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1
HILDYARD, John C. W., and Andrew P. HALESTRAP. "Identification of the mitochondrial pyruvate carrier in Saccharomyces cerevisiae." Biochemical Journal 374, no. 3 (September 15, 2003): 607–11. http://dx.doi.org/10.1042/bj20030995.
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Mitochondrial pyruvate transport is fundamental for metabolism and mediated by a specific inhibitable carrier. We have identified the yeast mitochondrial pyruvate carrier by measuring inhibitor-sensitive pyruvate uptake into mitochondria from 18 different Saccharomyces cerevisiae mutants, each lacking an unattributed member of the mitochondrial carrier family (MCF). Only mitochondria from the YIL006w deletion mutant exhibited no inhibitor-sensitive pyruvate transport, but otherwise behaved normally. YIL006w encodes a 41.9 kDa MCF member with homologous proteins present in both the human and mouse genomes.
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Tang, Bor Luen. "Targeting the Mitochondrial Pyruvate Carrier for Neuroprotection." Brain Sciences 9, no. 9 (September 18, 2019): 238. http://dx.doi.org/10.3390/brainsci9090238.
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The mitochondrial pyruvate carriers mediate pyruvate import into the mitochondria, which is key to the sustenance of the tricarboxylic cycle and oxidative phosphorylation. However, inhibition of mitochondria pyruvate carrier-mediated pyruvate transport was recently shown to be beneficial in experimental models of neurotoxicity pertaining to the context of Parkinson’s disease, and is also protective against excitotoxic neuronal death. These findings attested to the metabolic adaptability of neurons resulting from MPC inhibition, a phenomenon that has also been shown in other tissue types. In this short review, I discuss the mechanism and potential feasibility of mitochondrial pyruvate carrier inhibition as a neuroprotective strategy in neuronal injury and neurodegenerative diseases.
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Zangari, Joséphine, Francesco Petrelli, Benoît Maillot, and Jean-Claude Martinou. "The Multifaceted Pyruvate Metabolism: Role of the Mitochondrial Pyruvate Carrier." Biomolecules 10, no. 7 (July 17, 2020): 1068. http://dx.doi.org/10.3390/biom10071068.
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Pyruvate, the end product of glycolysis, plays a major role in cell metabolism. Produced in the cytosol, it is oxidized in the mitochondria where it fuels the citric acid cycle and boosts oxidative phosphorylation. Its sole entry point into mitochondria is through the recently identified mitochondrial pyruvate carrier (MPC). In this review, we report the latest findings on the physiology of the MPC and we discuss how a dysfunctional MPC can lead to diverse pathologies, including neurodegenerative diseases, metabolic disorders, and cancer.
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Reiter, Russel, Ramaswamy Sharma, Sergio Rosales-Corral, Walter Manucha, Luiz Gustavo de Almeida Chuffa, and Debora Aparecida Pires de Campos Zuccari. "Melatonin and Pathological Cell Interactions: Mitochondrial Glucose Processing in Cancer Cells." International Journal of Molecular Sciences 22, no. 22 (November 19, 2021): 12494. http://dx.doi.org/10.3390/ijms222212494.
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Melatonin is synthesized in the pineal gland at night. Since melatonin is produced in the mitochondria of all other cells in a non-circadian manner, the amount synthesized by the pineal gland is less than 5% of the total. Melatonin produced in mitochondria influences glucose metabolism in all cells. Many pathological cells adopt aerobic glycolysis (Warburg effect) in which pyruvate is excluded from the mitochondria and remains in the cytosol where it is metabolized to lactate. The entrance of pyruvate into the mitochondria of healthy cells allows it to be irreversibly decarboxylated by pyruvate dehydrogenase (PDH) to acetyl coenzyme A (acetyl-CoA). The exclusion of pyruvate from the mitochondria in pathological cells prevents the generation of acetyl-CoA from pyruvate. This is relevant to mitochondrial melatonin production, as acetyl-CoA is a required co-substrate/co-factor for melatonin synthesis. When PDH is inhibited during aerobic glycolysis or during intracellular hypoxia, the deficiency of acetyl-CoA likely prevents mitochondrial melatonin synthesis. When cells experiencing aerobic glycolysis or hypoxia with a diminished level of acetyl-CoA are supplemented with melatonin or receive it from another endogenous source (pineal-derived), pathological cells convert to a more normal phenotype and support the transport of pyruvate into the mitochondria, thereby re-establishing a healthier mitochondrial metabolic physiology.
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Moyes, C. D., L. T. Buck, P. W. Hochachka, and R. K. Suarez. "Oxidative properties of carp red and white muscle." Journal of Experimental Biology 143, no. 1 (May 1, 1989): 321–31. http://dx.doi.org/10.1242/jeb.143.1.321.
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Substrate preferences of isolated mitochondria and maximal enzyme activities were used to assess the oxidative capacities of red muscle (RM) and white muscle (WM) of carp (Cyprinus carpio). A 14-fold higher activity of citrate synthase (CS) in RM reflects the higher mitochondrial density in this tissue. RM mitochondria oxidize pyruvate and fatty acyl carnitines (8:O, 12:O, 16:O) at similarly high rates. WM mitochondria oxidize these fatty acyl carnitines at 35–70% the rate of pyruvate, depending on chain length. WM has only half the carnitine palmitoyl transferase/CS ratio of RM, but similar ratios of beta-hydroxyacyl CoA dehydrogenase/CS. Ketone bodies are poor substrates for mitochondria from both tissues. In both tissues mitochondrial alpha-glycerophosphate oxidation was minimal, and alpha-glycerophosphate dehydrogenase was present at low activities, suggesting the alpha-glycerophosphate shuttle is of minor significance in maintaining cytosolic redox balance in either tissue. The mitochondrial oxidation rates of other substrates relative to pyruvate are as follows: alpha-ketoglutarate 90% (RM and WM); glutamate 45% (WM) and 70% (RM); proline 20% (WM) and 45% (RM). Oxidation of neutral amino acids (serine, glycine, alanine, beta-alanine) was not consistently detectable. These data suggest that RM and WM differ in mitochondrial properties as well as mitochondrial abundance. Whereas RM mitochondria appear to be able to utilize a wide range of metabolic fuels (fatty acids, pyruvate, amino acids but not ketone bodies), WM mitochondria appear to be specialized to use pyruvate.
