Adenosine Triphosphate Metabolism Research Articles

Endothelial P2X7 promotes venous thromboembolism

Venous thromboembolism (VTE) affects 117 people per 100,000 each year and is an important cause of morbidity and mortality. VTE can lead to 1) death through pulmonary embolism, 2) the post-thrombotic syndrome, or 3) chronic pulmonary hypertension resulting in significant chronic respiratory compromise. Inflammatory pathways are intricately involved in thrombus formation and therefore in VTE mechanisms. In pathological conditions, adenosine triphosphate (ATP) is released in the extracellular compartment and is recognized as a danger signal by several cell types including endothelial cells. Recent data indicated that the CD39/CD73 system involved in the metabolism of ATP into AMP might protects against thrombosis by downregulating the pro-inflammatory pathway of the inflammasome. ATP can also interact with the P2X7 receptor involved in inflammation causing a wide range of responses. To determine how the endothelial P2X7 receptor contributes to venous thromboembolism. HUVECs were incubated with BzATP, thrombin or both. HUVECs were primed with TNF-alpha prior to stimulation. We used immunofluorescence, western blot and real-time quantitative PCR analyses to assess P2X7 expression by endothelial cells, activation of p38 and NFκB activation and expression of pro-inflammatory and pro-coagulant genes. We confirmed that endothelial cells expressed P2X7 in vitro in HUVECs and in vivo after induction of venous thrombosis by ligation of the inferior vena cava. Treatment of endothelial cells with BzATP and thrombin induced the activation of p38 and NFκB signaling pathways. This appears to be associated with an increased expression of IL-1β and downregulation of thrombomodulin expression. The expression of adhesion molecules, ICAM-1 and VCAM-1, tends to increase. Our data suggest that ATP released in the extracellular space following cell damage or activation induced a pro-inflammatory response in endothelial cells through P2X7 activation. P2X7 might have a pro-thrombotic role exacerbating venous thromboembolism.

The functional activity of donor kidneys is negatively regulated by microribonucleic acid-451 in different perfusion methods to inhibit adenosine triphosphate metabolism and the proliferation of HK2 cells

ABSTRACT This study explored the regulation of different perfusion methods on ischemia-reperfusion injury in donor kidneys. In this study, renal cortical/medullary tissue specimens were collected from porcine kidneys donors using different perfusion methods at various time points. Hematoxylin and eosin (H&E) staining was used to test the histological differences. Differentially expressed micro-ribonucleic acids (miRNAs) were identified by miRNA transcriptome sequencing. Reverse transcription-polymerase chain reaction (RT-PCR) tests were used to verify the changes in miRNAs in the kidney tissue taken from different perfusion groups. The related signaling pathways and the changes in the cell functions of different perfusion groups were analyzed by Kyoto Encyclopedia of Genes and Genomes (KEGG) /Gene Ontology (GO) bioinformatics analyses. The effects of miRNA overexpression on the metabolism and proliferation of HK2 cells were detected by ATP kit and MTT assay. The H&E staining results showed that there were essentially no differences in the tissue samples among different perfusion groups at and before 12 h compared with a control group. The quantitative PCR results revealed that there was essentially no change in the expression of ssc-miR-451, ssc-miR-1285, and ssc-miR-486 in the cis infusion or joint infusion kidney groups, and their expression was significantly down-regulated over time in the trans-infusion kidney group. The bioinformatics analysis showed that the cellular component, molecular function, and biological processes of the kidney tissue, which had been perfused using three methods, had been consistently affected. The most significant changes after perfusion occurred in the intracellular metabolism signaling pathways. Furthermore, the energy metabolism and proliferation of the HK2 cells were significantly inhibited after the overexpression of miR-451. Specific miRNA markers, such as miR-451, may play a negative regulatory role in cell metabolism following the perfusion of kidney transplants using different methods.

