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measurement of ATP

Massimo D'Agostino, Andreas Mayer
Fluorescence de-quenching can be used to analyze membrane lipid mixing during an in vitro fusion reaction. Here we describe a method to measure lipid mixing using vacuolar membranes purified from the yeast Saccharomyces cerevisiae. Labeling the isolated organelles with rhodamine-phosphatidylethanolamine allows to reveal ATP-dependent lipid mixing through fluorescence de-quenching in a spectrofluorometer. Combining this assay with content mixing indicators, such as the fusion-dependent maturation of a luminal vacuolar phosphatase, then permits the detection of hemifusion intermediates and the analysis of the requirements for fusion pore opening...
2019: Methods in Molecular Biology
Marzieh Rashedinia, Jamileh Saberzadeh, Tannaz Khosravi Bakhtiari, Solmaz Hozhabri, Rita Arabsolghar
Glycyrrhizic acid (GA) is the most effective ingredient in the root of licorice, with important pharmacological effects. We investigate the effects of GA on mitochondrial function and biogenesis in the aluminum toxicity in PC12 cell line. After pretreatment of PC12 cells with different concentrations of GA (5-100 μM), and specific concentration of aluminum maltolate (Almal,1000 μM) for 48 h, cell viability, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), mitochondria mass, cytochrome c oxidase enzyme activity, and the ATP level of the cells were measured...
October 13, 2018: Neurotoxicity Research
Matthew J Bird, Silvia Radenkovic, Pieter Vermeersch, David Cassiman
Here, we offer you a high-throughput assay to measure the ATP synthesis capacity in cells or isolated mitochondria. More specifically, the assay is linked to the mitochondrial' electron transport chain components of your interest being either through complex I (with or without a linkage to pyruvate dehydrogenase activity), through complex II, or through the electron transport flavoprotein and complex I (β-oxidation of fatty acids).
2019: Methods in Molecular Biology
Jia Jia, Na Li, Guofu Li, Bin Zang
OBJECTIVE: To investigate the pathogenesis of early biotrauma in ventilator-induced lung injury (VILI). METHODS: Twenty-four 8-week-old male specific-pathogen-free Sprague-Dawley (SD) rats weighing 250-300 g were randomly divided into sham group (S group), conventional mechanical ventilation group (L group) and high tidal volume (VT) mechanical ventilation group (H group) with 8 rats in each group. All rats received tracheostomy after anesthesia. Rats in S group received no mechanical ventilation but breathe room air spontaneously...
September 2018: Zhonghua Wei Zhong Bing Ji Jiu Yi Xue
Yue Yao, Qiang Li, Wei Wang, Jinchao Zhang, Ping Gao, Yi Xu
BACKGROUND/AIMS: Abnormal regulation of cholesterol homeostasis is associated with type 2 diabetes mellitus (T2DM) and multiple other diseases. Glucagon-like peptide-1 (GLP-1) has unique effects on modulating hepatic lipid metabolism. However, the mechanism behind these is largely unknown. The aim of this study was to investigate the effects of GLP-1 on cholesterol-induced lipotoxicity in hepatocytes and examine the underlying mechanisms. METHODS: Cell viability was determined by CCK-8...
October 11, 2018: Cellular Physiology and Biochemistry
Adelina Martín Salvador, Rafael Fernández Castillo, Inmaculada García García, María José Aguilar Cordero, Juan Bravo Soto
INTRODUCTION: the appearance of metabolic syndrome (MS) among renal recipients is one of the greatest post-transplant complications and is associated with an increased risk of graft failure and high rates of obesity and new onset diabetes. OBJECTIVE: the objective of this work is to identify the relationship between the glomerular filtration rate measured by two different methods and the components of the metabolic syndrome and their combinations in kidney transplant patients according to gender...
October 5, 2018: Nutrición Hospitalaria: Organo Oficial de la Sociedad Española de Nutrición Parenteral y Enteral
Petteri Rinne, Leo-Pekka Lyytikäinen, Emma Raitoharju, James J Kadiri, Ivana Kholova, Mika Kähönen, Terho Lehtimäki, Niku Oksala
α-melanocyte-stimulating hormone (α-MSH) is processed from pro-opiomelanocortin (POMC) and mediates anti-inflammatory actions in leukocytes. α-MSH also promotes macrophage reverse cholesterol transport by inducing ATP-binding cassette transporters ABCA1 and ABCG1. Here we investigated the regulation of POMC and α-MSH expression in atherosclerosis. First, transcript levels of POMC and its processing enzymes were analyzed in human arterial plaques (n = 68) and non-atherosclerotic controls (n = 24) as well as in whole blood samples from coronary artery disease patients (n = 55) and controls (n = 45) by microarray...