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Simard, Chloé, Andréa Lebel, Eric Pierre Allain, Mohamed Touaibia, Etienne Hebert-Chatelain, and Nicolas Pichaud. "Metabolic Characterization and Consequences of Mitochondrial Pyruvate Carrier Deficiency in Drosophila melanogaster." Metabolites 10, no. 9 (September 6, 2020): 363. http://dx.doi.org/10.3390/metabo10090363.
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In insect, pyruvate is generally the predominant oxidative substrate for mitochondria. This metabolite is transported inside mitochondria via the mitochondrial pyruvate carrier (MPC), but whether and how this transporter controls mitochondrial oxidative capacities in insects is still relatively unknown. Here, we characterize the importance of pyruvate transport as a metabolic control point for mitochondrial substrate oxidation in two genotypes of an insect model, Drosophila melanogaster, differently expressing MPC1, an essential protein for the MPC function. We evaluated the kinetics of pyruvate oxidation, mitochondrial oxygen consumption, metabolic profile, activities of metabolic enzymes, and climbing abilities of wild-type (WT) flies and flies harboring a deficiency in MPC1 (MPC1def). We hypothesized that MPC1 deficiency would cause a metabolic reprogramming that would favor the oxidation of alternative substrates. Our results show that the MPC1def flies display significantly reduced climbing capacity, pyruvate-induced oxygen consumption, and enzymatic activities of pyruvate kinase, alanine aminotransferase, and citrate synthase. Moreover, increased proline oxidation capacity was detected in MPC1def flies, which was associated with generally lower levels of several metabolites, and particularly those involved in amino acid catabolism such as ornithine, citrulline, and arginosuccinate. This study therefore reveals the flexibility of mitochondrial substrate oxidation allowing Drosophila to maintain cellular homeostasis.
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VALENTI, Daniela, Lidia de BARI, Anna ATLANTE, and Salvatore PASSARELLA. "l-Lactate transport into rat heart mitochondria and reconstruction of the l-lactate/pyruvate shuttle." Biochemical Journal 364, no. 1 (May 8, 2002): 101–4. http://dx.doi.org/10.1042/bj3640101.
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In vitro reconstruction of the l-lactate/pyruvate shuttle has been performed, which allows NADH oxidation outside rat heart mitochondria. Such a shuttle occurs due to the combined action of the novel mitochondrial l-lactate/pyruvate antiporter, which differs from the monocarboxylate carrier, and the mitochondrial l-lactate dehydrogenase. The rate of l-lactate/pyruvate antiport proved to regulate the shuttle in vitro.
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Fernandez-Caggiano, Mariana, and Philip Eaton. "Heart failure—emerging roles for the mitochondrial pyruvate carrier." Cell Death & Differentiation 28, no. 4 (January 20, 2021): 1149–58. http://dx.doi.org/10.1038/s41418-020-00729-0.
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AbstractThe mitochondrial pyruvate carrier (MPC) is the entry point for the glycolytic end-product pyruvate to the mitochondria. MPC activity, which is controlled by its abundance and post-translational regulation, determines whether pyruvate is oxidised in the mitochondria or metabolised in the cytosol. MPC serves as a crucial metabolic branch point that determines the fate of pyruvate in the cell, enabling metabolic adaptations during health, such as exercise, or as a result of disease. Decreased MPC expression in several cancers limits the mitochondrial oxidation of pyruvate and contributes to lactate accumulation in the cytosol, highlighting its role as a contributing, causal mediator of the Warburg effect. Pyruvate is handled similarly in the failing heart where a large proportion of it is reduced to lactate in the cytosol instead of being fully oxidised in the mitochondria. Several recent studies have found that the MPC abundance was also reduced in failing human and mouse hearts that were characterised by maladaptive hypertrophic growth, emulating the anabolic scenario observed in some cancer cells. In this review we discuss the evidence implicating the MPC as an important, perhaps causal, mediator of heart failure progression.
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Diers, Anne R., Katarzyna A. Broniowska, Ching-Fang Chang, and Neil Hogg. "Pyruvate fuels mitochondrial respiration and proliferation of breast cancer cells: effect of monocarboxylate transporter inhibition." Biochemical Journal 444, no. 3 (May 29, 2012): 561–71. http://dx.doi.org/10.1042/bj20120294.
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Recent studies have highlighted the fact that cancer cells have an altered metabolic phenotype, and this metabolic reprogramming is required to drive the biosynthesis pathways necessary for rapid replication and proliferation. Specifically, the importance of citric acid cycle-generated intermediates in the regulation of cancer cell proliferation has been recently appreciated. One function of MCTs (monocarboxylate transporters) is to transport the citric acid cycle substrate pyruvate across the plasma membrane and into mitochondria, and inhibition of MCTs has been proposed as a therapeutic strategy to target metabolic pathways in cancer. In the present paper, we examined the effect of different metabolic substrates (glucose and pyruvate) on mitochondrial function and proliferation in breast cancer cells. We demonstrated that cancer cells proliferate more rapidly in the presence of exogenous pyruvate when compared with lactate. Pyruvate supplementation fuelled mitochondrial oxygen consumption and the reserve respiratory capacity, and this increase in mitochondrial function correlated with proliferative potential. In addition, inhibition of cellular pyruvate uptake using the MCT inhibitor α-cyano-4-hydroxycinnamic acid impaired mitochondrial respiration and decreased cell growth. These data demonstrate the importance of mitochondrial metabolism in proliferative responses and highlight a novel mechanism of action for MCT inhibitors through suppression of pyruvate-fuelled mitochondrial respiration.
