Low glucose and high pyruvate reduce the production of 2-oxoaldehydes, improving mitochondrial efficiency, redox regulation and stallion sperm function

2021 ◽  
Author(s):  
J M Ortiz-Rodríguez ◽  
F E Martín-Cano ◽  
G Gaitskell-Phillips ◽  
A Silva ◽  
C Ortega-Ferrusola ◽  
...  
Keyword(s):  
Redox Regulation ◽  
Krebs Cycle ◽  
Tca Cycle ◽  
Mitochondrial Activity ◽  
Atp Production ◽  
Transport Chain ◽  
Low Glucose ◽  
Mitochondrial Efficiency ◽  
Metabolic Strategies ◽  
Go Terms

Abstract Energy metabolism in spermatozoa is complex and involves the metabolism of carbohydrate fatty acids and amino acids. The ATP produced in the electron transport chain (ETC) in the mitochondria appears to be crucial for both sperm motility and maintaining viability, while glycolytic enzymes in the flagella may contribute to ATP production to sustain motility and velocity. Stallion spermatozoa seemingly use diverse metabolic strategies, and in this regard, a study of the metabolic proteome showed that gene ontology (GO) terms and Reactome pathways related to pyruvate metabolism and the Krebs cycle were predominant. Following this, the hypothesis that low glucose concentrations can provide sufficient support for motility and velocity, and thus glucose concentration can be significantly reduced in the medium, was tested. Aliquots of stallion semen in four different media were stored for 48 h at 18°C; a commercial extender containing 67 mM glucose was used as a control. Stallion spermatozoa stored in media with low glucose (1 mM) and high pyruvate (10 mM) (LG-HP) sustained better motility and velocities than those stored in the commercial extender formulated with very high glucose (61.7 ± 1.2% in INRA 96 vs 76.2 ± 1.0% in LG-HP media after 48 h of incubation at 18°C P < 0.0001). Moreover, mitochondrial activity was superior in LG-HP extenders (24.1 ± 1.8% in INRA 96 vs 51.1 ± 0.7% in LG-HP of spermatozoa with active mitochondria after 48 h of storage at 18°C P < 0.0001). Low glucose concentrations may permit more efficient sperm metabolism and redox regulation when substrates for an efficient TCA cycle are provided. The improvement seen using low glucose extenders is due to reductions in the levels of glyoxal and methylglyoxal, 2-oxoaldehydes formed during glycolysis; these compounds are potent electrophiles able to react with proteins, lipids and DNA, causing sperm damage.

scholarly journals 010.Coordinating the transition from egg to embryo in mammals

2004 ◽  
Vol 16 (9) ◽  
pp. 10 ◽  
Author(s):  
J. Carroll
Keyword(s):  
Tca Cycle ◽  
Cell Types ◽  
Mitotic Division ◽  
Cortical Granule ◽  
Mitochondrial Activity ◽  
Mammalian Development ◽  
Atp Production ◽  
Downstream Target ◽  
Cell Functions ◽  
Cortical Granule Exocytosis

At fertilization of mammalian oocytes, the sperm induces a series of increases in the concentration of intracellular Ca2+. These Ca2+ oscillations trigger all the events of egg activation, including cortical granule exocytosis, completion of meiosis and entry into the first mitotic division. Thus, intracellular Ca2+ plays a pivotal role in coordinating the transition from egg to embryo. Our work is focussed on understanding how the oocyte prepares for fertilisation, how the Ca2+ oscillations are controlled and how Ca2+ stimulates signalling pathways that lead to optimal early embryonic development. In this lecture I will focus on the downstream pathways of Ca2+ signalling at fertilisation. Conventional Protein Kinase C (cPKC) is the major downstream target of Ca2+ in many cell functions. Using PKC-GFP fusion proteins we have found that cPKC is recruited to the membrane in a manner that is dependent on the frequency and amplitude of the Ca2+ oscillations. Recruitment of cPKC appears to promote the Ca2+ influx necessary to sustain the generation of long lasting Ca2+ oscillations. In other cell types cytosolic Ca2+ increases are known to stimulate mitochondrial respiration. We have found that maintenance of resting Ca2+ levels and sperm-induced Ca2+ oscillations are critically dependent on mitochondrial ATP production: a feature not shared by many cell types. Since Ca2+ release increases ATP consumption we investigated whether the Ca2+ transients increase mitochondrial activity so as to meet this increase in demand. Monitoring autofluorescence from NADH and flavoproteins reveals that Ca2+ transients stimulate a change in redox state of mitochondria, presumably by activating Ca2+-sensitive dehydrogenases of the TCA cycle. Thus, through activation of downstream pathways, including PKC, cyclin B degradation and mitochondrial activity, intracellular Ca2+ provides a signal that orchestrates the activation of early mammalian development.

