Differential utilization of vitamin B12-dependent and independent pathways for propionate metabolism across human cells.

Propionic acid links the oxidation of branched-chain amino acids and odd-chain fatty acids to the TCA cycle. Gut microbes ferment complex fiber remnants, generating high concentrations of short chain fatty acids, acetate, propionate and butyrate, which are shared with the host as fuel sources. Analysis of vitamin B12-dependent propionate utilization in skin biopsy samples has been used to characterize and diagnose underlying inborn errors of cobalamin (or B12) metabolism. In these cells, the B12-dependent enzyme, methylmalonyl-CoA mutase (MMUT), plays a central role in funneling propionate to the TCA cycle intermediate, succinate. Our understanding of the fate of propionate in other cell types, specifically, the involvement of the ß-oxidation-like and methylcitrate pathways, is limited. In this study, we have used [14C]-propionate tracing in combination with genetic ablation or inhibition of MMUT, to reveal the differential utilization of the B12-dependent and independent pathways for propionate metabolism in fibroblast versus colon cell lines. We demonstrate that itaconate can be used as a tool to investigate MMUT-dependent propionate metabolism in cultured cell lines. While MMUT gates the entry of propionate carbons into the TCA cycle in fibroblasts, colon-derived cell lines exhibit a quantitatively significant or exclusive reliance on the ß-oxidation-like pathway. Lipidomics and metabolomics analyses reveal that propionate elicits pleiotropic changes, including an increase in odd-chain glycerophospholipids, and perturbations in the purine nucleotide cycle and arginine/nitric oxide metabolism. The metabolic rationale and the regulatory mechanisms underlying the differential reliance on propionate utilization pathways at a cellular, and possibly tissue level, warrant further elucidation.

Acetyl-CoA carboxylase Inhibition increases retinal pigment epithelial cell fatty acid flux and restricts apolipoprotein efflux.

Lipid-rich deposits called drusen accumulate under the retinal pigment epithelium (RPE) in the eyes of patients with age-related macular degeneration (AMD) and Sorsby's fundus dystrophy (SFD). Drusen may contribute to photoreceptor and RPE degeneration in these blinding diseases. We hypothesize that stimulating ß-oxidation of fatty acids could decrease the availability of lipid with which RPE cells can generate drusen. Inhibitors of acetyl-CoA carboxylase (ACC) stimulate ß-oxidation and diminish lipid accumulation in fatty liver disease. In this report we test the hypothesis that an ACC inhibitor, Firsocostat, can diminish lipid deposition by RPE cells. We probed metabolism and cellular function in mouse RPE-choroid tissue and in cultured human RPE cells. We used 13C6-glucose, 13C16-palmitate, and gas chromatography-linked mass spectrometry to monitor effects of Firsocostat on glycolytic, Krebs cycle, and fatty acid metabolism. We quantified lipid abundance, apolipoprotein E (ApoE) and vascular endothelial growth factor (VEGF) release using liquid chromatography-mass spectrometry, enzyme-linked immunosorbent assays and localized ApoE deposits by immunostaining. RPE barrier function was assessed by trans-epithelial electrical resistance (TEER). Firsocostat-mediated ACC inhibition increases ß-oxidation, decreases intracellular lipid levels, diminishes lipoprotein release, and increases TEER. When human serum or outer segments are used to stimulate lipoprotein release, fewer lipoproteins are released in the presence of either lipid source and Firsocostat. In a culture model of SFD, Firsocostat stimulates fatty acid oxidation, increases TEER, and decreases ApoE release. We conclude that Firsocostat remodels RPE metabolism and can limit lipid deposition. This suggests that ACC inhibition could be an effective strategy for diminishing pathologic drusen in the eyes of patients with AMD or SFD.

ACAD10 and ACAD11 allow entry of 4-hydroxy fatty acids into ß-oxidation.