Endothelial P2X7 promotes venous thromboembolism

Venous thromboembolism (VTE) afflicts 117 people per 100,000 each year and is an important cause of morbidity and mortality. VTE can lead to 1) death through pulmonary embolism, 2) the post-thrombotic syndrome, or 3) chronic pulmonary hypertension resulting in significant chronic respiratory compromise. Inflammatory pathways are intricately involved in venous thromboembolism mechanisms. In pathological conditions, adenosine triphosphate (ATP) is released in the extracellular compartment and is recognized as a danger signal. Recent data indicated that the CD39/CD73 system involved in the metabolism of ATP into AMP might protect against thrombosis by downregulating the pro-inflammatory pathway of the inflammasome. ATP can also interact with the P2X7 receptor involved in inflammation causing a wide range of responses. Determining how the endothelial P2X7 receptor contribute to venous thromboembolism. HUVEC were incubated with BzATP, thrombin or both. In some experiment, HUVEC were primed with TNFaprior to stimulation. We used immunofluorescence, western blot and real-time quantitative PCR analyses to assess P2X7 expression by endothelial cells, activation of p38 and NFkB activation and expression of pro-inflammatory and pro- coagulant genes. We confirmed that endothelial cells expressed P2X7 in vitro in HUVECs and in vivo after induction of venous thrombosis by ligation of the inferior vena cava. Treatment of endothelial cells with BzATP and thrombin induced the activation of p38 and NFkB signaling pathways. This was associated with an increased expression of IL-1b and tissue factor and downregulation of thrombomodulin expression. Our data suggest that ATP released in the extracellular space following cell damage or activation induced the inflammasome in endothelial through P2X7 activation. P2X7 might have a pro-thrombotic role exacerbating venous thromboembolism.

Adenosinergic Signaling as a Key Modulator of the Glioma Microenvironment and Reactive Astrocytes.

Astrocytes are numerous glial cells of the central nervous system (CNS) and play important roles in brain homeostasis. These cells can directly communicate with neurons by releasing gliotransmitters, such as adenosine triphosphate (ATP) and glutamate, into the multipartite synapse. Moreover, astrocytes respond to tissue injury in the CNS environment. Recently, astrocytic heterogeneity and plasticity have been discussed by several authors, with studies proposing a spectrum of astrocytic activation characterized by A1/neurotoxic and A2/neuroprotective polarization extremes. The fundamental roles of astrocytes in communicating with other cells and sustaining homeostasis are regulated by purinergic signaling. In the CNS environment, the gliotransmitter ATP acts cooperatively with other glial signaling molecules, such as cytokines, which may impact CNS functions by facilitating/inhibiting neurotransmitter release. Adenosine (ADO), the main product of extracellular ATP metabolism, is an important homeostatic modulator and acts as a neuromodulator in synaptic transmission via P1 receptor sensitization. Furthermore, purinergic signaling is a key factor in the tumor microenvironment (TME), as damaged cells release ATP, leading to ADO accumulation in the TME through the ectonucleotidase cascade. Indeed, the enzyme CD73, which converts AMP to ADO, is overexpressed in glioblastoma cells; this upregulation is associated with tumor aggressiveness. Because of the crucial activity of CD73 in these cells, extracellular ADO accumulation in the TME contributes to sustaining glioblastoma immune escape while promoting A2-like activation. The present review describes the importance of ADO in modulating astrocyte polarization and simultaneously promoting tumor growth. We also discuss whether targeting of CD73 to block ADO production can be used as an alternative cancer therapy.

Study on community structure and metabolic mechanism of dominant polyphosphate-accumulating organisms (PAOs) and glycogen-accumulating organisms (GAOs) in suspended biofilm based on phosphate recovery

Biofilm sequencing batch reactor (BSBR) can achieve efficient phosphate (P) removal and enrichment, but its process performance and metabolic mechanisms for P removal and enrichment of municipal wastewater remain largely unclear. In the present study, we assessed the P removal and enrichment of municipal wastewater at influent P concentrations of 2.5 mg/L and 10 mg/L. The efficiency of P removal and enzyme activity in polyphosphate-accumulating organisms (PAOs) and glycogen-accumulating organisms (GAOs) were compared, and the growth and metabolic characteristics of dominant PAOs and GAOs at different influent P concentrations were studied with the macro-sequencing technology. The results showed that the P recovery efficiencies were 70.03% and 76.19% when the influent P concentration was 2.5 mg/L and 10 mg/L in BSBR, respectively, and the maximum P concentration of recovery liquid was 81.29 mg/L and 173.12 mg/L, respectively. There were no phosphate kinase (PPK) and phosphate hydrolase (PPX) in extracellular polymeric substances (EPS). The dominant PAOs were Candidatus_Contendobacter, Dechloromonas, and Flavobacterium, and the dominant GAO was Candidatus_Competibacter. The abundance of Candidatus_Contendobacter was the highest with the most potential contribution to P removal. PAOs had competitive advantages in carbon (C) source uptake, glycogen metabolism, P metabolism, and adenosine triphosphate (ATP) metabolism. HMP was unique to PAOs, EMP had the highest abundance in glycogen metabolism, and ED was contained in PAOs of BSBR. These results indicated that BSBR provided sufficient reducing power and ATP for PAOs through different glycogen decomposition pathways to promote P uptake and obtained competitive advantages in P metabolism, C source uptake, and ATP utilization to achieve efficient P removal and enrichment. Collectively, our current findings provided valuable insights into the P removal and enrichment mechanism of BSBR in municipal sewage.

