Aims/hypothesis Lysosomal acid lipase (LAL) hydrolyses cholesteryl esters and triacylglycerols (TG) within lysosomes to mobilise NEFA and cholesterol. Since LAL-deficient (Lal ^-/- ) mice suffer from progressive loss of adipose tissue and severe accumulation of lipids in hepatic lysosomes, we hypothesised that LAL deficiency triggers alternative energy pathway(s). Methods We studied metabolic adaptations in Lal ^-/- mice. Results Despite loss of adipose tissue, Lal ^-/- mice show enhanced glucose clearance during insulin and glucose tolerance tests and have increased uptake of [³H]2-deoxy-D-glucose into skeletal muscle compared with wild-type mice. In agreement, fasted Lal ^-/- mice exhibit reduced glucose and glycogen levels in skeletal muscle. We observed 84% decreased plasma leptin levels and significantly reduced hepatic ATP, glucose, glycogen and glutamine concentrations in fed Lal ^-/- mice. Markedly reduced hepatic acyl-CoA concentrations decrease the expression of peroxisome proliferator-activated receptor α (PPARα) target genes. However, treatment of Lal ^-/- mice with the PPARα agonist fenofibrate further decreased plasma TG (and hepatic glucose and glycogen) concentrations in Lal ^-/- mice. Depletion of hepatic nuclear factor 4α and forkhead box protein a2 in fasted Lal ^-/- mice might be responsible for reduced expression of microsomal TG transfer protein, defective VLDL synthesis and drastically reduced plasma TG levels. Conclusions/interpretation Our findings indicate that neither activation nor inactivation of PPARα per se but rather the availability of hepatic acyl-CoA concentrations regulates VLDL synthesis and subsequent metabolic adaptations in Lal ^-/- mice. We conclude that decreased plasma VLDL production enhances glucose uptake into skeletal muscle to compensate for the lack of energy supply.

Aims: Lipotoxic cardiomyopathy in diabetic and obese patients typically encompasses increased cardiac fatty acid (FA) uptake eventually surpassing the mitochondrial oxidative capacity. Lowering FA utilization via inhibition of lipolysis represents a strategy to counteract the development of lipotoxic heart dysfunction. However, defective cardiac triacylglycerol (TAG) catabolism and FA oxidation in humans (and mice) carrying mutated ATGL alleles provokes lipotoxic heart dysfunction questioning a therapeutic approach to decrease cardiac lipolysis. Interestingly, decreased lipolysis via cardiac overexpression of Perilipin 5 (Plin5), a binding partner of ATGL, is compatible with normal heart function and lifespan despite massive cardiac lipid accumulation. Herein, we decipher mechanisms that protect Plin5 transgenic mice from the development of heart dysfunction. Methods and results: We generated mice with cardiac-specific overexpression of Plin5 encoding a serine-155 to alanine exchange (Plin5-S155A) of the protein kinase A phosphorylation site, which has been suggested as a prerequisite to stimulate lipolysis and may play a crucial role in the preservation of heart function. Plin5-S155A mice showed a substantial increase in cardiac TAG and ceramide levels, which was comparable to mice overexpressing non-mutated Plin5. Lipid accumulation was compatible with normal heart function even under mild stress. Plin5-S155A mice showed reduced cardiac FA oxidation but normal ATP production and changes in the Plin5-S155A phosphoproteome compared to Plin5 transgenic mice. Interestingly, mitochondrial recruitment of dynamin-related protein 1 (Drp1) was markedly reduced in cardiac muscle of Plin5-S155A and Plin5 transgenic mice accompanied by decreased phosphorylation of mitochondrial fission factor, a mitochondrial receptor of Drp1. Conclusions: This study suggests that low cardiac lipolysis is associated with reduced mitochondrial fission and may represent a strategy to combat the development of lipotoxic heart dysfunction.