Hydroxylated fatty acids are important intermediates in lipid metabolism and signaling. Surprisingly, the metabolism of 4-hydroxy fatty acids remains largely unexplored. We found that both ACAD10 and ACAD11 unite two enzymatic activities to introduce these metabolites into mitochondrial and peroxisomal ß-oxidation, respectively. First, they phosphorylate 4-hydroxyacyl-CoAs via a kinase domain, followed by an elimination of the phosphate to form enoyl-CoAs catalyzed by an acyl-CoA dehydrogenase (ACAD) domain. Studies in knockout cell lines revealed that ACAD10 preferentially metabolizes shorter chain 4-hydroxy fatty acids than ACAD11 (i.e. 6 carbons versus 10 carbons). Yet, recombinant proteins showed comparable activity on the corresponding 4-hydroxyacyl-CoAs. This suggests that the localization of ACAD10 and ACAD11 to mitochondria and peroxisomes, respectively, might influence their physiological substrate spectrum. Interestingly, we observed that ACAD10 is cleaved internally during its maturation generating a C-terminal part consisting of the ACAD domain, and an N-terminal part comprising the kinase domain and a haloacid dehalogenase (HAD) domain. HAD domains often exhibit phosphatase activity, but negligible activity was observed in the case of ACAD10. Yet, inactivation of a presumptive key residue in this domain significantly increased the kinase activity, suggesting that this domain might have acquired a regulatory function to prevent accumulation of the phospho-hydroxyacyl-CoA intermediate. Taken together, our work reveals that 4-hydroxy fatty acids enter mitochondrial and peroxisomal fatty acid ß-oxidation via two enzymes with an overlapping substrate repertoire.

CLPX regulates mitochondrial fatty acid ß-oxidation in liver cells.

Mitochondrial fatty acid oxidation (ß-oxidation) is an essential metabolic process for energy production in eukaryotic cells, but the regulatory mechanisms of this pathway are largely unknown. In the present study, we found that several enzymes involved in ß-oxidation are associated with CLPX, the AAA+ unfoldase that is a component of the mitochondrial matrix protease ClpXP. The suppression of CLPX expression increased ß-oxidation activity in the HepG2 cell line and in primary human hepatocytes without glucagon treatment. However, the protein levels of enzymes involved in ß-oxidation did not significantly increase in CLPX-deleted HepG2 cells (CLPX-KO cells). Coimmunoprecipitation experiments revealed that the protein level in the immunoprecipitates of each antibody changed after the treatment of WT cells with glucagon, and a part of these changes was also observed in the comparison of WT and CLPX-KO cells without glucagon treatment. Although the exogenous expression of WT or ATP-hydrolysis mutant CLPX suppressed ß-oxidation activity in CLPX-KO cells, glucagon treatment induced ß-oxidation activity only in CLPX-KO cells expressing WT CLPX. These results suggest that the dissociation of CLPX from its target proteins is essential for the induction of ß-oxidation in HepG2 cells. Moreover, specific phosphorylation of AMP-activated protein kinase and a decrease in the expression of acetyl-CoA carboxylase 2 were observed in CLPX-KO cells, suggesting that CLPX might participate in the regulation of the cytosolic signaling pathway for ß-oxidation. The mechanism for AMP-activated protein kinase phosphorylation remains elusive; however, our results uncovered the hitherto unknown role of CLPX in mitochondrial ß-oxidation in human liver cells.

Crystal structure combined with metabolomics and biochemical studies indicates that FAM3A participates in fatty acid beta-oxidation upon binding of acyl-L-carnitine.

As the first member of the family with sequence similarity 3 (FAM3), FAM3A promotes synthesis of ATP in mitochondria of hepatic cells and cells from other organs. Dysregulations of FAM3A are involved in the development of diabetes and nonalcoholic fatty liver disease (NAFLD). So far, the molecule mechanism under the physiological and pathological functions of FAM3A is largely unexplored. Here, we determined the crystal structure of FAM3A at high resolution of 1.38Å, complexed with an unknown-source compound which was characterized through metabolomics and confirmed as methacholine by thermal shift assay and surface plasmon resonance (SPR). Exploration for natural ligands of FAM3A was conducted through the same molecular interaction assays. The observed binding of acyl-L-carnitine molecules indicated FAM3A participating in fatty acid beta-oxidation. Knockdown and rescue assays coupled with fatty acid oxidation determination confirmed the role of FAM3A in beta-oxidation. This investigation reveals the molecular mechanism for the biological function of FAM3A and would provide basis for identifying drug target for treatment of diabetes and NAFLD.

Lipid metabolism in focus: how the build-up and breakdown of lipids affects stem cells.