A Dual-Control Strategy by Phosphate Ions and Local Microviscosity for Tracking Adenosine Triphosphate Metabolism in Mitochondria and Cellular Activity Dynamically.

Adenosine triphosphate (ATP) acts as the main energy source for growth and development in organisms, and the disorder reflects the mitochondrial damage to a large extent. Therefore, an efficient tool for the evaluation of the ATP metabolic level is important to track mitochondrial health, providing an additional perspective for an in-depth long-term study on living activities. Herein, a twisted intramolecular charge transfer (TICT) framework is utilized to build up a sensitive receptor, Mito-VP, with a negligible background to target mitochondrial ATP metabolism by monitoring the phosphate ion (Pi) level upon ATP hydrolysis under the overall consideration of the structural and functional features of mitochondria. The responsive fluorescence could be lighted on under the dual control of Pi and local microviscosity, and the two steps of ATP hydrolysis could be captured through fluorescence. In addition to the well-behaved mitochondrial targeting, the energy metabolism at cellular and organism levels has been clarified via mitosis and zebrafish development, respectively.

Mitochondrial Heteroplasmy Shifting as a Potential Biomarker of Cancer Progression.

Cancer is a serious health problem with a high mortality rate worldwide. Given the relevance of mitochondria in numerous physiological and pathological mechanisms, such as adenosine triphosphate (ATP) synthesis, apoptosis, metabolism, cancer progression and drug resistance, mitochondrial genome (mtDNA) analysis has become of great interest in the study of human diseases, including cancer. To date, a high number of variants and mutations have been identified in different types of tumors, which coexist with normal alleles, a phenomenon named heteroplasmy. This mechanism is considered an intermediate state between the fixation or elimination of the acquired mutations. It is suggested that mutations, which confer adaptive advantages to tumor growth and invasion, are enriched in malignant cells. Notably, many recent studies have reported a heteroplasmy-shifting phenomenon as a potential shaper in tumor progression and treatment response, and we suggest that each cancer type also has a unique mitochondrial heteroplasmy-shifting profile. So far, a plethora of data evidencing correlations among heteroplasmy and cancer-related phenotypes are available, but still, not authentic demonstrations, and whether the heteroplasmy or the variation in mtDNA copy number (mtCNV) in cancer are cause or consequence remained unknown. Further studies are needed to support these findings and decipher their clinical implications and impact in the field of drug discovery aimed at treating human cancer.

Endothelial P2X7 promotes venous thromboembolism

Venous thromboembolism (VTE) afflicts 117 people per 100,000 each year and is an important cause of morbidity and mortality. VTE can lead to: – death through pulmonary embolism; – the post-thrombotic syndrome, or; – chronic pulmonary hypertension resulting in significant chronic respiratory compromise. Inflammatory pathways are intricately involved in venous thromboembolism mechanisms. In pathological conditions, adenosine triphosphate (ATP) is released in the extracellular compartment and is recognized as a danger signal. Recent data indicated that the CD39/CD73 system involved in the metabolism of ATP into AMP might protect against thrombosis by downregulating the pro-inflammatory pathway of the inflammasome. ATP can also interact with the P2X7 receptor involved in inflammation causing a wide range of responses. Determine how the endothelial P2X7 receptor contribute to venous thromboembolism. HUVEC were incubated with BzATP, thrombin or both. In some experiment, HUVEC were primed with TNFa prior to stimulation. We used immunofluorescence, western blot and real-time quantitative PCR analyses to assess P2X7 expression by endothelial cells, activation of p38 and NFkB activation and expression of pro-inflammatory and procoagulant genes. We confirmed that endothelial cells expressed P2X7 in vitro in HUVECs and in vivo after induction of venous thrombosis by ligation of the inferior vena cava. Treatment of endothelial cells with BzATP and thrombin induced the activation of p38 and NFkB signaling pathways. This was associated with an increased expression of IL-1b and tissue factor and downregulation of thrombomodulin expression. Our data suggest that ATP released in the extracellular space following cell damage or activation induced the inflammasome in endothelial through P2X7 activation. P2X7 might have a pro-thrombotic role exacerbating venous thromboembolism.

