that underlies diabetes, hepatic steatosis, and heart disease. By genetically engineering mice, we deleted the enzyme dihydroceramide desaturase 1 (DES1), which normally inserts a conserved double bond into the backbone of ceramides and other predominant sphingolipids. Ablation of DES1 from whole animals or tissue-specific deletion in the liver and/or adipose tissue resolved hepatic steatosis and insulin resistance in mice caused by leptin deficiency or obesogenic diets. Mechanistic studies revealed ceramide actions that promoted lipid uptake and storage and impaired glucose utilization, none of which could be recapitulated by (dihydro)ceramides that lacked the critical double bond. These studies suggest that inhibition of DES1 may provide a means of treating hepatic steatosis and metabolic disorders. Department of Nutrition and Integrative Physiology Diabetes and Metabolism Research Center University of Utah Merck Research Laboratories Merck School of Dentistry São Paulo State University (UNESP) Department of Biochemistry and the Diabetes and Metabolism Research Center University of Utah Baker IDI Heart and Diabetes Institute Faculty of Science University of Brunei Darussalam Sciex Howard Hughes Medical Institute Bristol Myers Squibb Morphic Therapeutic Johnson and Johnson School of Dentistry São Paulo State University (UNESP) FAPESP: 2014/17619-6 U.S. Department of Agriculture: 2019-67018-29250 Juvenile Diabetes Research Foundation United Kingdom: 3-SRA-2019-768-A-B Agricultural Research Service: 5T32DK091317 National Institutes of Health: DK108833 National Institutes of Health: DK112826 National Institutes of Health: DK115824 National Institutes of Health: DK116450 Norges Idrettshøgskole: P30DK020579

Peroxisomal Biogenesis Disorders (PBDs) are a class of inherited metabolic disorders with profound neurological and other phenotypes. The most severe PBDs are caused by mutations in peroxin genes, which result in nonfunctional peroxisomes typically through impaired protein import. In order to better understand the molecular causes of Zellweger Spectrum Disease (ZSD) -the most severe PBDs -, we investigated the fate of peroxisomal mRNAs and proteins in ZSD model systems. We found that loss of peroxisomal import has no effect on peroxin mRNA expression or translational efficiency. Instead, peroxin proteins—still produced at high levels— aberrantly accumulate on the mitochondrial membrane, impairing respiration and ATP generation. Finally, we rescued mitochondrial function in fibroblasts derived from human patients with ZSD by overexpressing ATAD1, an AAA-ATPase that functions in mitochondrial quality control. These findings might provide a new focus of PBD therapies in supporting quality control pathways that protect mitochondrial function.

Macrophages have a defined role in the pathogenesis of metabolic disease and cholesterol metabolism where alternative activation of macrophages is thought to be beneficial to both glucose and cholesterol metabolism during high fat diet induced disease. It is well established that helminth infection protects from metabolic disease, but the mechanisms underlying protection are not well understood. Here, we investigated the effects of Schistosoma mansoni infection and cytokine activation in the metabolic signatures of bone marrow derived macrophages using an approach that integrated transcriptomics, metabolomics, and lipidomics in a metabolic disease prone mouse model. We demonstrate that bone marrow derived macrophages (BMDM) from S. mansoni infected male ApoE-/- mice have dramatically increased mitochondrial respiration compared to those from uninfected mice. This change is associated with increased glucose and palmitate shuttling into TCA cycle intermediates, increased accumulation of free fatty acids, and decreased accumulation of cellular cholesterol esters, tri and diglycerides, and is dependent on mgll activity. Systemic injection of IL-4 complexes is unable to recapitulate either reductions in systemic glucose AUC or the re-programing of BMDM mitochondrial respiration seen in infected males. Importantly, the metabolic reprogramming of male myeloid cells is transferrable via bone marrow transplantation to an uninfected host, indicating maintenance of reprogramming in the absence of sustained antigen exposure. Finally, schistosome induced metabolic and bone marrow modulation is sex-dependent, with infection protecting male, but not female mice from glucose intolerance and obesity. Our findings identify a transferable, long-lasting sex-dependent reprograming of the metabolic signature of macrophages by helminth infection, providing key mechanistic insight into the factors regulating the beneficial roles of helminth infection in metabolic disease., Author summary Globally, helminth endemic regions have lower incidences of metabolic disease. Schistosomiasis in particular has been shown to protect male mice from the development of atherosclerosis, diabetes and obesity while altering the metabolism of liver macrophages. In this study we sought to understand if metabolic modulation occurs systemically. We have found for the first time that macrophages generated from bone marrow myeloid progenitors from infected male mice have long-lived increases in their basal metabolism of lipids. We have also found the type-2 polarizing cytokine IL-4 is not sufficient to replicate infection induced metabolic reprogramming. Importantly, we demonstrate that the changes to male macrophage metabolism can be transferred to an uninfected host. This suggests that metabolic reprogramming is long-lived without exposure to active infection, and develops a memory-like phenotype in myeloid cells.