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Li, Min, Shuang Zhou, Chaoyang Chen, Lingyun Ma, Daohuang Luo, Xin Tian, Xiu Dong, Ying Zhou, Yanling Yang, and Yimin Cui. "Therapeutic potential of pyruvate therapy for patients with mitochondrial diseases: a systematic review." Therapeutic Advances in Endocrinology and Metabolism 11 (January 2020): 204201882093824. http://dx.doi.org/10.1177/2042018820938240.
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Background: Mitochondrial disease is a term used to describe a set of heterogeneous genetic diseases caused by impaired structure or function of mitochondria. Pyruvate therapy for mitochondrial disease is promising from a clinical point of view. Methods: According to PRISMA guidelines, the following databases were searched to identify studies regarding pyruvate therapy for mitochondrial disease: PubMed, EMBASE, Cochrane Library, and Clinicaltrials. The search was up to April 2019. The endpoints were specific biomarkers (plasma level of lactate, plasma level of pyruvate, L/P ratio) and clinical rating scales [Japanese mitochondrial disease-rating scale (JMDRS), Newcastle Mitochondrial Disease Adult Scale (NMDAS), and others]. Two researchers independently screened articles, extracted data, and assessed the quality of the studies. Results: A total of six studies were included. Considerable differences were noted between studies in terms of study design, patient information, and outcome measures. The collected evidence may indicate an effective potential of pyruvate therapy on the improvement of mitochondrial disease. The majority of the common adverse events of pyruvate therapy were diarrhea and short irritation of the stomach. Conclusion: Pyruvate therapy with no serious adverse events may be a potential therapeutic candidate for patients with incurable mitochondrial diseases, such as Leigh syndrome. However, recent evidence taken from case series and case reports, and theoretical supports of basic research are not sufficient. The use of global registries to collect patient data and more adaptive trial designs with larger numbers of participants are necessary to clarify the efficacy of pyruvate therapy.
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Moyes, C. D., P. M. Schulte, and P. W. Hochachka. "Recovery metabolism of trout white muscle: role of mitochondria." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 262, no. 2 (February 1, 1992): R295—R304. http://dx.doi.org/10.1152/ajpregu.1992.262.2.r295.
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Recovery from burst exercise in fish is very slow. Lactate conversion to glycogen occurs primarily within white muscle and must be fueled by mitochondrially produced ATP. In a parallel study we characterized the changes in tissue metabolites associated with burst exercise and recovery in rainbow trout (Oncorhynchus mykiss) white muscle. The present study examines whether the mitochondrial capacity to produce ATP may limit the rate of recovery of trout white muscle. The cost (ATP.min-1.g-1) of glycogen resynthesis (0.05 mumol lactate converted.min-1.g tissue-1) was compared with the mitochondrial capacity to produce ATP. The cost of recovery can be met by only 3.5% of the maximal mitochondrial capacity. In fact, during recovery trout white muscle mitochondria operate at a small fraction of their in vitro maximum. This capacity is suppressed in vivo by highly inhibitory ATP/ADP and limiting phosphate. The primary signal for increased ATP synthesis associated with recovery is not a change in ATP/ADP but probably phosphate, elevated because of phosphocreatine hydrolysis and adenylate catabolism in the purine nucleotide cycle. At low ADP availability and suboptimal phosphate (less than 5 mM), acidosis enhances respiration. At high respiratory rates mitochondrial pyruvate oxidation is sensitive to pyruvate concentration over the physiological range (apparent Michaelis constant = 35-40 microM). This sensitivity is lost at the low rates that approximate in vivo respiration. Changes in lactate do not affect the kinetics of pyruvate oxidation. Fatty acid oxidation may spare pyruvate and lactate for use in glyconeogenesis, primarily through allosteric inhibition of pyruvate dehydrogenase rather than covalent modification.
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Le, Xuyen H., Chun-Pong Lee, and A. Harvey Millar. "The mitochondrial pyruvate carrier (MPC) complex mediates one of three pyruvate-supplying pathways that sustain Arabidopsis respiratory metabolism." Plant Cell 33, no. 8 (June 17, 2021): 2776–93. http://dx.doi.org/10.1093/plcell/koab148.
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Abstract Malate oxidation by plant mitochondria enables the generation of both oxaloacetate and pyruvate for tricarboxylic acid (TCA) cycle function, potentially eliminating the need for pyruvate transport into mitochondria in plants. Here, we show that the absence of the mitochondrial pyruvate carrier 1 (MPC1) causes the co-commitment loss of its putative orthologs, MPC3/MPC4, and eliminates pyruvate transport into Arabidopsis thaliana mitochondria, proving it is essential for MPC complex function. While the loss of either MPC or mitochondrial pyruvate-generating NAD-malic enzyme (NAD-ME) did not cause vegetative phenotypes, the lack of both reduced plant growth and caused an increase in cellular pyruvate levels, indicating a block in respiratory metabolism, and elevated the levels of branched-chain amino acids at night, a sign of alterative substrate provision for respiration. 13C-pyruvate feeding of leaves lacking MPC showed metabolic homeostasis was largely maintained except for alanine and glutamate, indicating that transamination contributes to the restoration of the metabolic network to an operating equilibrium by delivering pyruvate independently of MPC into the matrix. Inhibition of alanine aminotransferases when MPC1 is absent resulted in extremely retarded phenotypes in Arabidopsis, suggesting all pyruvate-supplying enzymes work synergistically to support the TCA cycle for sustained plant growth.
