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Aberrant expression of MYC transcription factor family members predicts poor clinical outcome in many human cancers. Oncogenic MYC profoundly alters metabolism and mediates an antioxidant response to maintain redox balance. Here we show that MYCN induces massive lipid peroxidation on depletion of cysteine, the rate-limiting amino acid for glutathione (GSH) biosynthesis, and sensitizes cells to ferroptosis, an oxidative, non-apoptotic and iron-dependent type of cell death. The high cysteine demand of MYCN-amplified childhood neuroblastoma is met by uptake and transsulfuration. When uptake is limited, cysteine usage for protein synthesis is maintained at the expense of GSH triggering ferroptosis and potentially contributing to spontaneous tumor regression in low-risk neuroblastomas. Pharmacological inhibition of both cystine uptake and transsulfuration combined with GPX4 inactivation resulted in tumor remission in an orthotopic MYCN-amplified neuroblastoma model. These findings provide a proof of concept of combining multiple ferroptosis targets as a promising therapeutic strategy for aggressive MYCN-amplified tumors.

By defining God as a maximally great being Plantinga is able to devise an ontological argument which validly infers from the possibility of there being a God that there necessarily is a God. In this article I shall argue that Plantinga’s argument is not only question-begging, as several critics have complained, but circular in the strongest sense of the term. Based on reflections on the relation between the notions of coherence and possibility, I shall defend two arguments, previously proposed by Tooley (Mind 90:422–427, 1981) and Guleserian (Nous 17:221–238, 1983), against the existence of a maximally great being. The article concludes with some critical remarks on Plantinga’s rationale for conceiving God as such a being.

The positivists defined analyticity as truth in virtue of meaning alone and advocated the view that the notion of analyticity so defined is co-extensive with both the notion of an a priori truth an...

Elevated blood lactate levels are frequently found in critically ill patients and thought to result from tissue hypoperfusion and cellular oxygen shortage. Considering the close relationship between immune cell function and intracellular metabolism, lactate is more than a glycolytic waste molecule but able to regulate the immune response. Our aim was to elucidate the temporal and mechanistic effect of extracellular lactate on monocytes. To this end, primary human monocytes and the human monocytic cell line MonoMac6 were stimulated with various toll-like-receptor agonists after priming with Na-L-lactate under constant pH conditions. As readout, cytokine production was measured, real-time assessment of intracellular energy pathways was performed, and intracellular metabolite concentrations were determined. Irrespective of the immunogenic stimulus, short-term Na-lactate-priming strongly reduced cytokine production capacity. Lactate and hexoses accumulated intracellularly and, together with a decreased glycolytic flux, indicate a lactate-triggered impairment of glycolysis. To counteract intracellular hyperglycemia, glucose is shunted into the branching polyol pathway, leading to sorbitol accumulation. In contrast, long-term priming with Na-L-lactate induced cellular adaption and abolished the suppressive effect. This lactate tolerance is characterized by a decreased cellular respiration due to a reduced complex-I activity. Our results indicate that exogenous lactate shapes monocyte function by altering the intracellular energy metabolism and acts as a metabolic checkpoint of monocyte activation.

Chronic hypoxia increases the resistance of pulmonary arteries by stimulating their contraction and augmenting their coverage by smooth muscle cells (SMCs). While these responses require adjustment of the vascular SMC transcriptome, regulatory elements are not well defined in this context. Here, we explored the functional role of the transcription factor nuclear factor of activated T-cells 5 (NFAT5/TonEBP) in the hypoxic lung. Regulatory functions of NFAT5 were investigated in cultured artery SMCs and lungs from control (Nfat5fl/fl) and SMC-specific Nfat5-deficient (Nfat5(SMC)−/−) mice. Exposure to hypoxia promoted the expression of genes associated with metabolism and mitochondrial oxidative phosphorylation (OXPHOS) in Nfat5(SMC)−/− versus Nfat5fl/fl lungs. In vitro, hypoxia-exposed Nfat5-deficient pulmonary artery SMCs elevated the level of OXPHOS-related transcripts, mitochondrial respiration, and production of reactive oxygen species (ROS). Right ventricular functions were impaired while pulmonary right ventricular systolic pressure (RVSP) was amplified in hypoxia-exposed Nfat5(SMC)−/− versus Nfat5fl/fl mice. Scavenging of mitochondrial ROS normalized the raise in RVSP. Our findings suggest a critical role for NFAT5 as a suppressor of OXPHOS-associated gene expression, mitochondrial respiration, and ROS production in pulmonary artery SMCs that is vital to limit ROS-dependent arterial resistance in a hypoxic environment.