Cells harbor two systems for fatty acid synthesis, one in the cytoplasm (catalyzed by fatty acid synthase, FASN) and one in the mitochondria (mtFAS). In contrast to FASN, mtFAS is poorly characterized, especially in higher eukaryotes, with the major product(s), metabolic roles, and cellular function(s) being essentially unknown. Here we show that hypomorphic mtFAS mutant mouse skeletal myoblast cell lines display a severe loss of electron transport chain (ETC) complexes and exhibit compensatory metabolic activities including reductive carboxylation. This effect on ETC complexes appears to be independent of protein lipoylation, the best characterized function of mtFAS, as mutants lacking lipoylation have an intact ETC. Finally, mtFAS impairment blocks the differentiation of skeletal myoblasts in vitro. Together, these data suggest that ETC activity in mammals is profoundly controlled by mtFAS function, thereby connecting anabolic fatty acid synthesis with the oxidation of carbon fuels., eLife digest In human, plant and other eukaryotic cells, fats are an important source of energy and also play many other roles including waterproofing, thermal insulation and energy storage. Eukaryotic cells have two systems that make the building blocks of fats (known as fatty acids) and one of these systems, called the mtFAS pathway, operates in small compartments known as mitochondria. This pathway only has one known product, a small fat molecule called lipoic acid, which mitochondria attach to several enzymes to allow them to work properly. The main role of mitochondria is to break down fats and other molecules to release chemical energy that powers many processes in cells. They achieve this using large groups of proteins known as ETC complexes. To build these complexes, families of proteins known as ETC assembly factors carefully coordinate the assembly of many proteins and small molecules into specific structures. However, it remains unclear precisely how this process works. Here, Nowinski et al. used a gene editing technique to mutate the genes encoding three enzymes in the mtFAS pathway in mammalian cells. The experiments found that the mutant cells had fewer ETC complexes and seemed to be less able to break down fats and other molecules than ‘normal’ cells. Furthermore, a family of ETC assembly factors were less stable in the mutant cells. These findings suggest that the mtFAS pathway controls how mitochondria assemble ETC complexes. Further experiments indicated that lipoic acid is not involved in the assembly of ETC complexes and that the mtFAS pathway produces another, as yet unidentified, product that regulates this process, instead. MEPAN syndrome is a rare neurological disorder that leads to progressive loss of control of movement, slurred speech and impaired vision in children. Patients with this syndrome have genetic mutations affecting components of the mtFAS pathway, therefore, a better understanding of how the pathway works may help researchers develop new treatments in the future. More broadly, these findings will have important ramifications for many other situations in which the activity of ETC complexes in mitochondria is modified.