scholarly journals Slow TCA flux implies low ATP production in tumors

2021 ◽  
Author(s):  
Caroline R. Bartman ◽  
Yihui Shen ◽  
Won Dong Lee ◽  
Tara TeSlaa ◽  
Connor S.R. Jankowski ◽  
...  
Keyword(s):  
Electron Transport ◽  
Warburg Effect ◽  
Electron Transport Chain ◽  
Aerobic Glycolysis ◽  
Tca Cycle ◽  
High Energy ◽  
Glycolytic Flux ◽  
Atp Production ◽  
Transport Chain ◽  
The Warburg Effect

SummaryThe tricarboxylic acid (TCA) cycle oxidizes carbon substrates to carbon dioxide, with the resulting high energy electrons fed into the electron transport chain to produce ATP by oxidative phosphorylation. Healthy tissues derive most of their ATP from oxidative metabolism, and the remainder from glycolysis. The corresponding balance in tumors remains unclear. Tumors upregulate aerobic glycolysis (the Warburg effect), yet they also typically require an intact TCA cycle and electron transport chain1–6. Recent studies have measured which nutrients contribute carbon to the tumor TCA metabolites7,8, but not tumor TCA flux: how fast the cycle turns. Here, we develop and validate an in vivo dynamic isotope tracing-mass spectrometry strategy for TCA flux quantitation, which we apply to all major mouse organs and to five tumor models. We show that, compared to the tissue of origin, tumor TCA flux is markedly suppressed. Complementary glycolytic flux measurements confirm tumor glycolysis acceleration, but the majority of tumor ATP is nevertheless made aerobically, and total tumor ATP production is suppressed compared to healthy tissues. In murine pancreatic cancer, this is accommodated by downregulation of the major energy-using pathway in the healthy exocrine pancreas, protein synthesis. Thus, instead of being hypermetabolic as commonly assumed, tumors apparently make ATP at a lower than normal rate. We propose that, as cells de-differentiate into cancer, they eschew ATP-intensive processes characteristic of the host tissue, and that the resulting suppressed ATP demand contributes to the Warburg effect and facilitates cancer growth in the nutrient-poor tumor microenvironment.

scholarly journals ACE overexpression in myeloid cells increases oxidative metabolism and cellular ATP

2019 ◽  
Vol 295 (5) ◽  
pp. 1369-1384 ◽  
Author(s):  
Duo-Yao Cao ◽  
Weston R. Spivia ◽  
Luciana C. Veiras ◽  
Zakir Khan ◽  
Zhenzi Peng ◽  
...  
Keyword(s):  
Ace Inhibitor ◽  
Cell Function ◽  
Krebs Cycle ◽  
Myeloid Cells ◽  
Myeloid Cell ◽  
Complex Iv ◽  
Atp Production ◽  
Transport Chain ◽  
Intermediate Metabolites ◽  
Krebs Cycle Intermediates