Determination of the ratio of reduced to oxidized glutathione is of profound clinical interest in assessing the oxidative status of tissues and body fluids. However, this ratio is not yet a routine clinical parameter due to the analytically challenging interconversion of reduced (free) glutathione to oxidized (bound) glutathione. We aimed to facilitate this ratio determination in order to aid its incorporation as a routine clinical parameter. To this end, we developed a simple derivatization route that yields different isotopologues of N-ethylmaleimide alkylated glutathione from reduced and oxidized glutathione (after its chemical reduction) for mass spectrometric analysis. A third isotopologue can be used as isotopic standard for simultaneous absolute quantification. As all isotopologues have similar chromatographic properties, matrix effects arising from different sample origins can only impact method sensitivity but not quantification accuracy. Robustness, simplified data analysis, cost effectiveness by one common standard, and highly improved mass spectrometric sensitivity by conversion of oxidized glutathione to an alkylated glutathione isotopologue are the main advantages of our approach. We present a method fully optimized for blood, plasma, serum, cell, and tissue samples. In addition, we propose production of N-ethylmaleimide customized blood collection tubes to even further facilitate the analysis in a clinical setting.

Cheap and renewable feedstocks such as the one-carbon substrate formate are emerging for sustainable production in a growing chemical industry. We investigated the acetogen Acetobacterium woodii as a potential host for bioproduction from formate alone and together with autotrophic and heterotrophic co-substrates by quantitatively analyzing physiology, transcriptome, and proteome in chemostat cultivations in combination with computational analyses. Continuous cultivations with a specific growth rate of 0.05 h^-1 on formate showed high specific substrate uptake rates (47 mmol g^-1 h^-1). Co-utilization of formate with H₂, CO, CO₂ or fructose was achieved without catabolite repression and with acetate as the sole metabolic product. A transcriptomic comparison of all growth conditions revealed a distinct adaptation of A. woodii to growth on formate as 570 genes were changed in their transcript level. Transcriptome and proteome showed higher expression of the Wood-Ljungdahl pathway during growth on formate and gaseous substrates, underlining its function during utilization of one-carbon substrates. Flux balance analysis showed varying flux levels for the WLP (0.7-16.4 mmol g^-1 h^-1) and major differences in redox and energy metabolism. Growth on formate, H₂/CO₂, and formate + H₂/CO₂ resulted in low energy availability (0.20-0.22 ATP/acetate) which was increased during co-utilization with CO or fructose (0.31 ATP/acetate for formate + H₂/CO/CO₂, 0.75 ATP/acetate for formate + fructose). Unitrophic and mixotrophic conversion of all substrates was further characterized by high energetic efficiencies. In silico analysis of bioproduction of ethanol and lactate from formate and autotrophic and heterotrophic co-substrates showed promising energetic efficiencies (70-92%). Collectively, our findings reveal A. woodii as a promising host for flexible and simultaneous bioconversion of multiple substrates, underline the potential of substrate co-utilization to improve the energy availability of acetogens and encourage metabolic engineering of acetogenic bacteria for the efficient synthesis of bulk chemicals and fuels from sustainable one carbon substrates.

Introduction: Development of specific biomarkers aiding early diagnosis of heart failure is an ongoing challenge. Biomarkers commonly used in clinical routine usually act as readouts of an already existing acute condition rather than disease initiation. Functional decline of cardiac muscle is greatly aggravated by increased oxidative stress and damage of proteins. Oxidative post-translational modifications occur already at early stages of tissue damage and are thus regarded as potential up-coming disease markers. Areas covered: Clinical practice regarding commonly used biomarkers for heart disease is briefly summarized. The types of oxidative post-translational modification in cardiac pathologies are discussed with a special focus on available quantitative techniques and characteristics of individual modifications with regard to their stability and analytical accessibility. As irreversible oxidative modifications trigger protein degradation pathways or cause protein aggregation, both influencing biomarker abundance, a chapter is dedicated to their regulation in the heart.