Aberrant expression of MYC family members predicts poor clinical outcome in many human cancers. Oncogenic MYC profoundly alters metabolism and mediates an antioxidant response to maintain redox balance. Here we show that MYC induces massive lipid peroxidation upon depletion of cysteine, the rate-limiting amino acid for glutathione biosynthesis and sensitizes cells to ferroptosis, an oxidative, non-apoptotic and irondependent type of cell death. In MYCN-amplified childhood neuroblastoma, MYCN mediates resistance to ferroptosis by activating transsulfuration of methionine to cysteine. MYCN may contribute to spontaneous tumor regression in low-risk neuroblastomas by promoting ferroptosis in cells with epigenetically silenced cystathionine-beta-synthase, the rate-limiting enzyme for transsulfuration. We identified enzymes and antiporter proteins crucial to ferroptotic escape, providing multiple previously unknown sites that may be acted on therapeutically.

According to Hume’s principle, a sentence of the form ⌜The number of Fs = the number of Gs⌝ is true if and only if the Fs are bijectively correlatable to the Gs. Neo-Fregeans maintain that this principle provides an implicit definition of the notion of cardinal number that vindicates a platonist construal of such numerical equations. Based on a clarification of the explanatory status of Hume’s principle, I will provide an argument in favour of a nominalist construal of numerical equations. The neo-Fregean objections to such a construal will be examined and rejected. And the implications of the nominalist construal for the use of numerals and for the understanding of ontological questions for the existence of numbers will be spelled out.

Background Hepatocellular carcinoma (HCC) is a major cause of cancer-related death. Paired related homeobox 1 (PRRX1) is a transcription factor that regulates cell growth and differentiation, but its importance in HCC is unclear. Methods We examined the expression pattern of PRRX1 in nine microarray datasets of human HCC tumour samples (n > 1100) and analyzed its function in HCC cell lines. In addition, we performed gene set enrichment, Kaplan-Meier overall survival analysis, metabolomics and functional assays. Results PRRX1 is frequently upregulated in human HCC. Pathway enrichment analysis predicted a direct correlation between PRRX1 and focal adhesion and epithelial-mesenchymal transition. High expression of PRRX1 and low ZEB1 or high ZEB2 significantly predicted better overall survival in HCC patients. In contrast, metabolic processes correlated inversely and transcriptional analyses revealed that glycolysis, TCA cycle and amino acid metabolism were affected. These findings were confirmed by metabolomics analysis. At the phenotypic level, PRRX1 knockdown accelerated proliferation and clonogenicity in HCC cell lines. Conclusions Our results suggest that PRRX1 controls metabolism, has a tumour suppressive role, and may function in cooperation with ZEB1/2. These findings have functional relevance in HCC, including in understanding transcriptional control of distinct cancer hallmarks.

Seeds preserve a far developed plant embryo in a quiescent state. Seed metabolism relies on stored resources and is reactivated to drive germination when the external conditions are favorable. Since the switchover from quiescence to reactivation provides a remarkable case of a cell physiological transition we investigated the earliest events in energy and redox metabolism of Arabidopsis seeds at imbibition. By developing fluorescent protein biosensing in intact seeds, we observed ATP accumulation and oxygen uptake within minutes, indicating rapid activation of mitochondrial respiration, which coincided with a sharp transition from an oxidizing to a more reducing thiol redox environment in the mitochondrial matrix. To identify individual operational protein thiol switches, we captured the fast release of metabolic quiescence in organello and devised quantitative iodoacetyl tandem mass tag (iodoTMT)-based thiol redox proteomics. The redox state across all Cys peptides was shifted toward reduction from 27.1% down to 13.0% oxidized thiol. A large number of Cys peptides (412) were redox switched, representing central pathways of mitochondrial energy metabolism, including the respiratory chain and each enzymatic step of the tricarboxylic acid (TCA) cycle. Active site Cys peptides of glutathione reductase 2, NADPH-thioredoxin reductase a/b, and thioredoxin-o1 showed the strongest responses. Germination of seeds lacking those redox proteins was associated with markedly enhanced respiration and deregulated TCA cycle dynamics suggesting decreased resource efficiency of energy metabolism. Germination in aged seeds was strongly impaired. We identify a global operation of thiol redox switches that is required for optimal usage of energy stores by the mitochondria to drive efficient germination.