The biophysical environment of membrane phospholipids affects structure, function, and stability of membrane-bound proteins.1,2 Obesity can disrupt membrane lipids, and in particular, alter the activity of sarco/endoplasmic reticulum (ER/SR) Ca2+-ATPase (SERCA) to affect cellular metabolism.3–5 Recent evidence suggests that transport efficiency (Ca2+ uptake / ATP hydrolysis) of skeletal muscle SERCA can be uncoupled to increase energy expenditure and protect mice from diet-induced obesity.6,7 In isolated SR vesicles, membrane phospholipid composition is known to modulate SERCA efficiency.8–11 Here we show that skeletal muscle SR phospholipids can be altered to decrease SERCA efficiency and increase whole-body metabolic rate. The absence of skeletal muscle phosphatidylethanolamine (PE) methyltransferase (PEMT) promotes an increase in skeletal muscle and whole-body metabolic rate to protect mice from diet-induced obesity. The elevation in metabolic rate is caused by a decrease in SERCA Ca2+-transport efficiency, whereas mitochondrial uncoupling is unaffected. Our findings support the hypothesis that skeletal muscle energy efficiency can be reduced to promote protection from obesity.

Cells harbor two systems for fatty acid synthesis, one in the cytoplasm (catalyzed by fatty acid synthase, FASN) and one in the mitochondria (mtFAS). In contrast to FASN, mtFAS is poorly characterized, especially in higher eukaryotes, with the major product(s), metabolic roles, and cellular function(s) being essentially unknown. Here we show that hypomorphic mtFAS mutants display a severe loss of electron transport chain (ETC) complexes and exhibit compensatory metabolic activities including reductive carboxylation. This effect on ETC complexes appears to be independent of protein lipoylation, the best characterized function of mtFAS, as mutants lacking lipoylation have an intact ETC. Finally, mtFAS impairment blocks the differentiation of skeletal myoblasts in vitro. Together, these data suggest that ETC activity in mammals is profoundly controlled by mtFAS function, thereby connecting anabolic fatty acid synthesis with the oxidation of carbon fuels.

The use of retention time is often critical for the identification of compounds in metabolomic and lipidomic studies. Standards are frequently unavailable for the retention time measurement of many metabolites, thus the ability to predict retention time for these compounds is highly valuable. A number of studies have applied machine learning to predict retention times, but applying a published machine learning model to different lab conditions is difficult. This is due to variation between chromatographic equipment, methods, and columns used for analysis. Recreating a machine learning model is likewise difficult without a dedicated bioinformatician. Herein we present QSRR Automator, a software package to automate retention time prediction model creation and demonstrate its utility by testing data from multiple chromatography columns from previous publications and in-house work. Analysis of these data sets shows similar accuracy to published models, demonstrating the software&rsquo, s utility in metabolomic and lipidomic studies.

Despite strong evidence that helminth infections are protective against the development of metabolic disease, a major gap exists in understanding the mechanism(s) underlying this. We have previously found that Schistosoma mansoni induces profound alterations to the metabolic transcriptome of hepatic macrophages and protects male ApoE -/- on high fat diet from the development of obesity, glucose intolerance, and atherosclerosis. Here we demonstrate that macrophages derived from the bone marrow (BMDM) of S. mansoni infected male ApoE -/- mice have dramatically increased mitochondrial respiration and mitochondrial mass compared to those from uninfected mice. This change is accompanied by increased glucose and palmitate shuttling into TCA cycle intermediates and decreased accumulation of cellular cholesterol esters. The systemic effects of metabolic modulation by schistosome infection are a function of biological sex, where schistosome infection protects ApoE -/- male mice from obesity, glucose intolerance, and increased serum triglycerides, but not female mice. The sex-dependent effects of infection extend to myeloid cells specifically, where metabolic reprogramming leads to opposite cholesterol phenotypes in BMDM from infected females and males. Finally, we demonstrate that the metabolic reprogramming of male myeloid cells is transferrable via bone marrow transplantation to an uninfected host, indicating maintenance of reprogramming in the absence of ongoing schistosome antigen exposure. This work provides strong evidence that S. mansoni systemically reprograms the metabolism of the myeloid compartment in a sex-dependent manner.