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Herzig, Sébastien, Etienne Raemy, Sylvie Montessuit, Jean-Luc Veuthey, Nicola Zamboni, Benedikt Westermann, Edmund R. S. Kunji, and Jean-Claude Martinou. "Identification and Functional Expression of the Mitochondrial Pyruvate Carrier." Science 337, no. 6090 (May 24, 2012): 93–96. http://dx.doi.org/10.1126/science.1218530.
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The transport of pyruvate, the end product of glycolysis, into mitochondria is an essential process that provides the organelle with a major oxidative fuel. Although the existence of a specific mitochondrial pyruvate carrier (MPC) has been anticipated, its molecular identity remained unknown. We report that MPC is a heterocomplex formed by two members of a family of previously uncharacterized membrane proteins that are conserved from yeast to mammals. Members of the MPC family were found in the inner mitochondrial membrane, and yeast mutants lacking MPC proteins showed severe defects in mitochondrial pyruvate uptake. Coexpression of mouse MPC1 and MPC2 in Lactococcus lactis promoted transport of pyruvate across the membrane. These observations firmly establish these proteins as essential components of the MPC.
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Bowman, Caitlyn E., Liang Zhao, Thomas Hartung, and Michael J. Wolfgang. "Requirement for the Mitochondrial Pyruvate Carrier in Mammalian Development Revealed by a Hypomorphic Allelic Series." Molecular and Cellular Biology 36, no. 15 (May 23, 2016): 2089–104. http://dx.doi.org/10.1128/mcb.00166-16.
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Glucose and oxygen are two of the most important molecules transferred from mother to fetus during eutherian pregnancy, and the metabolic fates of these nutrients converge at the transport and metabolism of pyruvate in mitochondria. Pyruvate enters the mitochondrial matrix through the mitochondrial pyruvate carrier (MPC), a complex in the inner mitochondrial membrane that consists of two essential components, MPC1 and MPC2. Here, we define the requirement for mitochondrial pyruvate metabolism during development with a progressive allelic series of Mpc1 deficiency in mouse. Mpc1 deletion was homozygous lethal in midgestation, but Mpc1 hypomorphs and tissue-specific deletion of Mpc1 presented as early perinatal lethality. The allelic series demonstrated that graded suppression of MPC resulted in dose-dependent metabolic and transcriptional changes. Steady-state metabolomics analysis of brain and liver from Mpc1 hypomorphic embryos identified compensatory changes in amino acid and lipid metabolism. Flux assays in Mpc1-deficient embryonic fibroblasts also reflected these changes, including a dramatic increase in mitochondrial alanine utilization. The mitochondrial alanine transaminase GPT2 was found to be necessary and sufficient for increased alanine flux upon MPC inhibition. These data show that impaired mitochondrial pyruvate transport results in biosynthetic deficiencies that can be mitigated in part by alternative anaplerotic substratesin utero.
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Johnston, I. A., H. Guderley, C. E. Franklin, T. Crockford, and C. Kamunde. "ARE MITOCHONDRIA SUBJECT TO EVOLUTIONARY TEMPERATURE ADAPTATION?" Journal of Experimental Biology 195, no. 1 (October 1, 1994): 293–306. http://dx.doi.org/10.1242/jeb.195.1.293.
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Thermal tolerance and the respiratory properties of isolated red muscle mitochondria were investigated in Oreochromis alcalicus grahami from the alkaline hot-springs, Lake Magadi, Kenya. Populations of O. a. grahami were resident in pools at 42.8 °C and migrated into water reaching temperatures of 44.8 °C for short periods. The maximum respiration rates of mitochondria with pyruvate as substrate were 217 and 284 natom O mg-1 mitochondrial protein min-1 at 37 °C and 42 °C, respectively (Q10=1.71). Fatty acyl carnitines (chain lengths C8, C12 and C16), malate and glutamate were oxidised at 70­80 % of the rate for pyruvate. In order to assess evolutionary temperature adaptation of maximum mitochondrial oxidative capacities, the rates of pyruvate and palmitoyl carnitine utilisation in red muscle mitochondria were measured from species living at other temperatures: Notothenia coriiceps from Antarctica (-1.5 to +1 °C); summer-caught Myoxocephalus scorpius from the North Sea (10­15 °C); and Oreochromis andersoni from African lakes and rivers (22­30 °C). State 3 respiration rates had Q10 values in the range 1.8­2.7. At the lower lethal temperature of O. andersoni (12.5 °C), isolated mitochondria utilised pyruvate at a similar rate to mitochondria from N. coriiceps at 2.5 °C (30 natom O mg-1 mitochondrial protein min-1). Rates of pyruvate oxidation by mitochondria from M. scorpius and N. coriiceps were similar and were higher at a given temperature than for O. andersoni. At their normal body temperature (-1.2 °C), mitochondria from the Antarctic fish oxidised pyruvate at 5.5 % and palmitoyl-dl-carnitine at 8.8 % of the rates of mitochondria from the hot-spring species at 42 °C. The results indicate only modest evolutionary adjustments in the maximal rates of mitochondrial respiration in fish living at different temperatures.
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Reyes, J., and D. J. Benos. "Specificity of gossypol uncoupling: a comparative study of liver and spermatogenic cells." American Journal of Physiology-Cell Physiology 254, no. 4 (April 1, 1988): C571—C576. http://dx.doi.org/10.1152/ajpcell.1988.254.4.c571.
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A comparative study of gossypol uncoupling of rat spermatogenic and liver cells shows that spermatogenic cells metabolizing pyruvate are two to three times more sensitive to gossypol uncoupling than either spermatogenic cells metabolizing glucose or liver cells metabolizing pyruvate or glucose. Direct measurements of in situ rat liver and spermatogenic cell mitochondrial respiration indicate that the pyruvate dependence of the gossypol uncoupling appears to be located in the spermatogenic cell mitochondria. A different type of mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone does not present the substrate-dependent uncoupling effect. This special interaction between spermatogenic cell pyruvate metabolism and gossypol uncoupling confers specificity to a bioenergetic model of gossypol action.