Cellular metabolism has recently emerged as a key regulator of stem cell behavior. Various studies have suggested that metabolic regulatory mechanisms are conserved in different stem cell niches, suggesting a common level of stem cell regulation across tissues. Although the balance between glycolysis and oxidative phosphorylation has been shown to be distinct in stem cells and their differentiated progeny, much less is known about lipid metabolism in stem cell regulation. In this Review, we focus on how stem cells are affected by two major lipid metabolic pathways: the build-up of lipids, called de novo lipogenesis, and the breakdown of lipids, called fatty acid beta-oxidation. We cover the recent literature on hematopoietic stem cells, intestinal stem cells, neural stem/progenitor cells and cancer stem cells, where these two lipid pathways have been studied in more depth.

Long-term prognosis of fatty-acid oxidation disorders in adults: Optimism despite the limited effective therapies available.

INTRODUCTION: Fatty-acid oxidation disorders (FAODs) are recessive genetic diseases. MATERIALS AND METHODS: We report here clinical and paraclinical data from a retrospective study of 44 adults with muscular FAODs from six French reference centers for neuromuscular or metabolic diseases. RESULTS: The study cohort consisted of 44 adult patients: 14 with carnitine palmitoyl transferase 2 deficiency (32%), nine with multiple acyl-CoA deficiency (20%), 13 with very long-chain acyl-CoA dehydrogenase deficiency (30%), three with long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (7%), and five with short-chain acyl-CoA dehydrogenase deficiency (11%). Disease onset occurred during childhood in the majority of patients (59%), with a mean age at onset of 15 years (range = 0.5-35) and a mean of 12.6 years (range = 0-58) from disease onset to diagnosis. The principal symptoms were acute muscle manifestations (rhabdomyolysis, exercise intolerance, myalgia), sometimes associated with permanent muscle weakness. Episodes of rhabdomyolysis were frequent (84%), with a mean creatinine kinase level of 68,958 U/L (range = 660-300,000). General metabolic complications were observed in 58% of patients, respiratory manifestations in 18% of cases, and cardiological manifestations in 9% of cases. Fasting acylcarnitine profile was used to orient genetic explorations in 65% of cases. After a mean follow-up of 10 years, 33% of patients were asymptomatic and 56% continued to display symptoms after exercise. The frequency of rhabdomyolysis decreased after diagnosis in 64% of cases. CONCLUSION: A standardized register would complete this cohort description of muscular forms of FAODs with exhaustive data, making it possible to assess the efficacy of therapeutic protocols in real-life conditions and during the long-term follow-up of patients.

Association between F2-Isoprostane Metabolites and Weight Change in Older Women: A Longitudinal Analysis.

INTRODUCTION: Theoretically, some metabolic traits may predispose older individuals to weight loss during aging, leading to increased all-cause mortality and many serious health issues. Biomarkers to robustly predict progressive weight loss during aging are, however, lacking. We prospectively assessed if urinary levels of F2-isoprostanes and their peroxisomal ß-oxidation metabolite, 2,3-dinor-5,6-dihydro-15-F2t-isoprostane (F2-IsoP-M), were associated with subsequent weight loss in middle-aged and older women. METHODS: Included in the analysis were 2,066 women aged 40-70 years, a subset of a prospective cohort study. F2-isoprostanes (F2-IsoPs) and its ß-oxidation metabolite, F2-IsoP-M, were measured in urine using gas chromatography-mass spectrometry. Measurements of anthropometry and exposures to major determinants of body weight were performed at baseline and repeated thrice over 15-year follow-up. The longitudinal associations of F2-IsoP-M and the F2-IsoP-M to its parent compound, F2-IsoP, ratio (MPR) with repeatedly measured weight changes were examined using linear mixed-effect models. RESULTS: After adjusting for time-varying covariates: energy intake, physical activity, and comorbidity index, among others, levels of F2-IsoP-M and the MPR were both inversely associated with percentage of weight change. Weight in the highest quartile of these two biomarkers was 1.33% (95% CI = -2.41, -0.24) and 1.09% (95% CI = -2.16, -0.02) lower than those in the lowest quartile group, with p for trend of 0.01 and 0.03, respectively. The inverse association was consistently seen across follow-up periods, although appearing stronger with prolonged follow-up. There was no association between the parent compound, F2-IsoPs, and weight change. CONCLUSION: This study demonstrates the first piece of evidence to associate F2-IsoP metabolism, peroxisomal ß-oxidation, with weight loss in older women. Further investigations into the role of lipid peroxidation and peroxisomal ß-oxidation in weight change among older individuals are warranted.