October 10, 2018: Scientific Reports
Ying Zhang, Jin Liu, Xiao-Qiang Chen, C-Y Oliver Chen
Coenzyme Q10 (CoQ10) exerts its functions in the body through the ability of its benzoquinone head group to accept and donate electrons. The primary functions are to relay electrons for ATP production in the electron transport chain and to act as an important lipophilic antioxidant. Ubiquinone, the oxidized form of CoQ10, is commonly formulated in commercial supplements, and it must be reduced to ubiquinol to exert CoQ10's functions after consumption. Thus, we aimed to examine whether as compared to ubiquinone, ubiquinol would be more effective to enhance the CoQ10 status in older men...
October 10, 2018: Food & Function
Bedoor Qabazard, Mariam H M Yousif, Oludotun A Phillips
GYY4137 is a novel hydrogen sulfide (H2 S) releasing molecule with vasodilator activity. The objectives of this study were to investigate: (1) the pharmacological effect of GYY4137 on the reactivity of the corpus cavernosum (CC) from normal and diabetic rats; (2) the contribution of ATP-sensitive potassium (K-ATP) channels and nitric oxide (NO) pathway; (3) the reactivity to vasoactive agonists following ex vivo incubation of the diabetic rat CC with GYY4137. Longitudinal strips of CC from control and diabetic male Sprague-Dawley (SD) rats (n = 5-6 animals per group) were suspended in organ-baths...
October 9, 2018: International Journal of Impotence Research
Wenjie Li, Min Yao, Ruonan Wang, Yun Shi, Lianguo Hou, Ziyuan Hou, Kaoqi Lian, Nan Zhang, Yaqi Wang, Weiwei Li, Wei Wang, Lingling Jiang
BACKGROUND: Lipotoxicity contributes to diabetic myocardial disease. In this study, we investigated the lipid species contributing to lipotoxicity and the relationship with peroxisomal β-oxidation in the heart of diabetic mice. METHODS: Male C57BL/6 mice were randomly divided into a Diabetic group (intraperitoneal injection of STZ) and a Control group (saline). Cardiac function indexes [ejection fraction (EF%) and fractional shortening (FS%)] were evaluated by echocardiography...
October 9, 2018: Lipids in Health and Disease
Pedro Cisternas, Juan M Zolezzi, Milka Martinez, Viviana I Torres, G William Wong, Nibaldo C Inestrosa
Dysregulated Wnt signaling is linked to major neurodegenerative diseases, including Alzheimer disease (AD). In mouse models of AD, activation of the canonical Wnt signaling pathway improves learning/memory, but the mechanism for this remains unclear. The decline in brain function in AD patients correlates with reduced glucose utilization by neurons. Here, we test whether improvements in glucose metabolism mediate the neuroprotective effects of Wnt in AD mouse model. APPswe/PS1dE9 transgenic mice were used to model AD, Andrographolide or Lithium was used to activate Wnt signaling, and cytochalasin B was used to block glucose uptake...
October 9, 2018: Journal of Neurochemistry
Marisa Iborra, Inés Moret, Francisco Rausell, Enrique Busó, Elena Cerrillo, Esteban Sáez-González, Pilar Nos, Belén Beltrán
BACKGROUND/AIMS: Increased oxidative stress and decreased immune cell apoptosis have been reported to be important factors in the pathogenesis of Crohn's disease (CD). Our aim was to characterize the genetic expression of molecules implicated in the regulation of oxidative stress and apoptosis in peripheral white mononuclear cells of 18 healthy volunteers (controls) and 20 patients at the onset of CD (active CD [aCD]): 10 who achieved remission (inactive CD [iCD]) and 10 who did not present a complete and deep response to treatment (aCD-T)...
October 9, 2018: Digestion
Juan Qiao, Chuanfang Chen, Dihua Shangguan, Xiaoyu Mu, Shutao Wang, Lei Jiang, Li Qi
Real-time monitoring of the distribution of energy released during oxidative phosphorylation (OXPHOS) in living cells would advance the understanding of metabolic pathways and cell biology. However, the relationship between intracellular temperature and ATP fluctuation during the OXPHOS process is rarely studied due to the limitation of the sensing approach. Novel fluorescent polymer probes were developed for accurate simultaneous measurements of intracellular temperature and ATP. Utilizing the fluorescence imaging techniques, it was demonstrated for the first time that the temperature in mitochondria increased 2...
October 8, 2018: Analytical Chemistry
Paul G Weightman Potter, Julia M Vlachaki Walker, Josephine L Robb, John K Chilton, Ritchie Williamson, Andrew D Randall, Kate L J Ellacott, Craig Beall
AIMS/HYPOTHESIS: Hypoglycaemia is a major barrier to good glucose control in type 1 diabetes. Frequent hypoglycaemic episodes impair awareness of subsequent hypoglycaemic bouts. Neural changes underpinning awareness of hypoglycaemia are poorly defined and molecular mechanisms by which glial cells contribute to hypoglycaemia sensing and glucose counterregulation require further investigation. The aim of the current study was to examine whether, and by what mechanism, human primary astrocyte (HPA) function was altered by acute and recurrent low glucose (RLG)...