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Thomas, A. P., and R. M. Denton. "Use of toluene-permeabilized mitochondria to study the regulation of adipose tissue pyruvate dehydrogenase in situ. Further evidence that insulin acts through stimulation of pyruvate dehydrogenase phosphate phosphatase." Biochemical Journal 238, no. 1 (August 15, 1986): 93–101. http://dx.doi.org/10.1042/bj2380093.
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Rat epididymal-adipose-tissue mitochondria were made selectively permeable to small molecules without the loss of matrix enzymes by treating the mitochondria with toluene under controlled conditions. With this preparation the entire pyruvate dehydrogenase system was shown to be retained within the mitochondrial matrix and to retain its normal catalytic activity. By using dilute suspensions of these permeabilized mitochondria maintained in the cuvette of a spectrophotometer, it was possible to monitor changes of pyruvate dehydrogenase activity continuously while the activities of the interconverting kinase and phosphatase could be independently manipulated. Permeabilized mitochondria were prepared from control and insulin-treated adipose tissue, and the properties of both the pyruvate dehydrogenase kinase and the phosphatase were compared in situ. No difference in kinase activity was detected, but increases in phosphatase activity were observed in permeabilized mitochondria from insulin-treated tissue. Further studies showed that the main effect of insulin treatment was a decrease in the apparent Ka of the phosphatase for Mg2+, in agreement with earlier studies with mitochondria made permeable to Mg2+ by using the ionophore A23187 [Thomas, Diggle & Denton (1986) Biochem. J. 238, 83-91]. No effects of spermine were detected, although spermine diminishes the Ka of purified phosphatase preparations for Mg2+. Since effects of insulin on pyruvate dehydrogenase phosphatase activity are not evident in mitochondrial extracts, it is concluded that insulin may act by altering some high-Mr component which interacts with the pyruvate dehydrogenase system within intact or permeabilized mitochondria, but not when the mitochondrial membranes are disrupted.
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Grenell, Allison, Yekai Wang, Michelle Yam, Aditi Swarup, Tanya L. Dilan, Allison Hauer, Jonathan D. Linton, et al. "Loss of MPC1 reprograms retinal metabolism to impair visual function." Proceedings of the National Academy of Sciences 116, no. 9 (February 11, 2019): 3530–35. http://dx.doi.org/10.1073/pnas.1812941116.
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Glucose metabolism in vertebrate retinas is dominated by aerobic glycolysis (the “Warburg Effect”), which allows only a small fraction of glucose-derived pyruvate to enter mitochondria. Here, we report evidence that the small fraction of pyruvate in photoreceptors that does get oxidized by their mitochondria is required for visual function, photoreceptor structure and viability, normal neuron–glial interaction, and homeostasis of retinal metabolism. The mitochondrial pyruvate carrier (MPC) links glycolysis and mitochondrial metabolism. Retina-specific deletion of MPC1 results in progressive retinal degeneration and decline of visual function in both rod and cone photoreceptors. Using targeted-metabolomics and 13C tracers, we found that MPC1 is required for cytosolic reducing power maintenance, glutamine/glutamate metabolism, and flexibility in fuel utilization.
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Gao, Qun, and Michael S. Wolin. "Effects of hypoxia on relationships between cytosolic and mitochondrial NAD(P)H redox and superoxide generation in coronary arterial smooth muscle." American Journal of Physiology-Heart and Circulatory Physiology 295, no. 3 (September 2008): H978—H989. http://dx.doi.org/10.1152/ajpheart.00316.2008.
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Since controversy exists on how hypoxia influences vascular reactive oxygen species (ROS) generation, and our previous work provided evidence that it relaxes endothelium-denuded bovine coronary arteries (BCA) in a ROS-independent manner by promoting cytosolic NADPH oxidation, we examined how hypoxia alters relationships between cytosolic and mitochondrial NAD(P)H redox and superoxide generation in BCA. Methods were developed to image and interpret the effects of hypoxia on NAD(P)H redox based on its autofluorescence in the cytosolic, mitochondrial, and nuclear regions of smooth muscle cells isolated from BCA. Aspects of anaerobic glycolysis and cytosolic NADH redox in BCA were assessed from measurements of lactate and pyruvate. Imaging changes in mitosox and dehydroethidium fluorescence were used to detect changes in mitochondrial and cytosolic-nuclear superoxide, respectively. Hypoxia appeared to increase mitochondrial and decrease cytosolic-nuclear superoxide under conditions associated with increased cytosolic NADH (lactate/pyruvate), mitochondrial NAD(P)H, and hyperpolarization of mitochondria detected by tetramethylrhodamine methyl-ester perchlorate fluorescence. Rotenone appeared to increase mitochondrial NAD(P)H and superoxide, suggesting hypoxia could increase superoxide generation by complex I. However, hypoxia decreased mitochondrial superoxide in the presence of contraction to 30 mM KCl, associated with decreased mitochondrial NAD(P)H. Thus, while hypoxia augments NAD(P)H redox associated with increased mitochondrial superoxide, contraction with KCl reverses these effects of hypoxia on mitochondrial superoxide, suggesting mitochondrial ROS increases do not mediate hypoxic relaxation in BCA. Since hypoxia lowers pyruvate, and pyruvate inhibits hypoxia-elicited relaxation and NADPH oxidation in BCA, mitochondrial control of pyruvate metabolism associated with cytosolic NADPH redox regulation could contribute to sensing hypoxia.