Hypoxia-induced miR-27 and miR-195 regulate ATP consumption, viability, and metabolism of rat cardiomyocytes by targeting PPARγ and FASN expression.

This study examined whether hypoxia-induced microRNA (miRNA) upregulation was related to the inhibition of chondriosome aliphatic acid oxidation in myocardial cells under anoxia. We showed that anoxia induced high expression of hypoxia-inducible factor-1-alpha, muscle carnitine palmitoyltransferase I, and vascular endothelial growth factor in cardiomyocytes. Meanwhile, miR-27 and miR-195 were also upregulated in hypoxia-induced cardiomyocytes. Furthermore, hypoxia induction led to reductions in the adenosine triphosphate (ATP) consumption rate and oxidative metabolism as well as an increase in cardiomyocyte glycolysis. Metabolic reprogramming was reduced by hypoxia, as evidenced by the downregulation of sirtuin 1, forkhead box protein O1, sterol regulatory element-binding protein 1c, ATP citrate lyase, acetyl-coenzyme A carboxylase 2, adiponutrin, adipose triglyceride lipase, and glucose transporter type 4, while miR-27 and miR-195 inhibition partially recovered the expression of these transcription factors. In addition, hypoxia induction reduced cell viability and survival by triggering apoptosis; however, miR-27 and miR-195 inhibition partially increased cell viability. Moreover, miR-27 and miR-195 targeted the 3’untranslated regions of two key lipid-associated metabolic players, peroxisome proliferator-activated receptor gamma and fatty acid synthase. In conclusion, miR-27 and miR-195 are related to hypoxia-mediated ATP levels, glycolysis, oxidation, cell survival, and a cascade of transcription factors that control metabolism in cardiomyocytes.

Utilizing Pork Exudate Metabolomics to Reveal the Impact of Aging on Meat Quality.

This study was performed to assess the changes in meat quality and metabolome profiles of meat exudate during postmortem aging. At 24 h postmortem, longissimus lumborum muscles were collected from 10 pork carcasses, cut into three sections, and randomly assigned to three aging period groups (2, 9, and 16 d). Meat quality and chemical analyses, along with the metabolomics of meat exudates using ultra-high-performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometer (UHPLC-QTOF-MS) platform, were conducted. Results indicated a declined (p < 0.05) display color stability, and increased (p < 0.05) purge loss, meat tenderness, and lipid oxidation as aging extended. The principal component analysis and hierarchical clustering analysis exhibited distinct clusters of the exudate metabolome of each aging treatment. A total of 39 significantly changed features were tentatively identified via matching them to METLIN database according to their MS/MS information. Some of those features are associated with adenosine triphosphate metabolism (creatine and hypoxanthine), antioxidation (oxidized glutathione and carnosine), and proteolysis (dipeptides and tripeptides). The findings provide valuable information that reflects the meat quality’s attributes and could be used as a source of potential biomarkers for predicting aging times and meat quality changes.

Effects of notoginseng leaf triterpenes on small molecule metabolism after cerebral ischemia/reperfusion injury assessed using MALDI-MS imaging.