Angiotensin-converting enzyme (ACE) affects blood pressure. In addition, ACE overexpression in myeloid cells increases their immune function. Using MS and chemical analysis, we identified marked changes of intermediate metabolites in ACE-overexpressing macrophages and neutrophils, with increased cellular ATP (1.7–3.0-fold) and Krebs cycle intermediates, including citrate, isocitrate, succinate, and malate (1.4–3.9-fold). Increased ATP is due to ACE C-domain catalytic activity; it is reversed by an ACE inhibitor but not by an angiotensin II AT1 receptor antagonist. In contrast, macrophages from ACE knockout (null) mice averaged only 28% of the ATP levels found in WT mice. ACE overexpression does not change cell or mitochondrial size or number. However, expression levels of the electron transport chain proteins NDUFB8 (complex I), ATP5A, and ATP5β (complex V) are significantly increased in macrophages and neutrophils, and COX1 and COX2 (complex IV) are increased in macrophages overexpressing ACE. Macrophages overexpressing ACE have increased mitochondrial membrane potential (24% higher), ATP production rates (29% higher), and maximal respiratory rates (37% higher) compared with WT cells. Increased cellular ATP underpins increased myeloid cell superoxide production and phagocytosis associated with increased ACE expression. Myeloid cells overexpressing ACE indicate the existence of a novel pathway in which myeloid cell function can be enhanced, with a key feature being increased cellular ATP.

scholarly journals Rescue of TCA Cycle Dysfunction for Cancer Therapy

2019 ◽  
Vol 8 (12) ◽  
pp. 2161 ◽  
Author(s):  
Jubert Marquez ◽  
Jessa Flores ◽  
Amy Hyein Kim ◽  
Bayalagmaa Nyamaa ◽  
Anh Thi Tuyet Nguyen ◽  
...  
Keyword(s):  
Tca Cycle ◽  
Mitochondrial Enzymes ◽  
Mitochondrial Enzyme ◽  
Therapeutic Approaches ◽  
Atp Production ◽  
Tca Cycle Enzymes ◽  
Subcellular Organelle ◽  
The Tca Cycle ◽  
Transport Chain ◽  
Cancer Types

Mitochondrion, a maternally hereditary, subcellular organelle, is the site of the tricarboxylic acid (TCA) cycle, electron transport chain (ETC), and oxidative phosphorylation (OXPHOS)—the basic processes of ATP production. Mitochondrial function plays a pivotal role in the development and pathology of different cancers. Disruption in its activity, like mutations in its TCA cycle enzymes, leads to physiological imbalances and metabolic shifts of the cell, which contributes to the progression of cancer. In this review, we explored the different significant mutations in the mitochondrial enzymes participating in the TCA cycle and the diseases, especially cancer types, that these malfunctions are closely associated with. In addition, this paper also discussed the different therapeutic approaches which are currently being developed to address these diseases caused by mitochondrial enzyme malfunction.

scholarly journals Photobiomodulation and Oxidative Stress: 980 nm Diode Laser Light Regulates Mitochondrial Activity and Reactive Oxygen Species Production

2021 ◽  
Vol 2021 ◽  
pp. 1-11 ◽  
Author(s):  
Andrea Amaroli ◽  
Claudio Pasquale ◽  
Angelina Zekiy ◽  
Anatoliy Utyuzh ◽  
Stefano Benedicenti ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Reactive Oxygen Species ◽  
Oxygen Consumption ◽  
Respiratory Chain ◽  
Laser Light ◽  
Krebs Cycle ◽  
Mitochondrial Activity ◽  
Oxygen Species ◽  
Atp Production ◽  
And Oxidative Stress