From the CH(2)Cl(2) extract of the Antarctic sponge Dendrilla antarctica we found spongian diterpenes, including previously reported aplysulphurin (1), tetrahydroaplysulphurin-1 (2), membranolide (3), and darwinolide (4), utilizing a CH(2)Cl(2)/MeOH extraction scheme. However, the extracts also yielded diterpenes bearing one or more methyl acetal functionalities (5–9), two of which are previously unreported, while others are revised here. Further investigation of diterpene reactivity led to additional new metabolites (10–12), which identified them as well as the methyl acetals as artifacts from methanolysis of aplysulphurin. The bioactivity of the methanolysis products, membranoids A–H (5–12), as well as natural products 1–4, were assessed for activity against Leishmania donovani-infected J774A.1 macrophages, revealing insights into their structure/activity relationships. Four diterpenes, tetrahydroaplysulphurin-1 (2) as well as membranoids B (6), D (8), and G (11), displayed low micromolar activity against L. donovani with no discernible cytotoxicity against uninfected J774A.1 cells. Leishmaniasis is a neglected tropical disease that affects one million people every year and can be fatal if left untreated.

There are conflicting epidemiologic data on whether chronic aspirin (ASA) use may reduce melanoma risk in humans. Potential anticancer effects of ASA may be mediated by its ability to suppress prostaglandin E2 (PGE2) production and activate 5′-adenosine monophosphate–activated protein kinase (AMPK). We investigated the inhibitory effects of ASA in a panel of melanoma and transformed melanocyte cell lines, and on tumor growth in a preclinical model. ASA and the COX-2 inhibitor celecoxib did not affect melanoma cell viability, but significantly reduced colony formation, cell motility, and pigmentation (melanin production) in vitro at concentrations of 1 mmol/L and 20 μmol/L, respectively. ASA-mediated inhibition of cell migration and pigmentation was rescued by exogenous PGE2 or Compound C, which inhibits AMPK activation. Levels of tyrosinase, MITF, and p-ERK were unaffected by ASA exposure. Following a single oral dose of 0.4 mg ASA to NOD/SCID mice, salicylate was detected in plasma and skin at 4 hours and PGE2 levels were reduced up to 24 hours. Some human melanoma tumors xenografted into NOD/SCID mice were sensitive to chronic daily ASA administration, exhibiting reduced growth and proliferation. ASA-treated mice bearing sensitive and resistant tumors exhibited both decreased PGE2 in plasma and tumors and increased phosphorylated AMPK in tumors. We conclude that ASA inhibits colony formation, cell motility, and pigmentation through suppression of PGE2 and activation of AMPK and reduces growth of some melanoma tumors in vivo. This preclinical model could be used for further tumor and biomarker studies to support future melanoma chemoprevention trials in humans. Cancer Prev Res; 11(10); 629–42. ©2018 AACR.

Aberrant lipid metabolism promotes the development of skeletal muscle insulin resistance, but the exact identity of lipid-mediated mechanisms relevant to human obesity remains unclear. A comprehensive lipidomic analysis of primary myocytes from individuals who were insulin-sensitive and lean (LN) or insulin-resistant with obesity (OB) revealed several species of lysophospholipids (lyso-PLs) that were differentially abundant. These changes coincided with greater expression of lysophosphatidylcholine acyltransferase 3 (LPCAT3), an enzyme involved in phospholipid transacylation (Lands cycle). Strikingly, mice with skeletal muscle-specific knockout of LPCAT3 (LPCAT3-MKO) exhibited greater muscle lysophosphatidylcholine/phosphatidylcholine, concomitant with improved skeletal muscle insulin sensitivity. Conversely, skeletal muscle-specific overexpression of LPCAT3 (LPCAT3-MKI) promoted glucose intolerance. The absence of LPCAT3 reduced phospholipid packing of cellular membranes and increased plasma membrane lipid clustering, suggesting that LPCAT3 affects insulin receptor phosphorylation by modulating plasma membrane lipid organization. In conclusion, obesity accelerates the skeletal muscle Lands cycle, whose consequence might induce the disruption of plasma membrane organization that suppresses muscle insulin action.