October 6, 2018: Diabetologia
Dick van der Kooij, Harm R Veenendaal, Ronald Italiaander, Ed J van der Mark, Marco Dignum
Slow sand filtration with extensive pre-treatment reduces the microbial growth potential of drinking water to a minimum level at four surface water supplies in The Netherlands. The potential of these slow sand filtrates (SSFs) to promote microbial growth in warm tap water installations was assessed by measuring biofilm formation and growth of Legionella on surfaces of glass and chlorinated polyvinylchloride (CPVC) exposed to SSF of 37 ± 2°C in a model system for up to six months. The steady-state biofilm concentration on glass ranged from 230 to 3980 pg ATP cm-2 and 1...
October 5, 2018: Applied and Environmental Microbiology
Merlin G Butler, Waheeda A Hossain, Robert Tessman, Partha C Krishnamurthy
Prader-Willi syndrome (PWS) is a complex multisystem disorder because of errors in genomic imprinting with severe hypotonia, decreased muscle mass, poor suckling, feeding problems and failure to thrive during infancy, growth and other hormone deficiency, childhood-onset hyperphagia, and subsequent obesity. Decreased energy expenditure in PWS is thought to contribute to reduced muscle mass and physical activity but may also relate to cellular metabolism and disturbances in mitochondrial function. We established fibroblast cell lines from six children and adults with PWS and six healthy controls for mitochondrial assays...
October 5, 2018: American Journal of Medical Genetics. Part A
Leonie Durnin, Benjamin Kwok, Priya Kukadia, Roisin McAvera, Robert D Corrigan, Sean M Ward, Ying Zhang, Qi Chen, Sang Don Koh, Kenton M Sanders, Violeta N Mutafova-Yambolieva
A KEY POINT LIST: Studies of urothelial cells, bladder sheets or lumens of filled bladders have suggested that mediators released from urothelium into suburothelium (SubU)/lamina propria (LP) activate mechanisms controlling detrusor excitability. None of these approaches, however, has enabled direct assessment of availability of mediators at SubU/LP during filling. We developed an ex vivo mouse bladder preparation with intact urothelium and SubU/LP but no detrusor, which allows direct access to the SubU/LP surface of urothelium during filling...
October 5, 2018: Journal of Physiology
Ondřej Gahura, Brian Panicucci, Hana Váchová, John E Walker, Alena Zíková
Hydrolysis of ATP by the mitochondrial F-ATPase is inhibited by a protein called IF1 . In the parasitic flagellate, Trypanosoma brucei, this protein, known as TbIF1 , is expressed exclusively in the procyclic stage, where the F-ATPase is synthesizing ATP. In the bloodstream stage, where TbIF1 is absent, the F-ATPase hydrolyzes ATP made by glycolysis and compensates for the absence of a proton pumping respiratory chain by translocating protons into the intermembrane space, thereby maintaining the essential mitochondrial membrane potential...
October 4, 2018: FEBS Journal
Lei Wang, Yugo Iwasaki, Kiran K Andra, Kalpana Pandey, Anant K Menon, Peter Bütikofer
Members of the G protein-coupled receptor and transmembrane protein 16 (TMEM16) protein families are phospholipid scramblases that facilitate rapid, bidirectional movement of phospholipids across a membrane bilayer in an ATP-independent manner. On reconstitution into large unilamellar vesicles, these proteins scramble more than 10,000 lipids per protein per second as measured with core-constituted fluorescent nitrobenzoxadiazole (NBD)-labeled phospholipids. Although NBD-labeled phospholipids are ubiquitously used as reporters of scramblase activity, it remains unclear whether the NBD modification influences the quantitative outcomes of the scramblase assay...
October 4, 2018: Journal of Biological Chemistry
Miriam Saiz-Rodríguez, Dolores Ochoa, Coral Herrador, Carmen Belmonte, Manuel Román, Enrique Alday, Dora Koller, Pablo Zubiaur, Gina Mejía, María Hernández-Martínez, Francisco Abad-Santos
Fentanyl is an agonist of the μ-opioid receptor commonly used in the treatment of moderate-severe pain. In order to study whether pharmacogenetics explain some of the variability in the response to fentanyl, several genes related to fentanyl receptors, transporters and metabolic enzymes have been analysed. Thirty-five healthy volunteers (19 men and 16 women) receiving a single 300 μg oral dose of fentanyl were genotyped for 9 polymorphisms in cytochrome P450 (CYP) enzymes (CYP3A4 and CYP3A5), ATP binding cassette subfamily B member 1 (ABCB1), opioid receptor mu 1 (OPRM1), catechol-O-methyltransferase (COMT) and adrenoceptor beta 2 (ADRB2) by real-time PCR...
October 3, 2018: Basic & Clinical Pharmacology & Toxicology
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