BackgroundNotoginseng leaf triterpenes (PNGL) is believed to have neuroprotective effects via the inhibition of inflammatory response and neuronal apoptosis. However, its mechanisms underlying the anti-ischemia/reperfusion (I/R) injury effects on the regulation of small molecule metabolism in rat brain remains unclear. The purpose of this study was thus to explore the mechanisms of PNGL on the regulation of small molecule metabolism in rat brain after I/R injury using matrix-assisted laser desorption ionization-mass spectrometry imaging (MALDI-MSI).MethodsAs a model of in vivo cerebral I/R injury, male Sprague-Dawley (SD) rats were established with a middle cerebral artery occlusion/reperfusion (MCAO/R) model after PNGL administration with 40 mg·kg-1 through intraperitoneal injection (i.p.) for 7 days. We assessed the neurological behavior, regional cerebral blood flow (r CBF), neuron injury, and spatial distribution of metabolic small molecules.ResultsOur in vivo results suggested that PNGL increased cerebral blood flow and relieved neurological dysfunction. Furthermore, using MALDI-MSI, we demonstrated that PNGL regulated 16 endogenous small molecules implicated in metabolic networks including tricarboxylic acid (TCA) cycle, adenosine triphosphate (ATP) metabolism, malate-aspartate shuttle, metal ions, and antioxidants underwent noticeable changes after reperfusion for 24 h.ConclusionsPNGL is a novel cerebrovascular agent that can improve cerebral blood flow and attenuate adverse neurological disorders. The mechanisms are closely correlated with relative metabolic pathways, which offers insight into exploring new mechanisms in PNGL for the treatment of cerebral I/R injury.

Variability of mitochondrial energy balance across brain regions.

Brain is not homogenous and neurons from various brain regions are known to have different vulnerabilities to mitochondrial mutations and mitochondrial toxins. However, it is not clear if this vulnerability is connected to different energy metabolism in specific brain regions. Here, using live-cell imaging, we compared mitochondrial membrane potential and nicotinamide adenine dinucleotide (NADH) redox balance in acute rat brain slices in different brain regions and further detailed the mitochondrial metabolism in primary neurons and astrocytes from rat cortex, midbrain and cerebellum. We have found that mitochondrial membrane potential is higher in brain slices from the hippocampus and brain stem. In primary co-cultures, mitochondrial membrane potential in astrocytes was lower than in neurons, whereas in midbrain cells it was higher than in cortex and cerebellum. The rate of NADH production and mitochondrial NADH pool were highest in acute slices from midbrain and midbrain primary neurons and astrocytes. Although the level of adenosine tri phosphate (ATP) was similar among primary neurons and astrocytes from cortex, midbrain and cerebellum, the rate of ATP consumption was highest in midbrain cells that lead to faster neuronal and astrocytic collapse in response to inhibitors of ATP production. Thus, midbrain neurons and astrocytes have a higher metabolic rate and ATP consumption that makes them more vulnerable to energy deprivation.

Breast Cancer Subtypes Underlying EMT-Mediated Catabolic Metabolism.

Efficient catabolic metabolism of adenosine triphosphate (ATP) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) is essentially required for cancer cell survival, especially in metastatic cancer progression. Epithelial–mesenchymal transition (EMT) plays an important role in metabolic rewiring of cancer cells as well as in phenotypic conversion and therapeutic resistance. Snail (SNAI1), a well-known inducer of cancer EMT, is critical in providing ATP and NADPH via suppression of several gatekeeper genes involving catabolic metabolism, such as phosphofructokinase 1 (PFK1), fructose-1,6-bisphosphatase 1 (FBP1), and acetyl-CoA carboxylase 2 (ACC2). Paradoxically, PFK1 and FBP1 are counter-opposing and rate-limiting reaction enzymes of glycolysis and gluconeogenesis, respectively. In this study, we report a distinct metabolic circuit of catabolic metabolism in breast cancer subtypes. Interestingly, PFKP and FBP1 are inversely correlated in clinical samples, indicating different metabolic subsets of breast cancer. The luminal types of breast cancer consist of the pentose phosphate pathway (PPP) subset by suppression of PFKP while the basal-like subtype (also known as triple negative breast cancer, TNBC) mainly utilizes glycolysis and mitochondrial fatty acid oxidation (FAO) by loss of FBP1 and ACC2. Notably, PPP remains active via upregulation of TIGAR in the FBP1-loss basal-like subset, indicating the importance of PPP in catabolic cancer metabolism. These results indicate different catabolic metabolic circuits and thus therapeutic strategies in breast cancer subsets.