Photobiomodulation with 808 nm laser light electively stimulates Complexes III and IV of the mitochondrial respiratory chain, while Complexes I and II are not affected. At the wavelength of 1064 nm, Complexes I, III, and IV are excited, while Complex II and some mitochondrial matrix enzymes seem to be not receptive to photons at that wavelength. Complex IV was also activated by 633 nm. The mechanism of action of wavelengths in the range 900–1000 nm on mitochondria is less understood or not described. Oxidative stress from reactive oxygen species (ROS) generated by mitochondrial activity is an inescapable consequence of aerobic metabolism. The antioxidant enzyme system for ROS scavenging can keep them under control. However, alterations in mitochondrial activity can cause an increment of ROS production. ROS and ATP can play a role in cell death, cell proliferation, and cell cycle arrest. In our work, bovine liver isolated mitochondria were irradiated for 60 sec, in continuous wave mode with 980 nm and powers from 0.1 to 1.4 W (0.1 W increment at every step) to generate energies from 6 to 84 J, fluences from 7.7 to 107.7 J/cm2, power densities from 0.13 to 1.79 W/cm2, and spot size 0.78 cm2. The control was equal to 0 W. The activity of the mitochondria’s complexes, Krebs cycle enzymes, ATP production, oxygen consumption, generation of ROS, and oxidative stress were detected. Lower powers (0.1–0.2 W) showed an inhibitory effect; those that were intermediate (0.3–0.7 W) did not display an effect, and the higher powers (0.8–1.1 W) induced an increment of ATP synthesis. Increasing the power (1.2–1.4 W) recovered the ATP production to the control level. The interaction occurred on Complexes III and IV, as well as ATP production and oxygen consumption. Results showed that 0.1 W uncoupled the respiratory chain and induced higher oxidative stress and drastic inhibition of ATP production. Conversely, 0.8 W kept mitochondria coupled and induced an increase of ATP production by increments of Complex III and IV activities. An augmentation of oxidative stress was also observed, probably as a consequence of the increased oxygen consumption and mitochondrial isolation experimental conditions. No effect was observed using 0.5 W, and no effect was observed on the enzymes of the Krebs cycle.

scholarly journals Monitoring cytosolic H2O2 fluctuations arising from altered plasma membrane gradients or from mitochondrial activity

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Mercè Carmona ◽  
Laura de Cubas ◽  
Eric Bautista ◽  
Marta Moral-Blanch ◽  
Iria Medraño-Fernández ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Steady State ◽  
Fission Yeast ◽  
Electron Transport Chain ◽  
Metabolic Reprogramming ◽  
Mitochondrial Activity ◽  
Transport Chain ◽  
Steady State Levels ◽  
Low Glucose ◽  
Living Organisms

Abstract Genetically encoded probes monitoring H2O2 fluctuations in living organisms are key to decipher redox signaling events. Here we use a new probe, roGFP2-Tpx1.C169S, to monitor pre-toxic fluctuations of peroxides in fission yeast, where the concentrations linked to signaling or to toxicity have been established. This probe is able to detect nanomolar fluctuations of intracellular H2O2 caused by extracellular peroxides; expression of human aquaporin 8 channels H2O2 entry into fission yeast decreasing membrane gradients. The probe also detects H2O2 bursts from mitochondria after addition of electron transport chain inhibitors, the extent of probe oxidation being proportional to the mitochondrial activity. The oxidation of this probe is an indicator of steady-state levels of H2O2 in different genetic backgrounds. Metabolic reprogramming during growth in low-glucose media causes probe reduction due to the activation of antioxidant cascades. We demonstrate how peroxiredoxin-based probes can be used to monitor physiological H2O2 fluctuations.

scholarly journals Bioenergetic and metabolic impairments in Duchenne Muscular Dystrophy (DMD) patients' iPSC-derived cardiomyocytes

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
L Willi ◽  
B Agranovich ◽  
I Abramovich ◽  
D Freimark ◽  
M Arad ◽  
...  
Keyword(s):  
Stress Test ◽  
Young Male ◽  
Induced Pluripotent Stem Cell ◽  
Dystrophin Gene ◽  
Energy System ◽  
Basal Respiration ◽  
Mitochondrial Activity ◽  
Funding Source ◽  
Atp Production ◽  
Metabolic Impairments