Molecular characteristics of the multi-functional FAO enzyme ACAD9 illustrate the importance of FADH2 /NADH ratios for mitochondrial ROS formation.

A decade ago I postulated that ROS formation in mitochondria was influenced by different FADH2 /NADH (F/N) ratios of catabolic substrates. Thus, fatty acid oxidation (FAO) would give higher ROS formation than glucose oxidation. Both the emergence of peroxisomes and neurons not using FAO, could be explained thus. ROS formation in NADH:ubiquinone oxidoreductase (Complex I) comes about by reverse electron transport (RET) due to high QH2 levels, and scarcity of its electron-acceptor (Q) during FAO. The then new, unexpected, finding of an FAO enzyme, ACAD9, being involved in complex I biogenesis, hinted at connections in line with the hypothesis. Recent findings about ACAD9's role in regulation of respiration fit with predictions the model makes: cementing connections between ROS production and F/N ratios. I describe how ACAD9 might be central to reversing the oxidative damage in complex I resulting from FAO. This seems to involve two distinct, but intimately connected, ACAD9 characteristics: (i) its upregulation of complex I biogenesis, and (ii) releasing FADH2 , with possible conversion into FMN, the crucial prosthetic group of complex I. Also see the video abstract here: https://youtu.be/N7AT_HBNumg.

Structural insights into the pathogenicity of point mutations in human acyl-CoA dehydrogenase homotetramers.

Acyl-CoA dehydrogenase deficiency (ACAD) is an inherited and potentially fatal disorder with variable clinical symptoms. The relationship between pathogenicity and deleterious point mutations is investigated here in ACAD structures of short (SCAD) and medium-chain (MCAD) types. Structures and dynamic features of native and mutant forms of enzymes models were compared. A total of 2.88 µs molecular dynamics simulations were performed at four different temperatures. Total energy, RMSD, protein ligand interactions and affinity, RMSF measures, secondary structure changes, and important interactions were studied. Mutations in the three main domains of ACADs are pathogenic, while those located at linker turns are not. Mutations affect mostly tetramer formations, secondary structures, and many contacts and interactions. In R206H (MCAD mutant) which is experimentally known to cause a huge turnover decrease, the lack of a single H-bond between substrate and FAD was observed. Secondary structures showed temperature-dependent changes, and SCAD activity was found to be highly correlated to the enzyme helix 3-10 content. Finally, RMSF patterns pointed to one important loop that maintains the substrate close to the active site and is a cause of substrate wobbling upon mutation. Despite similar structure, function, and cellular location, SCAD and MCAD may have different optimum temperatures that are related to the structure taken at that specific temperature. In conclusion, new insight has been provided on the effect of various SCAD and MCAD pathogenic mutations on the structure and dynamical features of the enzymes.

Myo-inositol alters the effects of glucose, leptin and insulin on placental palmitic acid and oleic acid metabolism.