Acute exposure to environmentally relevant concentrations of copper affects branchial and hepatic phosphoryl transfer network of Cichlasoma amazonarum: Impacts on bioenergetics homeostasis

The toxic effects of copper (Cu) are linked to dysfunction of metabolism and depletion of adenosine triphosphate (ATP). Nevertheless, the effects related to phosphoryl transfer network, a network of enzymes to precise coupling of the ATP-production and ATP-consuming process for maintenance of bioenergetic, remain unknown. Therefore, the aim of this study was to determine whether the phosphoryl transfer network could be one pathway involved in the bioenergetic imbalance of Cichlasoma amazonarum exposed for 96 h to environmentally relevant concentrations of Cu found in Amazonia water around mines. Branchial mitochondrial creatine kinase (CK) activity was significantly lower in fish exposed to 1500 μg/L Cu than in the control group, while branchial cytosolic CK activity was significantly greater. Branchial (exposed to 750 and 1500 μg/L Cu) and hepatic (exposed to 1500 μg/L Cu) pyruvate kinase (PK) activity was significantly lower in fish exposed to Cu than in the control group. Branchial and hepatic ATP levels were significantly lower in fish exposed to 1500 μg/L than in the control group. Branchial reactive oxygen species (ROS) and lipid peroxidation (LPO) levels were significantly higher in fish exposed to 750 and 1500 μg/L Cu compared to control. Hepatic ROS and LPO levels were significantly higher in fish exposed to 1500 μg/L than in the control group. Branchial and hepatic Cu levels were significantly higher in fish exposed to 1500 μg/L compared to other groups. Exposure to 750 and 1500 μg/L Cu impairs bioenergetics homeostasis, which appears to be mediated by ROS overproduction and lipid peroxidation.

Tau Contributes to Sevoflurane-induced Neurocognitive Impairment in Neonatal Mice.

Sevoflurane anesthesia induces Tau phosphorylation and cognitive impairment in neonatal but not in adult mice. This study tested the hypothesis that differences in brain Tau amounts and in the activity of mitochondria-adenosine triphosphate (ATP)-Nuak1-Tau cascade between the neonatal and adult mice contribute to the age-dependent effects of sevoflurane on cognitive function. 6- and 60-day-old mice of both sexes received anesthesia with 3% sevoflurane for 2 h daily for 3 days. Biochemical methods were used to measure amounts of Tau, phosphorylated Tau, Nuak1, ATP concentrations, and mitochondrial metabolism in the cerebral cortex and hippocampus. The Morris water maze test was used to evaluate cognitive function in the neonatal and adult mice. Under baseline conditions and compared with 60-day-old mice, 6-day-old mice had higher amounts of Tau (2.6 ± 0.4 [arbitrary units, mean ± SD] vs. 1.3 ± 0.2; P < 0.001), Tau oligomer (0.3 ± 0.1 vs. 0.1 ± 0.1; P = 0.008), and Nuak1 (0.9 ± 0.3 vs. 0.3 ± 0.1; P = 0.025) but lesser amounts of ATP (0.8 ± 0.1 vs. 1.5 ± 0.1; P < 0.001) and mitochondrial metabolism (74.8 ± 14.1 [pmol/min] vs. 169.6 ± 15.3; P < 0.001) in the cerebral cortex. Compared with baseline conditions, sevoflurane anesthesia induced Tau phosphorylation at its serine 202/threonine 205 residues (1.1 ± 0.4 vs. 0.2 ± 0.1; P < 0.001) in the 6-day-old mice but not in the 60-day-old mice (0.05 ± 0.04 vs. 0.03 ± 0.01; P = 0.186). The sevoflurane-induced Tau phosphorylation and cognitive impairment in the neonatal mice were both attenuated by the inhibition of Nuak1 and the treatment of vitamin K2. Higher brain Tau concentrations and lower brain mitochondrial metabolism in neonatal compared with adult mice contribute to developmental stage-dependent cognitive dysfunction after sevoflurane anesthesia.

Purine Metabolism in Platelets and Heart Cells of Hyperlipidemic Rats.