Aberrant lipid metabolism promotes the development of skeletal muscle insulin resistance, but the exact identity of lipid-mediated mechanisms relevant to human obesity remains unclear. A comprehensive lipidomic analyses of primary myocytes from lean insulin-sensitive (LN) and obese insulin-resistant (OB) individuals revealed several species of lysophospholipids (lyso-PL) that were differentially-abundant. These changes coincided with greater expression of lysophosphatidylcholine acyltransferase 3 (LPCAT3), an enzyme involved in phospholipid transacylation (Lands cycle). Strikingly, mice with skeletal muscle-specific knockout of LPCAT3 (LPCAT3-MKO) exhibited greater muscle lyso-PC/PC, concomitant with greater insulin sensitivityin vivoand insulin-stimulated skeletal muscle glucose uptakeex vivo. Absence of LPCAT3 reduced phospholipid packing of the cellular membranes and increased plasma membrane lipid clustering, suggesting that LPCAT3 affects insulin receptor phosphorylation by modulating plasma membrane lipid organization. In conclusion, obesity accelerates the skeletal muscle Lands cycle, whose consequence might induce the disruption of plasma membrane organization that suppresses muscle insulin action.

Mitochondrial membrane lipids adapt to exercise or disuse, underscoring an essential role for PE in energy transduction., Exercise capacity is a strong predictor of all-cause mortality. Skeletal muscle mitochondrial respiratory capacity, its biggest contributor, adapts robustly to changes in energy demands induced by contractile activity. While transcriptional regulation of mitochondrial enzymes has been extensively studied, there is limited information on how mitochondrial membrane lipids are regulated. Here, we show that exercise training or muscle disuse alters mitochondrial membrane phospholipids including phosphatidylethanolamine (PE). Addition of PE promoted, whereas removal of PE diminished, mitochondrial respiratory capacity. Unexpectedly, skeletal muscle–specific inhibition of mitochondria-autonomous synthesis of PE caused respiratory failure because of metabolic insults in the diaphragm muscle. While mitochondrial PE deficiency coincided with increased oxidative stress, neutralization of the latter did not rescue lethality. These findings highlight the previously underappreciated role of mitochondrial membrane phospholipids in dynamically controlling skeletal muscle energetics and function.

Objective Phosphatidylethanolamine methyltransferase (PEMT) generates phosphatidylcholine (PC), the most abundant phospholipid in the mitochondria and an important acyl chain donor for cardiolipin (CL) biosynthesis. Mice lacking PEMT (PEMTKO) are cold-intolerant when fed a high-fat diet (HFD) due to unclear mechanisms. The purpose of this study was to determine whether PEMT-derived phospholipids are important for the function of uncoupling protein 1 (UCP1) and thus for maintenance of core temperature. Methods To test whether PEMT-derived phospholipids are important for UCP1 function, we examined cold-tolerance and brown adipose (BAT) mitochondria from PEMTKO mice with or without HFD feeding. We complemented these studies with experiments on mice lacking functional CL due to tafazzin knockdown (TAZKD). We generated several conditional mouse models to study the tissue-specific roles of PEMT, including mice with BAT-specific knockout of PEMT (PEMT-BKO). Results Chow- and HFD-fed PEMTKO mice completely lacked UCP1 protein in BAT, despite a lack of difference in mRNA levels, and the mice were accordingly cold-intolerant. While HFD-fed PEMTKO mice exhibited reduced mitochondrial CL content, this was not observed in chow-fed PEMTKO mice or TAZKD mice, indicating that the lack of UCP1 was not attributable to CL deficiency. Surprisingly, the PEMT-BKO mice exhibited normal UCP1 protein levels. Knockout of PEMT in the adipose tissue (PEMT-AKO), liver (PEMT-LKO), or skeletal muscle (PEMT-MKO) also did not affect UCP1 protein levels, suggesting that lack of PEMT in other non-UCP1-expressing cells communicates to BAT to suppress UCP1. Instead, we identified an untranslated UCP1 splice variant that was triggered during the perinatal period in the PEMTKO mice. Conclusions PEMT is required for UCP1 splicing that yields functional protein. This effect is derived by PEMT in nonadipocytes that communicates to BAT during embryonic development. Future research will focus on identifying the non-cell-autonomous PEMT-dependent mechanism of UCP1 splicing., Highlights • Whole-body ablation of PEMT results in a complete loss of UCP1 protein. • While PEMT-derived PC may participate in CL biosynthesis, changes in mitochondrial PC or CL do not explain the loss of UCP1 protein. • PEMT regulates brown adipose UCP1 in a non-cell-autonomous manner. • PEMT deletion promotes the use of an alternative 3’ splice junction in exon 5 of UCP1, preventing UCP1 protein expression.