Well-regulated placental palmitic acid (PA) and oleic acid (OA) metabolism is vital for optimal placental function and fetal development, but dysregulation occurs with gestational diabetes (GDM). We hypothesized that such dysregulation might arise from increased maternofetal glucose, leptin or insulin concentrations present in GDM, and that dysregulated PA and OA lipid metabolism could be moderated by myo-inositol, a natural polyol and potential GDM intervention. Placental explants from 21 women were incubated with stable isotope-labelled 13 C-PA or 13 C-OA for 48 h. Explants were treated with glucose (5, 10 mm) or leptin (13 nm) or insulin (150 nm) in combination with myo-inositol (0.3, 30, 60 µm). Forty-seven 13 C-PA lipids and 37 13 C-OA lipids were measured by liquid chromatography-mass spectrometry (LCMS). Compared with controls (5 mm glucose), glucose (10 mm) increased 19 13 C-OA lipids and nine 13 C-PA lipids, but decreased 13 C-OA phosphatidylethanolamine 38:5 and 13 C-PA phosphatidylethanolamine 36:4. The effects of leptin and insulin were less prominent than glucose, with leptin increasing 13 C-OA acylcarnitine 18:1, and insulin increasing four 13 C-PA triacylglycerides. Most glucose, leptin and insulin-induced alterations in lipids were attenuated by co-incubation with myo-inositol (30 or 60 µm), with attenuation also occurring in all subgroups stratified by GDM status and fetal sex. However, glucose-induced increases in acylcarnitine were not attenuated by myo-inositol and were even exaggerated in some instances. Myo-inositol therefore appears to generally act as a moderator, suppressing the perturbation of lipid metabolic processes by glucose, leptin and insulin in placenta in vitro. Whether myo-inositol protects the fetus and pregnancy from unfavourable outcomes requires further research. KEY POINTS: Incubation of placental explants with additional glucose, or to a lesser extent insulin or leptin, alters the placental production of 13 C-lipids from 13 C-palmitic acid (PA) and 13 C-oleic acid (OA) in vitro compared with untreated controls from the same placenta. Co-incubation with myo-inositol attenuated most alterations induced by glucose, insulin or leptin in 13 C-lipids, but did not affect alterations in 13 C-acylcarnitines. Alterations induced by glucose and leptin in 13 C-PA triacylglycerides and 13 C-PA phospholipids were influenced by fetal sex and gestational diabetes status, but were all still attenuated by myo-inositol co-incubation. Insulin differently affected 13 C-PA triacylglycerides and 13 C-PA phospholipids depending on fetal sex, with alterations also attenuated by myo-inositol co-incubation.

Engineering the biological conversion of formate into crotonate in Cupriavidus necator.

To advance the sustainability of the biobased economy, our society needs to develop novel bioprocesses based on truly renewable resources. The C1-molecule formate is increasingly proposed as carbon and energy source for microbial fermentations, as it can be efficiently generated electrochemically from CO2 and renewable energy. Yet, its biotechnological conversion into value-added compounds has been limited to a handful of examples. In this work, we engineered the natural formatotrophic bacterium C. necator as cell factory to enable biological conversion of formate into crotonate, a platform short-chain unsaturated carboxylic acid of biotechnological relevance. First, we developed a small-scale (150-mL working volume) cultivation setup for growing C. necator in minimal medium using formate as only carbon and energy source. By using a fed-batch strategy with automatic feeding of formic acid, we could increase final biomass concentrations 15-fold compared to batch cultivations in flasks. Then, we engineered a heterologous crotonate pathway in the bacterium via a modular approach, where each pathway section was assessed using multiple candidates. The best performing modules included a malonyl-CoA bypass for increasing the thermodynamic drive towards the intermediate acetoacetyl-CoA and subsequent conversion to crotonyl-CoA through partial reverse ß-oxidation. This pathway architecture was then tested for formate-based biosynthesis in our fed-batch setup, resulting in a two-fold higher titer, three-fold higher productivity, and five-fold higher yield compared to the strain not harboring the bypass. Eventually, we reached a maximum product titer of 148.0 ± 6.8 mg/L. Altogether, this work consists in a proof-of-principle integrating bioprocess and metabolic engineering approaches for the biological upgrading of formate into a value-added platform chemical.

Transcriptome analysis of Aurantiochytrium limacinum under low salt conditions.

Aurantiochytrium limacinum can accumulate high amounts of omega-3 polyunsaturated fatty acids, especially docosahexaenoic acid (DHA). Although salinity affects the DHA content, its impact on the metabolic pathway responsible for DHA production in A. limacinum is not completely understood. To address this issue, we investigated the transcriptional profile of A. limacinum under hypoosmotic stress. We first cultured A. limacinum under typical and low salinity for RNA sequencing, respectively. Transcriptome analyses revealed that 933 genes exhibited significant changes in expression under hypoosmotic conditions, of which 81.4% were downregulated. Strikingly, A. limacinum downregulated genes related to polyketide synthesis and fatty acid synthase pathways, while upregulating ß-oxidation-related genes. In accordance with this, DHA production significantly decreased under hypoosmotic conditions, while antioxidant-related genes were significantly upregulated. Considering that ß-oxidation of fatty acids generates energy and reactive oxygen species (ROS), our results suggest that A. limacinum utilizes fatty acids for energy to survive under hypoosmotic conditions and detoxifies ROS using antioxidant systems.