Hyperlipidemia, characterized by an increase in circulating lipid levels, doubles the chance of developing cardiovascular diseases. It prompts inflammation, immune activation, and oxidative stress in the bloodstream and organs of rats. Thus, we theorized that the metabolism of purines, an immunomodulatory mechanism, is altered in cells involved in the development of cardiovascular diseases. Therefore, we induced acute hyperlipidemia in Wistar rats with Poloxamer-407 and euthanized the animals 36h later. The leucocyte differential, the rate of purine metabolism on the surface of platelets and heart cells, and markers of oxidative stress in the heart tissue were evaluated. These parameters were also assessed in animals pretreated for 30days with curcumin and/or rutin. Hyperlipidemia increased the hydrolyses of adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) in platelets. In heart cells, the metabolism of ATP and adenosine (ADO) were increased, while ADP hydrolysis was reduced. Additionally, lipid damage and antioxidant defenses were increased in heart homogenates. Hyperlipidemic rats alsoexhibited a reduced percentage of eosinophils and lymphocytes. Together, these findings are indicative of an increased risk of developing cardiovascular diseases in hyperlipidemic rats. The pretreatments with antioxidants reverted some of the changes prompted by hyperlipidemia preventing detrimental changes in the cells and tissues. Graphical Abstract.

Role of SUMOylation in Ethanol‐Modulated Mitochondria Homeostasis

ProposeAlcohol‐induced liver disease (ALD) is a major health concern of alcohol abuse and a leading cause of liver‐related morbidity and mortality. Alterations in mitochondrial morphology and function are a hallmark of ALD. Mitochondria synthesize most of the adenosine triphosphate (ATP) needed by mammalian cells and are intimately involved in the generation of and defense against reactive oxygen species (ROS). Evidence support the importance of ROS production and oxidative stress in the development of ALD. SUMOylation is a dynamic and reversible posttranscriptional modification involved in many cellular pathways including transcription, intracellular transport, DNA repair, replication, and cell signaling. We recently demonstrated that UBC9, the sole E2 protein required by SUMOylation machinery, is upregulated in Intragastric ethanol‐fed mouse model (IE) and cirrhotic tissues. We also found that UBC9 is phosphorylated and this is correlated with high level of SUMOylation in lipopolysaccharides‐activated Kupffer cells that lead inflammation development. In addition, we described a novel key role of SUMOylation inducing the enzymatic activity and protein stability of Cytochrome P450 2E1 (CYP2E1), that is the major component of the microsomal ethanol‐oxidizing system (MEOS), in NIAAA mouse model. This study aims to investigate whether ethanol induces mitochondrial dysfunction in ALD regulating the SUMOylation status of key respiratory chain proteins and explore the molecular mechanisms.MethodsPrimary mouse hepatocytes and livers from NIAAA mouse model were used. mRNA and protein levels were analyzed by Real‐Time PCR and Western Blotting, respectively. OCR (oxygen consumption rate) and Δψm (mitochondrial membrane potential) were measured by Seahorse mito stress test and JC‐1 staining by confocal microscopy, respectively.ResultsIn order to explore the role of SUMOylation in ALD, Mass Spectrometry (MS) was performed to identify SUMOylated proteins in 10‐day ethanol‐feeding+1 Binge ethanol (NIAAA) model, where SUMOs binding columns were used to purify SUMOylated proteins from total livers. Interestingly, we found that ethanol induces changes in SUMOylation state of several mitochondrial proteins involved in ATP synthesis (ATP5B, SOD2, CLC25A5, HSPD1, FBP1, CYCS), ATP metabolism (ATP5A1, ATP5B, ATP5C1, ATP5F1, ATP5J2, ASPA8, AK3), and mitochondria disorder (UQCRC2, PC, HMGCS2, ECHS1, COX6B1, NDUFV3, COX6C). These finding could suggest a potential role of SUMOylation in oxidative phosphorylation, electron transport chain (ETC) and respiratory control ratio may be modulating the ability of these proteins to form the complex above and/or regulating their activity. Specifically, ethanol treatment decreases ATP production and maximal respiration in primary mouse hepatocytes, while UBC9 knockdown prevents the loss of respiration. Furthermore, UBC9 silencing prevents the drop of Δψm when ethanol is added. This finding clearly indicates that SUMOylation is involved in mitochondrial physiology.Conclusionsthese novel finding provides highly novel information on the role of SUMOylation in the development of ALD and may provide new therapeutic strategies, which is of high public health relevance.Support or Funding InformationNIH/NIAAA R21AA027325‐01A1