Prostate cell metabolism exhibits distinct profiles pre- and post-malignancy. The malignant metabolic shift converts prostate cells from "citrate-producing" to "citrate-oxidizing" cells, thereby enhancing glucose metabolism, a phenotype that contrasts classical tumoral Warburg metabolism. An on-line biosensor chip system (BIONAS 2500) was used to monitor metabolic changes (glycolysis and respiration) in response to the putative anti-cancer nutraceutical 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], in different prostate cancer (PCa) cell lines (LNCaP, VCaP, DU145 and PC3). LNCaP cells exhibited profound metabolic responsiveness to the treatment and thus extensive analysis of metabolism-modulating effects of 1,25(OH)2D3 were performed, including mRNA expression analysis of key metabolic genes (e.g. GLUT1 and PDHK1), analysis of TCA cycle metabolites, glucose uptake/consumption measurements, ATP production, and mitochondrial biogenesis/activity. Altogether, data demonstrate a vivid disruption of glucose metabolism by 1,25(OH)2D3, illustrated by a decreased glucose uptake and an accumulation of citrate/isocitrate due to TCA cycle truncation. Depletion of glycolytic intermediates led to a consistent decrease in TXNIP expression in response to 1,25(OH)2D3, an effect that coincided with the activation of AMPK signaling and a reduction in c-MYC expression. Reduction in TXNIP levels in response to 1,25(OH)2D3 was rescued by an AMPK signaling inhibitor and mimicked by a MYC inhibitor highlighting the possible involvement of both pathways in mediating 1,25(OH)2D3's metabolic effects in PCa cells. Furthermore, pharmacological and genetic modulation of the androgen receptor showed similar and disparate effects on metabolic parameters compared to 1,25(OH)2D3 treatment, highlighting the AR-independent nature of 1,25(OH)2D3's metabolism-modulating effects.

The increased formation of methylglyoxal (MG) under hyperglycemia is associated with the development of microvascular complications in patients with diabetes mellitus; however, the effects of elevated MG levels in vivo are poorly understood. In zebrafish, a transient knockdown of glyoxalase 1, the main MG detoxifying system, led to the elevation of endogenous MG levels and blood vessel alterations. To evaluate effects of a permanent knockout of glyoxalase 1 in vivo, glo1(–/–) zebrafish mutants were generated using CRISPR/Cas9. In addition, a diet-induced–obesity zebrafish model was used to analyze glo1(–/–) zebrafish under high nutrient intake. Glo1(–/–) zebrafish survived until adulthood without growth deficit and showed increased tissue MG concentrations. Impaired glucose tolerance developed in adult glo1(–/–) zebrafish and was indicated by increased postprandial blood glucose levels and postprandial S6 kinase activation. Challenged by an overfeeding period, fasting blood glucose levels in glo1(–/–) zebrafish were increased which translated into retinal blood vessel alterations. Thus, the data have identified a defective MG detoxification as a metabolic prerequisite and glyoxalase 1 alterations as a genetic susceptibility to the development of type 2 diabetes mellitus under high nutrition intake.

Pyrophosphate (PPi), a byproduct of macromolecule biosynthesis is maintained at low levels by soluble inorganic pyrophosphatases (sPPase) found in all eukaryotes. In plants, H+-pumping pyrophosphatases (H+-PPase) convert the substantial energy present in PPi into an electrochemical gradient. We show here, that both cold- and heat stress sensitivity of fugu5 mutants lacking the major H+-PPase isoform AVP1 is caused by reduced SUMOylation. In addition, we show that increased PPi concentrations interfere with SUMOylation in yeast and we provide evidence that SUMO activating E1-enzymes are inhibited by micromolar concentrations of PPi in a non-competitive manner. Taken together, our results do not only provide a mechanistic explanation for the beneficial effects of AVP1 overexpression in plants but they also highlight PPi as an important integrator of metabolism and stress tolerance in eukaryotes.