Comparative survey of mitochondrial ultrastructure in IDH1-mutant astrocytoma and IDH1-wildtype glioblastoma (GBM).

Gliomas are the most common malignant brain tumors with poor prognosis. The WHO's classification recognizes isocitrate dehydrogenase 1 (IDH1) mutant astrocytoma and IDH1-wildtype glioblastoma (GBM). The IDH1 mutation confers a survival advantage over the wildtype. There are several explanations for the metabolic advantage of the IDH1 mutation, some involve mitochondrial implications. Since an ultrastructural comparison of both tumor genotypes is still lacking, we surveyed the ultrastructural effects of the IDH1 mutation on the mitochondria of the IDH1-mutant astrocytoma (n = 15) and IDH1-wildtype glioblastoma (n = 15) tumors. Our results show that both IDH1 genotypes have degenerate and uncoupled mitochondria; this has not been reported before. The presence of ample lipid inclusions and lipid droplets in the cytoplasm of both genotypes support our conclusion of dysfunctional uncoupled mitochondria. Thus, the IDH1 mutation may have no ultrastructural consequences on the mitochondria, and the aberrant mitochondria in both genotypes may be the result of other unknown mutations. The status of the mitochondria in these genotypes portends a clinical challenge since tumor cells with uncoupled mitochondria are more primitive, aggressive, and considerably treatment resistant.

The Role of Acyl-CoA ß-Oxidation in Brain Metabolism and Neurodegenerative Diseases.

This review highlights the complex role of fatty acid ß-oxidation in brain metabolism. It demonstrates the fundamental importance of fatty acid degradation as a fuel in energy balance and as an essential component in lipid homeostasis, brain aging, and neurodegenerative disorders.

The Physiological and Pathological Role of Acyl-CoA Oxidation.

Fatty acid metabolism, including ß-oxidation (ßOX), plays an important role in human physiology and pathology. ßOX is an essential process in the energy metabolism of most human cells. Moreover, ßOX is also the source of acetyl-CoA, the substrate for (a) ketone bodies synthesis, (b) cholesterol synthesis, (c) phase II detoxication, (d) protein acetylation, and (d) the synthesis of many other compounds, including N-acetylglutamate-an important regulator of urea synthesis. This review describes the current knowledge on the importance of the mitochondrial and peroxisomal ßOX in various organs, including the liver, heart, kidney, lung, gastrointestinal tract, peripheral white blood cells, and other cells. In addition, the diseases associated with a disturbance of fatty acid oxidation (FAO) in the liver, heart, kidney, lung, alimentary tract, and other organs or cells are presented. Special attention was paid to abnormalities of FAO in cancer cells and the diseases caused by mutations in gene-encoding enzymes involved in FAO. Finally, issues related to α- and ω- fatty acid oxidation are discussed.

Altered Fatty Acid Oxidation in Lymphocyte Populations of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome.

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a disabling multisystem illness in which individuals are plagued with fatigue, inflammatory symptoms, cognitive dysfunction, and the hallmark symptom, post-exertional malaise. While the cause of this disease remains unknown, there is evidence of a potential infectious component that, along with patient symptoms and common onsets of the disease, implicates immune system dysfunction. To further our understanding of the state of ME/CFS lymphocytes, we characterized the role of fatty acids in isolated Natural Killer cells, CD4+ T cells, and CD8+ T cells in circulation and after overnight stimulation, through implicit perturbations to fatty acid oxidation. We examined samples obtained from at least 8 and as many as 20 subjects for immune cell fatty acid characterization in a variety of experiments and found that all three isolated cell types increased their utilization of lipids and levels of pertinent proteins involved in this metabolic pathway in ME/CFS samples, particularly during higher energy demands and activation. In T cells, we characterized the cell populations contributing to these metabolic shifts, which included CD4+ memory cells, CD4+ effector cells, CD8+ naïve cells, and CD8+ memory cells. We also discovered that patients with ME/CFS and healthy control samples had significant correlations between measurements of CD4+ T cell fatty acid metabolism and demographic data. These findings provide support for metabolic dysfunction in ME/CFS immune cells. We further hypothesize about the consequences that these altered fuel dependencies may have on T and NK cell effector function, which may shed light on the illness's mechanism of action.