Abstract Introduction DMD, an X-linked muscle degenerative fatal disease, is caused by mutations in the dystrophin gene. Dilated cardiomyopathy (DCM) is a major cause of morbidity and mortality in DMD patients. Treatments for DCM in DMD are limited to steroids and standard heart failure medications such as β-blockers and ACE-inhibitors, and therefore novel therapeutic modalities are urgently needed. Purpose We hypothesized that dystrophin mutations in DMD lead to cardiomyopathy-causing bioenergetic/metabolic impairments, which can be therapeutically targeted for improving cardiac function. Methods Induced Pluripotent Stem Cell-derived cardiomyocytes (iPSC-CMs) were generated from healthy volunteer and 3 DMD patients: young male (YM), adult male (AM) and adult female (AF). We investigated the bioenergetics, electrophysiology, mitochondrial and metabolic features of healthy and DMD iPSC-CMs using the Seahorse Flux analyzer, patch clamp, confocal fluorescence microscopy and Liquid chromatography mass spectrometry (LC-MS) technologies, respectively. Results To test the hypothesis, we measured respiration and glycolytic rates of healthy and DMD iPSC-CMs. Compared to healthy iPSC-CMs, in both AM and AF DMD, but not in YM DMD cardiomyocytes, there was a 75% decrease in ATP production, and 80% and 45% decrease in basal respiration, respectively. In agreement with the healthy-like bioenergetic status of YM, the iPSC-CMs showed no arrhythmias, in contrast to the prominent arrhythmias in AM and AF cardiomyocytes. To determine whether the impairment in the phosphorylation pathway (OXPHOS) affects glycolysis, we measured the cardiomyocytes' response to glycolytic stress test. These experiments showed that the glycolytic rates were similar in healthy and DMD iPSC-CMs. In agreement with impaired OXPHOS, mitochondrial activity measured by 3D life confocal microscopy was attenuated in the DMD male by 35%, compared to healthy cardiomyocytes. Furthermore, the metabolomic LC-MS analyses demonstrated significant differences in metabolite levels in YM, AM and AF DMD iPSC-CMs relative to healthy iPSC-CMs. For example, compared to healthy iPSC-CMs, there was a dramatic fall to undetected levels in phosphocreatine in both AM and AF, but not in YM DMD, indicating a dysfunctional phosphocreatine energy system. Conclusions DMD iPSC-CMs exhibit bioenergetic/metabolic impairments, which constitute novel targets for alleviating the cardiomyopathy in DMD patients. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): ISF - Israel Science Foundation

scholarly journals Cellular and mitochondrial mechanisms of atrial fibrillation

2020 ◽  
Vol 115 (6) ◽  
Author(s):  
Fleur E. Mason ◽  
Julius Ryan D. Pronto ◽  
Khaled Alhussini ◽  
Christoph Maack ◽  
Niels Voigt
Keyword(s):  
Atrial Fibrillation ◽  
Nicotinamide Adenine Dinucleotide ◽  
Molecular Mechanisms ◽  
Treatment Options ◽  
Specific Treatment ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Nicotinamide Adenine ◽  
Mitochondrial Activity ◽  
Atp Production

AbstractThe molecular mechanisms underlying atrial fibrillation (AF), the most common form of arrhythmia, are poorly understood and therefore target-specific treatment options remain an unmet clinical need. Excitation–contraction coupling in cardiac myocytes requires high amounts of adenosine triphosphate (ATP), which is replenished by oxidative phosphorylation in mitochondria. Calcium (Ca2+) is a key regulator of mitochondrial function by stimulating the Krebs cycle, which produces nicotinamide adenine dinucleotide for ATP production at the electron transport chain and nicotinamide adenine dinucleotide phosphate for the elimination of reactive oxygen species (ROS). While it is now well established that mitochondrial dysfunction plays an important role in the pathophysiology of heart failure, this has been less investigated in atrial myocytes in AF. Considering the high prevalence of AF, investigating the role of mitochondria in this disease may guide the path towards new therapeutic targets. In this review, we discuss the importance of mitochondrial Ca2+ handling in regulating ATP production and mitochondrial ROS emission and how alterations, particularly in these aspects of mitochondrial activity, may play a role in AF. In addition to describing research advances, we highlight areas in which further studies are required to elucidate the role of mitochondria in AF.