The gene CNDP1 was associated with the development of diabetic nephropathy. Its enzyme carnosinase 1 (CN1) primarily hydrolyzes the histidine-containing dipeptide carnosine but other organ and metabolic functions are mainly unknown. In our study we generated CNDP1 knockout zebrafish, which showed strongly decreased CN1 activity and increased intracellular carnosine levels. Vasculature and kidneys of CNDP1-/- zebrafish were not affected, except for a transient glomerular alteration. Amino acid profiling showed a decrease of certain amino acids in CNDP1-/- zebrafish, suggesting a specific function for CN1 in the amino acid metabolisms. Indeed, we identified a CN1 activity for Ala-His and Ser-His. Under diabetic conditions increased carnosine levels in CNDP1-/- embryos could not protect from respective organ alterations. Although, weight gain through overfeeding was restrained by CNDP1 loss. Together, zebrafish exhibits CN1 functions, while CNDP1 knockout alters the amino acid metabolism, attenuates weight gain but cannot protect organs from diabetic complications.

The paper provides an interpretation of Wittgenstein’s claim that a mathematical proof must be surveyable. It will be argued that this claim specifies a precondition for the applicability of the word ‘proof’. Accordingly, the latter is applicable to a proof-pattern only if we can come to agree by mere observation whether or not the pattern possesses the relevant structural features. The claim is problematic. It does not imply any questionable finitist doctrine. But it cannot be said to articulate a feature of our actual usage of the word ‘proof’. The claim can be dissociated, however, from two tenable doctrines of Wittgenstein, namely that proofs can be used as paradigms for corresponding proof concepts and that a proof is not an experiment.

The author critically examines Wittgenstein’s and Waismann’s objections to the explanation of equinumerosity in terms of one-one correlations. They are right to maintain that the actuality of a one-one correlation is not a necessary condition for equinumerosity. But, contrary to what they claim, the possibility of such a correlation is equivalent with equinumerosity. The author shows how this equivalence can be used to explain the use of ‘just as many’, thereby also responding to Husserl’s objection that this phrase is indefinable. And against all three authors it is argued that there are no semantically relevant equinumerosity criteria besides one-one correlatability.

Fine bases his influential conception of essence on a particular account of definitions. And he complements it with a specific account of analyticity. I will argue that Fine's conception of relative analyticity confuses the idea of a sentence's being true in virtue of a term's definition with the idea of a sentence's being true in virtue of a term's meaning. His idea that correct definitions specify essential properties of meanings is mistaken. The correctness of definitions can only be assessed by reference to the actual usage of the terms involved. The resulting conception of definitions leads to a deflationary interpretation of claims about essences.

The molecular structure of matter defines its properties and function. This is especially true for biological macromolecules such as proteins, which participate in virtually all biochemical processes. A three dimensional structural model of a protein is thus essential for the detailed understanding of its physiological function and the characterization of essential properties such as ligand binding and reaction mechanism. X-ray crystallography is a well-established technique that has been used for many years, but it is still by far the most widely used method for structure determination. A particular strength of this technique is the elucidation of atomic details of molecular interactions, thus providing an invaluable tool for a multitude of scientific projects ranging from the structural classification of macromolecules over the validation of enzymatic mechanisms or the understanding of host–pathogen interactions to structure-guided drug design. In the first part of this review, we describe essential methodological and practical aspects of X-ray crystallography. We provide some pointers that should allow researchers without a background in structural biology to assess the overall quality and reliability of a crystal structure. To highlight its potential, we then survey the impact X-ray crystallography has had on advancing an understanding of a class of enzymes that modify the bacterial cell wall. A substantial number of different bacterial amidase structures have been solved, mostly by X-ray crystallography. Comparison of these structures highlights conserved as well as divergent features. In combination with functional analyses, structural information on these enzymes has therefore proven to be a valuable template not only for understanding their mechanism of catalysis, but also for targeted interference with substrate binding.