Hormone-sensitive lipase protects adipose triglyceride lipase-deficient mice from lethal lipotoxic cardiomyopathy.

Lipid droplets (LDs) are multifunctional organelles that regulate energy storage and cellular homeostasis. The first step of triacylglycerol hydrolysis in LDs is catalyzed by adipose triglyceride lipase (ATGL), deficiency of which results in lethal cardiac steatosis. Although hormone-sensitive lipase (HSL) functions as a diacylglycerol lipase in the heart, we hypothesized that activation of HSL might compensate for ATGL deficiency. To test this hypothesis, we crossed ATGL-KO (AKO) mice and cardiac-specific HSL-overexpressing mice (cHSL) to establish homozygous AKO mice and AKO mice with cardiac-specific HSL overexpression (AKO+cHSL). We found that cardiac triacylglycerol content was 160-fold higher in AKO relative to Wt mice, whereas that of AKO+cHSL mice was comparable to the latter. In addition, AKO cardiac tissues exhibited reduced mRNA expression of PPARα-regulated genes and upregulation of genes involved in inflammation, fibrosis, and cardiac stress. In contrast, AKO+cHSL cardiac tissues exhibited expression levels similar to those observed in Wt mice. AKO cardiac tissues also exhibited macrophage infiltration, apoptosis, interstitial fibrosis, impaired systolic function, and marked increases in ceramide and diacylglycerol contents, whereas no such pathological alterations were observed in AKO+cHSL tissues. Furthermore, electron microscopy revealed considerable LDs, damaged mitochondria, and disrupted intercalated discs in AKO cardiomyocytes, none of which were noted in AKO+cHSL cardiomyocytes. Importantly, the life span of AKO+cHSL mice was comparable to that of Wt mice. HSL overexpression normalizes lipotoxic cardiomyopathy in AKO mice and the findings highlight the applicability of cardiac HSL activation as a therapeutic strategy for ATGL deficiency-associated lipotoxic cardiomyopathies.

Impaired beta-oxidation increases vulnerability to influenza A infection.

Influenza A virus (IAV) infection casts a significant burden on society. It has particularly high morbidity and mortality rates in patients suffering from metabolic disorders. The aim of this study was to relate metabolic changes with IAV susceptibility using well-characterized inbred mouse models. We compared the highly susceptible DBA/2J (D2) mouse strain for which IAV infection is lethal with the C57BL/6J (B6) strain, which exhibits a moderate course of disease and survives IAV infection. Previous studies showed that D2 has higher insulin and glucose levels and is predisposed to develop diet-induced type 2 diabetes. Using high-resolution liquid chromatography-coupled MS, the plasma metabolomes of individual animals were repeatedly measured up to 30 days postinfection. The biggest metabolic difference between these strains in healthy and infected states was in the levels of malonylcarnitine, which was consistently increased 5-fold in D2. Other interstrain and intrastrain differences in healthy and infected animals were observed for acylcarnitines, glucose, branched-chain amino acids, and oxidized fatty acids. By mapping metabolic changes to canonical pathways, we found that mitochondrial beta-oxidation is likely disturbed in D2 animals. In noninfected D2 mice, this leads to increased glycerolipid production and reduced acylcarnitine production, whereas in infected D2 animals, peroxisomal beta-oxidation becomes strongly increased. From these studies, we conclude that metabolic changes caused by a distortion of mitochondrial and peroxisomal metabolism might impact the innate immune response in D2, leading to high viral titers and mortality.

New insights into cellular links between sodium-glucose cotransporter-2 inhibitors and ketogenesis.

Sodium-glucose cotransporter-2 inhibitors (SGLT2is) are a newly developed class of highly effective antidiabetic therapies that normalize hyperglycemia via urinary glucose excretion. However, they may be accompanied by certain side effects that negatively impact their therapeutic benefits. SGLT2is induce a metabolic shift from glucose to fatty acids and thus increase lipolysis which, in turn, induces ketogenesis. The complete pathways linking SGLT2is to ketoacidosis have not yet been fully elucidated, though much is now known. Therefore, in this mechanistic study, we present the current knowledge and shed light upon the possible cellular pathways involved. A deeper understanding of the possible links between SGLT2is and ketogenesis could help to prevent adverse side effects in diabetic patients treated with these drugs.