scholarly journals Complex II subunit SDHD is critical for cell growth and metabolism, which can be partially restored with a synthetic ubiquinone analog

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Aloka B. Bandara ◽  
Joshua C. Drake ◽  
David A. Brown
Keyword(s):  
Mammalian Cells ◽  
Krebs Cycle ◽  
Atp Synthesis ◽  
Hek293 Cells ◽  
Knockout Mutant ◽  
Complex Ii ◽  
Growth Defects ◽  
Transport Chain ◽  
Mutant Cells

Abstract Background Succinate dehydrogenase (Complex II) plays a dual role in respiration by catalyzing the oxidation of succinate to fumarate in the mitochondrial Krebs cycle and transferring electrons from succinate to ubiquinone in the mitochondrial electron transport chain (ETC). Mutations in Complex II are associated with a number of pathologies. SDHD, one of the four subunits of Complex II, serves by anchoring the complex to the inner-membrane and transferring electrons from the complex to ubiquinone. Thus, modeling SDHD dysfunction could be a valuable tool for understanding its importance in metabolism and developing novel therapeutics, however no suitable models exist. Results Via CRISPR/Cas9, we mutated SDHD in HEK293 cells and investigated the in vitro role of SDHD in metabolism. Compared to the parent HEK293, the knockout mutant HEK293ΔSDHD produced significantly less number of cells in culture. The mutant cells predictably had suppressed Complex II-mediated mitochondrial respiration, but also Complex I-mediated respiration. SDHD mutation also adversely affected glycolytic capacity and ATP synthesis. Mutant cells were more apoptotic and susceptible to necrosis. Treatment with the mitochondrial therapeutic idebenone partially improved oxygen consumption and growth of mutant cells. Conclusions Overall, our results suggest that SDHD is vital for growth and metabolism of mammalian cells, and that respiratory and growth defects can be partially restored with treatment of a ubiquinone analog. This is the first report to use CRISPR/Cas9 approach to construct a knockout SDHD cell line and evaluate the efficacy of an established mitochondrial therapeutic candidate to improve bioenergetic capacity.

scholarly journals The Krebs Cycle Enzyme α-Ketoglutarate Decarboxylase Is an Essential Glycosomal Protein in Bloodstream African Trypanosomes

2014 ◽  
Vol 14 (3) ◽  
pp. 206-215 ◽  
Author(s):  
Steven Sykes ◽  
Anthony Szempruch ◽  
Stephen Hajduk
Keyword(s):  
Plasma Membranes ◽  
Fluorescent Protein ◽  
Krebs Cycle ◽  
Content Type ◽  
African Trypanosomes ◽  
Atp Production ◽  
Cycle Enzyme ◽  
A Cell ◽  
Direct Localization ◽  
Green Fluorescent

ABSTRACT α-Ketoglutarate decarboxylase (α-KDE1) is a Krebs cycle enzyme found in the mitochondrion of the procyclic form (PF) of Trypanosoma brucei . The bloodstream form (BF) of T. brucei lacks a functional Krebs cycle and relies exclusively on glycolysis for ATP production. Despite the lack of a functional Krebs cycle, α-KDE1 was expressed in BF T. brucei and RNA interference knockdown of α-KDE1 mRNA resulted in rapid growth arrest and killing. Cell death was preceded by progressive swelling of the flagellar pocket as a consequence of recruitment of both flagellar and plasma membranes into the pocket. BF T. brucei expressing an epitope-tagged copy of α-KDE1 showed localization to glycosomes and not the mitochondrion. We used a cell line transfected with a reporter construct containing the N-terminal sequence of α-KDE1 fused to green fluorescent protein to examine the requirements for glycosome targeting. We found that the N-terminal 18 amino acids of α-KDE1 contain overlapping mitochondrion- and peroxisome-targeting sequences and are sufficient to direct localization to the glycosome in BF T. brucei . These results suggest that α-KDE1 has a novel moonlighting function outside the mitochondrion in BF T. brucei .

Sign in / Sign up

Export Citation Format

Share Document