Traumatic brain injury (TBI) is the leading cause of acquired neurologic disability in children, particularly in those under four years old. During this period, rapid brain growth demands higher Docosahexaenoic Acid (DHA) intake. DHA is an essential fatty acid and brain cell component derived almost entirely from the diet. DHA improved neurologic outcomes and decreased inflammation after controlled cortical impact (CCI) in 17-day old (P17) rats, our established model of pediatric TBI. In adult rodents, TBI decreases brain DHA. We hypothesized that CCI would decrease rat brain DHA at post injury day (PID) 60, blunted by 0.1% DHA diet. We quantitated fatty acids using Gas Chromatography-Mass Spectrometry. We provided 0.1% DHA before CCI to ensure high DHA in dam milk. We compared brain DHA in rats after 60 days of regular (REG) or DHA diet to SHAM pups on REG diet. Brain DHA decreased in REGCCI, not in DHACCI, relative to SHAMREG. In a subsequent experiment, we gave rat pups DHA or vehicle intraperitoneally after CCI followed by DHA or REG diet for 60 days. REG increased brain Docosapentaenoic Acid (n-6 DPA, a brain DHA deficiency marker) relative to SHAMDHA and DHACCI pups (p 0.001, diet effect). DHA diet nearly doubled DHA and decreased n-6 DPA in blood but did not increase brain DHA content (p 0.0001, diet effect). We concluded that CCI or craniotomy alone induces a mild DHA deficit as shown by increased brain DPA.

A conventional metabolic pathway leads to a specific product. In stark contrast, there are diversity-generating metabolic pathways that naturally produce different chemicals, sometimes of great diversity. We demonstrate that for one such pathway, tru, each ensuing metabolic step is slower, in parallel with the increasing potential chemical divergence generated as the pathway proceeds. Intermediates are long lived and accumulate progressively, in contrast with conventional metabolic pathways, in which the first step is rate-limiting and metabolic intermediates are short-lived. Understanding these fundamental differences enables several different practical applications, such as combinatorial biosynthesis, some of which we demonstrate here. We propose that these principles may provide a unifying framework underlying diversity-generating metabolism in many different biosynthetic pathways.