Orbital blunt trauma is common, and the diagnosis of a fracture should be made by computed tomographic (CT) scan. However, this will expose patients to ionising radiation. Our objective was to identify clinical predictors of orbital fracture, in particular the presence of a black eye, to minimise unnecessary exposure to radiation. A 10-year retrospective study was made of the medical records of all patients with minor head trauma who presented with one or two black eyes to our emergency department between May 2000 and April 2010. Each of the patients had a CT scan, was over 16 years old, and had a Glasgow Coma Score (GCS) of 13-15. The primary outcome was whether the black eye was a valuable predictor of a fracture. Accompanying clinical signs were considered as a secondary outcome. A total of 1676 patients (mean (SD) age 51 (22) years) and minor head trauma with either one or two black eyes were included. In 1144 the CT scan showed a fracture of the maxillofacial skeleton, which gave an incidence of 68.3% in whom a black eye was the obvious symptom. Specificity for facial fractures was particularly high for other clinical signs, such as diminished skin sensation (specificity 96.4%), diplopia or occulomotility disorders (89.3%), fracture steps (99.8%), epistaxis (95.5%), subconjunctival haemorrhage (90.4%), and emphysema (99.6%). Sensitivity for the same signs ranged from 10.8% to 22.2%. The most striking fact was that 68.3% of all patients with a black eye had an underlying fracture. We therefore conclude that a CT scan should be recommended for every patient with minor head injury who presents with a black eye.

The purpose of this article is to describe a technique for secondary reconstruction of traumatic orbital wall defects using titanium implants that act as three-dimensional (3D) puzzle pieces. We present three cases of large defect reconstruction using implants produced by Xilloc Medical B.V. (Maastricht, the Netherlands) with a 3D printer manufactured by LayerWise (3D Systems; Heverlee, Belgium), and designed using the biomedical engineering software programs ProPlan and 3-Matic (Materialise, Heverlee, Belgium). The smaller size of the implants allowed sequential implantation for the reconstruction of extensive two-wall defects via a limited transconjunctival incision. The precise fit of the implants with regard to the surrounding ledges and each other was confirmed by intraoperative 3D imaging (Mobile C-arm Systems B.V. Pulsera, Philips Medical Systems, Eindhoven, the Netherlands). The patients showed near-complete restoration of orbital volume and ocular motility. However, challenges remain, including traumatic fat atrophy and fibrosis.

Subcellular sugar partitioning in plants is strongly regulated in response to developmental cues and changes in external conditions. Besides transitory starch, the vacuolar sugars represent a highly dynamic pool of instantly accessible metabolites that serve as energy source and osmoprotectant. Here, we present the molecular identification and functional characterization of the vacuolar glucose (Glc) exporter Arabidopsis (Arabidopsis thaliana) Early Responsive to Dehydration-Like6 (AtERDL6). We demonstrate tonoplast localization of AtERDL6 in plants. In Arabidopsis, AtERDL6 expression is induced in response to factors that activate vacuolar Glc pools, like darkness, heat stress, and wounding. On the other hand, AtERDL6 transcript levels drop during conditions that trigger Glc accumulation in the vacuole, like cold stress and external sugar supply. Accordingly, sugar analyses revealed that Aterdl6 mutants have elevated vacuolar Glc levels and that Glc flux across the tonoplast is impaired under stress conditions. Interestingly, overexpressor lines indicated a very similar function for the ERDL6 ortholog Integral Membrane Protein from sugar beet (Beta vulgaris). Aterdl6 mutant plants display increased sensitivity against external Glc, and mutant seeds exhibit a 10% increase in seed weight due to enhanced levels of seed sugars, proteins, and lipids. Our findings underline the importance of vacuolar Glc export during the regulation of cellular Glc homeostasis and the composition of seed reserves.

Graphite has been subjected to surface damage by Ar+ ion bombardment. It has been found by scanning tunneling microscopy (STM) that deposited Cs atoms preferentially cluster at the artificially-produced vacancy clusters in the basal plane at 300 K. Density functional theory calculations show that Cs is more strongly bound at these defect sites than at basal plane step edges, providing a plausible explanation for the experimental observation. STM reveals that Cs intercalation into graphite occurs by diffusion through these vacancy clusters, acting as entry ports to the interior, at 700 K. DFT calculations show that these defects must consist of more than four contiguous atoms in order for Cs intercalation to be energetically easy. Experimentally, Cs atom nucleation at defect sites seems to culminate at Cs cluster heights near 1 nm and cluster diameters near 5 nm. Intercalated Cs produces coherent single Cs islands in the galleries as well as layered structures caused by superposition of overlapping individual islands. Oxygen exposure to graphite containing Ar+-produced defects does not influence Cs clustering at the defects or Cs entry into galleries beneath the defect. However, large oxygen exposures these clusters results in diminution of Cs intercalation, probably due to surface oxidation of Cs clusters.