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CIVELEK, Vildan N., Jude T. DEENEY, Nicholas J. SHALOSKY, Keith TORNHEIM, Richard G. HANSFORD, Marc PRENTKI, and Barbara E. CORKEY. "Regulation of pancreatic β-cell mitochondrial metabolism: influence of Ca2+, substrate and ADP." Biochemical Journal 318, no. 2 (September 1, 1996): 615–21. http://dx.doi.org/10.1042/bj3180615.
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To gain insight into the regulation of pancreatic β-cell mitochondrial metabolism, the direct effects on respiration of different mitochondrial substrates, variations in the ATP/ADP ratio and free Ca2+ were examined using isolated mitochondria and permeabilized clonal pancreatic β-cells (HIT). Respiration from pyruvate was high and not influenced by Ca2+ in State 3 or under various redox states and fixed values of the ATP/ADP ratio; nevertheless, high Ca2+ elevated pyridine nucleotide fluorescence, indicating activation of pyruvate dehydrogenase by Ca2+. Furthermore, in the presence of pyruvate, elevated Ca2+ stimulated CO2 production from pyruvate, increased citrate production and efflux from the mitochondria and inhibited CO2 production from palmitate. The latter observation suggests that β-cell fatty acid oxidation is not regulated exclusively by malonyl-CoA but also by the mitochondrial redox state. α-Glycerophosphate (α-GP) oxidation was Ca2+-dependent with a half-maximal rate observed at around 300 nM Ca2+. We have recently demonstrated that increases in respiration precede increases in Ca2+ in glucose-stimulated clonal pancreatic β-cells (HIT), indicating that Ca2+ is not responsible for the initial stimulation of respiration [Civelek, Deeney, Kubik, Schultz, Tornheim and Corkey (1996) Biochem. J. 315, 1015–1019]. It is suggested that respiration is stimulated by increased substrate (α-GP and pyruvate) supply together with oscillatory increases in ADP [Nilsson, Schultz, Berggren, Corkey and Tornheim (1996) Biochem. J. 314, 91–94]. The rise in Ca2+, which in itself may not significantly increase net respiration, could have the important functions of (1) activating the α-GP shuttle, to maintain an oxidized cytosol and high glycolytic flux; (2) activating pyruvate dehydrogenase, and indirectly pyruvate carboxylase, to sustain production of citrate and hence the putative signal coupling factors, malonyl-CoA and acyl-CoA; and (3) increasing mitochondrial redox state to implement the switch from fatty acid to pyruvate oxidation.
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Koh, Eunjin, Young Kyung Kim, Daye Shin, and Kyung-Sup Kim. "MPC1 is essential for PGC-1α-induced mitochondrial respiration and biogenesis." Biochemical Journal 475, no. 10 (May 18, 2018): 1687–99. http://dx.doi.org/10.1042/bcj20170967.
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Mitochondrial pyruvate carrier (MPC), which is essential for mitochondrial pyruvate usage, mediates the transport of cytosolic pyruvate into mitochondria. Low MPC expression is associated with various cancers, and functionally associated with glycolytic metabolism and stemness. However, the mechanism by which MPC expression is regulated is largely unknown. In this study, we showed that MPC1 is down-regulated in human renal cell carcinoma (RCC) due to strong suppression of peroxisome proliferator-activated receptor-gamma co-activator (PGC)-1 alpha (PGC-1α). We also demonstrated that overexpression of PGC-1α stimulates MPC1 transcription, while depletion of PGC-1α by siRNA suppresses MPC expression. We found that PGC-1α interacts with estrogen-related receptor-alpha (ERR-α) and recruits it to the ERR-α response element motif located in the proximal MPC1 promoter, resulting in efficient activation of MPC1 expression. Furthermore, the MPC inhibitor, UK5099, blocked PGC-1α-induced pyruvate-dependent mitochondrial oxygen consumption. Taken together, our results suggest that MPC1 is a novel target gene of PGC-1α. In addition, low expression of PGC-1α in human RCC might contribute to the reduced expression of MPC, resulting in impaired mitochondrial respiratory capacity in RCC by limiting the transport of pyruvate into the mitochondrial matrix.
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Messer, Jeffrey I., Matthew R. Jackman, and Wayne T. Willis. "Pyruvate and citric acid cycle carbon requirements in isolated skeletal muscle mitochondria." American Journal of Physiology-Cell Physiology 286, no. 3 (March 2004): C565—C572. http://dx.doi.org/10.1152/ajpcell.00146.2003.
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Carbohydrate depletion precipitates fatigue in skeletal muscle, but, because pyruvate provides both acetyl-CoA for mainline oxidation and anaplerotic carbon to the citric acid cycle (CAC), the mechanism remains obscure. Thus pyruvate and CAC kinetic parameters were independently quantified in mitochondria isolated from rat mixed skeletal muscle. Mitochondrial oxygen consumption rate ( Jo) was measured polarographically while either pyruvate or malate was added stepwise in the presence of a saturating concentration of the other substrate. These substrate titrations were carried out across a physiological range of fixed extramitochondrial ATP free energy states (ΔGP), established with a creatine kinase energy clamp, and also at saturating [ADP]. The apparent Km,malate for mitochondrial Jo ranged from 21 to 32 μM, and the apparent Km,pyruvate ranged from 12 to 26 μM, with both substrate Km values increasing as ΔGP declined. Vmax for both substrates also increased as ΔGP fell, reflecting thermodynamic control of Jo. Reported in vivo skeletal muscle [malate] are >10-fold greater than the Km,malate determined in this study. In marked contrast, the Km,pyruvate determined is near the [pyruvate] reported in muscle approaching exhaustion associated with glycogen depletion. When data were evaluated in the context of a linear thermodynamic force-flow (ΔGP- Jo) relationship, the ΔGP- Jo slope was essentially insensitive to changes in [malate] in the range observed in vivo but decreased markedly with declining [pyruvate] across the physiological range. Mitochondrial respiration is particularly sensitive to variations in [pyruvate] in the physiological range. In contrast, physiological [malate] exerts very little, if any, influence on mitochondrial pyruvate oxidation measured in vitro.