There are conflicting epidemiologic data on whether chronic aspirin (ASA) use may reduce melanoma risk in humans. Potential anticancer effects of ASA may be mediated by its ability to suppress prostaglandin E(2) (PGE(2)) production and activate 5’-adenosine monophosphate–activated protein kinase (AMPK). We investigated the inhibitory effects of ASA in a panel of melanoma and transformed melanocyte cell lines, and on tumor growth in a preclinical model. ASA and the COX-2 inhibitor celecoxib did not affect melanoma cell viability, but significantly reduced colony formation, cell motility, and pigmentation (melanin production) in vitro at concentrations of 1 mmol/L and 20 μmol/L, respectively. ASA-mediated inhibition of cell migration and pigmentation was rescued by exogenous PGE(2) or Compound C, which inhibits AMPK activation. Levels of tyrosinase, MITF, and p-ERK were unaffected by ASA exposure. Following a single oral dose of 0.4 mg ASA to NOD/SCID mice, salicylate was detected in plasma and skin at 4 hours and PGE(2) levels were reduced up to 24 hours. Some human melanoma tumors xenografted into NOD/SCID mice were sensitive to chronic daily ASA administration, exhibiting reduced growth and proliferation. ASA-treated mice bearing sensitive and resistant tumors exhibited both decreased PGE(2) in plasma and tumors and increased phosphorylated AMPK in tumors. We conclude that ASA inhibits colony formation, cell motility, and pigmentation through suppression of PGE(2) and activation of AMPK and reduces growth of some melanoma tumors in vivo. This preclinical model could be used for further tumor and biomarker studies to support future melanoma chemoprevention trials in humans.

Bacterial biofilms are responsible for many persistent infections by many clinically relevant pathogens such as Staphylococcus aureus and Pseudomonas aeruginosa. Biofilms are much more resistant to conventional antibiotics than their planktonic counterparts. Quorum sensing, an intercellular communication system, controls pathogenesis and biofilm formation in most bacterial species. Quorum sensing provides an important pharmacological target since its inhibition does not provide a selective pressure for resistance. In this study, we investigated the quorum sensing and biofilm inhibitory activities of 126 plant extracts from 71 species collected from neotropical rainforests in Costa Rica. Quorum sensing and biofilm interference were assessed using a modified disc diffusion bioassay with Chromobacterium violaceum ATCC 12 472 and a spectrophotometric bioassay with Pseudomonas aeruginosa PA14, respectively. Species with significant anti-quorum sensing and/or anti-biofilm activities belonged to the Meliaceae, Melastomataceae, Lepidobotryaceae, Sapindaceae, and Simaroubaceae families. IC50 values ranged from 45 to 266 µg/mL. Extracts of these active species could lead to future development of botanical treatments for biofilm-associated infections.

Host defenses against pathogens are energetically expensive, leading ecological immunologists to postulate that they might participate in energetic trade-offs with other maintenance programs. However, the metabolic costs of immunity and the nature of physiologic trade-offs it engages are largely unknown. We report here that activation of immunity causes an energetic trade-off with the homeothermy (the stable maintenance of core temperature), resulting in hypometabolism and hypothermia. This immunity-induced physiologic trade-off was independent of sickness behaviors but required hematopoietic sensing of lipopolysaccharide (LPS) via the toll-like receptor 4 (TLR4). Metabolomics and genome-wide expression profiling revealed that distinct metabolic programs supported entry and recovery from the energy-conserving hypometabolic state. During bacterial infections, hypometabolic states, which could be elicited by competition for energy between maintenance programs or energy restriction, promoted disease tolerance. Together, our findings suggest that energy-conserving hypometabolic states, such as dormancy, might have evolved as a mechanism of tissue tolerance.

Detailed investigation of the Antarctic nudibranch Austrodoris kerguelenensis has resulted in the isolation of a diverse suite of new diterpenoid glyceride esters (1–16) related to the palmadorins (17–19), including one (palmadorin L (9)) that is the first halogenated diterpene from this well-studied nudibranch. Previous collections of A. kerguelenensis from McMurdo Sound, the Weddell Sea and the Western Antarctic Peninsula have afforded related diterpene glycerides, a natural product class implicated as a chemical defense in nudibranchs. In this paper we describe the isolation, structure elucidation, and stereochemical analysis of sixteen new palmadorins using a combination of one- and two-dimensional NMR techniques. Palmadorin A (17), B (18), D (1), M (10), N (11), and O (12) inhibit human erythroleukemia (HEL) cells with low micromolar IC50's, and palmadorin M inhibits Jak2, STAT5, and Erk1/2 activation in HEL cells and causes apoptosis, at 5 μM.