The vacuolar membrane is involved in solute uptake into and release from the vacuole, which is the largest plant organelle. In addition to inorganic ions and metabolites, large quantities of protons and sugars are shuttled across this membrane. Current models suggest that the proton gradient across the membrane drives the accumulation and/or release of sugars. Recent studies have associated AtSUC4 with the vacuolar membrane. Some members of the SUC family are plasma membrane proton/sucrose symporters. In addition, the sugar transporters TMT1 and TMT2, which are localized to the vacuolar membrane, have been suggested to function in proton-driven glucose antiport. Here we used the patch-clamp technique to monitor carrier-mediated sucrose transport by AtSUC4 and AtTMTs in intact Arabidopsis thaliana mesophyll vacuoles. In the whole-vacuole configuration with wild-type material, cytosolic sucrose-induced proton currents were associated with a proton/sucrose antiport mechanism. To identify the related transporter on one hand, and to enable the recording of symporter-mediated currents on the other hand, we electrophysiologically characterized vacuolar proteins recognized by Arabidopsis mutants of partially impaired sugar compartmentation. To our surprise, the intrinsic sucrose/proton antiporter activity was greatly reduced when vacuoles were isolated from plants lacking the monosaccharide transporter AtTMT1/TMT2. Transient expression of AtSUC4 in this mutant background resulted in proton/sucrose symport activity. From these studies, we conclude that, in the natural environment within the Arabidopsis cell, AtSUC4 most likely catalyses proton-coupled sucrose export from the vacuole. However, TMT1/2 probably represents a proton-coupled antiporter capable of high-capacity loading of glucose and sucrose into the vacuole.

To understand carbon partitioning in roots and nodules of Datisca glomerata, activities of sucrose-degrading enzymes and sugar transporter expression patterns were analyzed in both organs, and plasmodesmal connections between nodule cortical cells were examined by transmission electron microscopy. The results indicate that in nodules, the contribution of symplastic transport processes is increased in comparison to roots, specifically in infected cells which develop many secondary plasmodesmata. Invertase activities are dramatically reduced in nodules as compared to roots, indicating that here the main enzyme responsible for the cleavage of sucrose is sucrose synthase. A high-affinity, low-specificity monosaccharide transporter whose expression is induced in infected cells prior to the onset of bacterial nitrogen fixation, and which has an unusually low pH optimum and may be involved in turgor control or hexose retrieval during infection thread growth. Electronic supplementary material The online version of this article (doi:10.1007/s00425-010-1285-8) contains supplementary material, which is available to authorized users.

The Arabidopsis sugar transporter (AtSTP) family is one of the best characterised families within the monosaccharide transporter (MST)-like genes. However, several aspects are still poorly investigated or not yet addressed experimentally, such as post-translational modifications and other factors affecting transport activity. This mini-review summarises recent advances in the AtSTP family as well as objectives for future studies.

The doping of single-walled carbon nanotubes (SWNTs) under ultrahigh vacuum by Li atoms has been explored experimentally and theoretically. The chemical effect of Li in breaking the C−Cl bond in chloromethane has been observed. Temperature programmed desorption (TPD) experiments show that at low coverage CH3Cl is physisorbed to the undoped SWNT sample, exhibiting a desorption process near 178 K. The CH3Cl desorption peak shifts to about 240 K for lithiated SWNTs, indicating an increase in binding energy of about 0.16 eV. More importantly, the integrated intensity of the CH3Cl desorption peak is dramatically reduced in the lithiated SWNT case, and CH3, C2H6, or related species are not observed in significant quantities in the TPD experiments up to a temperature of 500 K. This strongly indicates that CH3Cl reacts on lithiated SWNTs to produce an irreversibly bound species. Products LiCl and Li2Cl2 are observed to desorb near 700 K. Density functional theory calculations present possible reaction mechanisms ...