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Willis, W. T., M. R. Jackman, M. E. Bizeau, M. J. Pagliassotti, and J. R. Hazel. "Hyperthermia impairs liver mitochondrial function in vitro." American Journal of Physiology-Regulatory, Integrative and Comparative Physiology 278, no. 5 (May 1, 2000): R1240—R1246. http://dx.doi.org/10.1152/ajpregu.2000.278.5.r1240.
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The effects of temperature on the relationships among the rates of pyruvate carboxylation, O2 uptake ( J o), oxidative phosphorylation ( J p), and the free energy of ATP hydrolysis ( G p) were studied in liver mitochondria isolated from 250-g female rats. Pyruvate carboxylation was evaluated at 37, 40, and 43°C. In disrupted mitochondria, pyruvate carboxylase maximal reaction velocity increased from 37 to 43°C with an apparent Q10 of 2.25. A reduction in ATP/ADP ratio decreased enzyme activity at all three temperatures. In contrast, in intact mitochondria, increasing temperature failed to increase pyruvate carboxylation (malate + citrate accumulation) but did result in increased J o and decreased extramitochondrial G p. J p was studied in respiring mitochondria at 37 and 43°C at various fractions of state 3 respiration, elicited with a glucose + hexokinase ADP-regenerating system. The relationship between J o and G p was similar at both temperatures. However, hyperthermia (43°C) reduced the J p/ J o ratio, resulting in lower G p for a given J p. Fluorescent measurements of membrane phospholipid polarization revealed a transition in membrane order between 40 and 43°C, a finding consistent with increased membrane proton conductance. It is concluded that hyperthermia augments nonspecific proton leaking across the inner mitochondrial membrane, and the resultant degraded energy state offsets temperature stimulation of pyruvate carboxylase. As a consequence, at high temperatures approaching 43°C, the pyruvate carboxylation rate of intact liver mitochondria may fail to exhibit a Q10effect.
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DÜFER, Martina, Peter KRIPPEIT-DREWS, Linas BUNTINAS, Detlef SIEMEN, and Gisela DREWS. "Methyl pyruvate stimulates pancreatic β-cells by a direct effect on KATP channels, and not as a mitochondrial substrate." Biochemical Journal 368, no. 3 (December 15, 2002): 817–25. http://dx.doi.org/10.1042/bj20020657.
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In pancreatic β-cells, methyl pyruvate is a potent secretagogue and is widely used to study stimulus—secretion coupling. In contrast with pyruvate, which barely stimulates insulin secretion, methyl pyruvate was suggested to act as an effective mitochondrial substrate. We show that methyl pyruvate elicited electrical activity in the presence of 0.5mM glucose, in contrast with pyruvate. Accordingly, methyl pyruvate increased the cytosolic free Ca2+ concentration after an initial decrease, similar to glucose. The initial decrease was inhibited by thapsigargin, suggesting that methyl pyruvate stimulates ATP production. This assumption is supported by the observation that methyl pyruvate hyperpolarized the mitochondrial membrane potential, similar to glucose. However, in contrast with glucose, methyl pyruvate even slightly decreased NAD(P)H autofluorescence and did not influence ATP production or the ATP/ADP ratio. This observation questions the suggestion that methyl pyruvate acts as a powerful mitochondrial substrate. The finding that methyl pyruvate directly inhibited a cation current across the inner membrane of Jurkat T-lymphocyte mitochondria suggests that this metabolite may increase ATP production in β-cells by activating the respiratory chains without providing reduction equivalents. We conclude that this mechanism may account for a slight and transient increase in ATP production. We further show that methyl pyruvate inhibited the KATP current measured in the standard whole-cell configuration, an effect that was at least partly antagonized by diazoxide. Accordingly, single-channel currents in inside-out patches were blocked by methyl pyruvate. We conclude that inhibition of KATP channels, and not activation of metabolism, mediates the induction of electrical activity in pancreatic β-cells by methyl pyruvate.
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Reel, Jessica Morgan, Hazzar M. Abysalamah, and Christopher R. Lupfer. "Sodium pyruvate reduces immune signaling during influenza A virus infection in macrophages." Journal of Immunology 204, no. 1_Supplement (May 1, 2020): 93.20. http://dx.doi.org/10.4049/jimmunol.204.supp.93.20.
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Abstract Pyruvate is a key metabolite for energy synthesis. After the production of pyruvate through glycolysis, the molecule is shuttled into the mitochondria. Once there, it is modified for use in the TCA cycle and the energy derived from pyruvate is eventually converted into ATP. While pyruvate is a key metabolite, it also appears to have anti-inflammatory properties. In models of sterile inflammation, like ischemia, pyruvate limits inflammation. We observed that infecting murine bone marrow derived macrophages (BMDM) with influenza A virus (IAV) and treating those macrophages with sodium pyruvate results in lower inflammasome activation and reactive oxygen species (ROS) production. However, this was specific to IAV infection as inflammasome activation and ROS were not affected by sodium pyruvate during E. coli infection or lipopolysaccharide (LPS) and ATP treatment of BMDMs. Our results show that IAV induces a potent and unique metabolic reprograming of infected cells. The addition of sodium pyruvate facilitates ATP production, which correlates with less mitochondrial damage and reduced inflammasome activation. Thus, pyruvate deserves additional examination as an anti-inflammatory treatment in diseases where mitochondrial metabolic stress is a factor.
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Khan, Dilshad H., Michael Mullokandov, Yan Wu, Veronique Voisin, Marcela Gronda, Rose Hurren, Xiaoming Wang, et al. "Mitochondrial carrier homolog 2 is necessary for AML survival." Blood 136, no. 1 (July 2, 2020): 81–92. http://dx.doi.org/10.1182/blood.2019000106.