Oral dextrose reduced procedural pain without altering cellular ATP metabolism in preterm neonates: a prospective randomized trial

ObjectiveTo examine the effects of 30% oral dextrose on biochemical markers of pain, adenosine triphosphate (ATP) degradation, and oxidative stress in preterm neonates experiencing a clinically required heel lance.Study designUtilizing a prospective study design, preterm neonates that met study criteria (n = 169) were randomized to receive either (1) 30% oral dextrose, (2) facilitated tucking, or (3) 30% oral dextrose and facilitated tucking 2 min before heel lance. Plasma markers of ATP degradation (hypoxanthine, uric acid) and oxidative stress (allantoin) were measured before and after the heel lance. Pain was measured using the premature infant pain profile-revised (PIPP-R).ResultsOral dextrose, administered alone or with facilitated tucking, did not alter plasma markers of ATP utilization and oxidative stress.ConclusionA single dose of 30% oral dextrose, given before a clinically required heel lance, decreased signs of pain without increasing ATP utilization and oxidative stress in premature neonates.

CD73 or CD39 Deletion Reveals Different Mechanisms of Formation for Spontaneous and Mechanically Stimulated Adenosine and Sex Specific Compensations in ATP Degradation.

Adenosine is important for local neuromodulation, and rapid adenosine signaling can occur spontaneously or after mechanical stimulation, but little is known about how adenosine is formed in the extracellular space for those stimulations. Here, we studied mechanically stimulated and spontaneous adenosine to determine if rapid adenosine is formed by extracellular breakdown of adenosine triphosphate (ATP) using mice globally deficient in extracellular breakdown enzymes, either CD39 (nucleoside triphosphate diphosphohydrolase 1, NTPDase1) or CD73 (ecto-5'-nucleotidase). CD39 knockout (KO) mice have a lower frequency of spontaneous adenosine events than wild-type (WT, C57BL/6). Surprisingly, CD73KO mice demonstrate sex differences in spontaneous adenosine; males maintain similar event frequencies as WT, but females have significantly fewer events and lower concentrations. Examining the mRNA expression of other enzymes that metabolize ATP revealed tissue nonspecific alkaline phosphatase (TNAP) was upregulated in male CD73KO mice, but not secreted prostatic acid phosphatase (PAP) or transmembrane PAP. Thus, TNAP upregulation compensates for CD73 loss in males but not in females. These sex differences highlight that spontaneous adenosine is formed by metabolism of extracellular ATP by many enzymes. For mechanically stimulated adenosine, CD39KO or CD73KO did not change stimulation frequency, concentration, or t1/2. Thus, the mechanism of formation for mechanically stimulated adenosine is likely direct release of adenosine, different than spontaneous adenosine. Understanding these different mechanisms of rapid adenosine formation will help to develop pharmacological treatments that differentially target modes of rapid adenosine signaling, and all treatments should be studied in both sexes, given possible differences in extracellular ATP degradation.

Single-Cell Adenosine Triphosphate Content Monitoring during Hyperthermia Cell Death by Using Plasmonic Fluorescent Nanoflare.

Gold nanorods-based plasmonic photothermal therapy (AuNRs-PPTT) is a prospective anticancer approach in which AuNRs absorb near-infrared (NIR) light and convert it into heat, leading to cell death. Investigating the molecular energy metabolism of single cells, especially cancer cells, during the hyperthermia cell death process is therefore of great significance, as it can help us to better understand the photothermal lethal mechanism of cancer cells and design new photothermal probes more rationally. However, during the AuNRs-PPTT process, how the cells respond to heat stimulation and how their energy metabolism changes have rarely been studied. Herein, we selected adenosine triphosphate (ATP) as a target molecule, and by preparing a plasmonic and turn-on type fluorescent nanoprobe, we examined the ATP metabolism difference between cancerous cells and normal cells during the AuNRs-PPTT process. We found that the fluorescence intensity increased ∼60% after 5 min laser irradiation as compared to the initial intensity in single HeLa cells, but only ∼20% increase was observed for single H8 cells; obviously, the increase of ATP content in cancerous cells was notably higher than that in normal cells during the hyperthermia cell death.