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Abstract Through a clustered regularly insterspaced short palindromic repeats (CRISPR) screen to identify mitochondrial genes necessary for the growth of acute myeloid leukemia (AML) cells, we identified the mitochondrial outer membrane protein mitochondrial carrier homolog 2 (MTCH2). In AML, knockdown of MTCH2 decreased growth, reduced engraftment potential of stem cells, and induced differentiation. Inhibiting MTCH2 in AML cells increased nuclear pyruvate and pyruvate dehydrogenase (PDH), which induced histone acetylation and subsequently promoted the differentiation of AML cells. Thus, we have defined a new mechanism by which mitochondria and metabolism regulate AML stem cells and gene expression.
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Johnston, J. D., and M. D. Brand. "Stimulation of the respiration rate of rat liver mitochondria by sub-micromolar concentrations of extramitochondrial Ca2+." Biochemical Journal 245, no. 1 (July 1, 1987): 217–22. http://dx.doi.org/10.1042/bj2450217.
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1. The respiration rate of rat liver mitochondria was stimulated by up to 70% when the extramitochondrial Ca2+ concentration was raised from 103 to 820 nM. This occurred when pyruvate, 2-oxoglutarate, or threo-(Ds)-isocitrate was employed as substrate, but not when succinate was used. 2. Ruthenium Red prevented the stimulation of mitochondrial respiration by extramitochondrial Ca2+, showing that the effect required Ca2+ uptake into the mitochondrial matrix. 3. Starvation of rats for 48 h abolished the stimulation of mitochondrial respiration by extramitochondrial Ca2+ when pyruvate was used as substrate, but did not affect the stimulation of 2-oxoglutarate oxidation by extramitochondrial Ca2+. 4. Our findings are in accord with proposals that oxidative metabolism in liver mitochondria may be stimulated by Ca2+ activation of intramitochondrial dehydrogenases.
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McCommis, Kyle S., and Brian N. Finck. "Mitochondrial pyruvate transport: a historical perspective and future research directions." Biochemical Journal 466, no. 3 (March 6, 2015): 443–54. http://dx.doi.org/10.1042/bj20141171.
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Pyruvate is the end-product of glycolysis, a major substrate for oxidative metabolism, and a branching point for glucose, lactate, fatty acid and amino acid synthesis. The mitochondrial enzymes that metabolize pyruvate are physically separated from cytosolic pyruvate pools and rely on a membrane transport system to shuttle pyruvate across the impermeable inner mitochondrial membrane (IMM). Despite long-standing acceptance that transport of pyruvate into the mitochondrial matrix by a carrier-mediated process is required for the bulk of its metabolism, it has taken almost 40 years to determine the molecular identity of an IMM pyruvate carrier. Our current understanding is that two proteins, mitochondrial pyruvate carriers MPC1 and MPC2, form a hetero-oligomeric complex in the IMM to facilitate pyruvate transport. This step is required for mitochondrial pyruvate oxidation and carboxylation–critical reactions in intermediary metabolism that are dysregulated in several common diseases. The identification of these transporter constituents opens the door to the identification of novel compounds that modulate MPC activity, with potential utility for treating diabetes, cardiovascular disease, cancer, neurodegenerative diseases, and other common causes of morbidity and mortality. The purpose of the present review is to detail the historical, current and future research investigations concerning mitochondrial pyruvate transport, and discuss the possible consequences of altered pyruvate transport in various metabolic tissues.
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Le, Catherine H., Lindsay G. Benage, Kalyn S. Specht, Lance C. Li Puma, Christopher M. Mulligan, Adam L. Heuberger, Jessica E. Prenni, et al. "Tafazzin deficiency impairs CoA-dependent oxidative metabolism in cardiac mitochondria." Journal of Biological Chemistry 295, no. 35 (July 14, 2020): 12485–97. http://dx.doi.org/10.1074/jbc.ra119.011229.
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Barth syndrome is a mitochondrial myopathy resulting from mutations in the tafazzin (TAZ) gene encoding a phospholipid transacylase required for cardiolipin remodeling. Cardiolipin is a phospholipid of the inner mitochondrial membrane essential for the function of numerous mitochondrial proteins and processes. However, it is unclear how tafazzin deficiency impacts cardiac mitochondrial metabolism. To address this question while avoiding confounding effects of cardiomyopathy on mitochondrial phenotype, we utilized Taz-shRNA knockdown (TazKD) mice, which exhibit defective cardiolipin remodeling and respiratory supercomplex instability characteristic of human Barth syndrome but normal cardiac function into adulthood. Consistent with previous reports from other models, mitochondrial H2O2 emission and oxidative damage were greater in TazKD than in wild-type (WT) hearts, but there were no differences in oxidative phosphorylation coupling efficiency or membrane potential. Fatty acid and pyruvate oxidation capacities were 40–60% lower in TazKD mitochondria, but an up-regulation of glutamate oxidation supported respiration rates approximating those with pyruvate and palmitoylcarnitine in WT. Deficiencies in mitochondrial CoA and shifts in the cardiac acyl-CoA profile paralleled changes in fatty acid oxidation enzymes and acyl-CoA thioesterases, suggesting limitations of CoA availability or “trapping” in TazKD mitochondrial metabolism. Incubation of TazKD mitochondria with exogenous CoA partially rescued pyruvate and palmitoylcarnitine oxidation capacities, implicating dysregulation of CoA-dependent intermediary metabolism rather than respiratory chain defects in the bioenergetic impacts of tafazzin deficiency. These findings support links among cardiolipin abnormalities, respiratory supercomplex instability, and mitochondrial oxidant production and shed new light on the distinct metabolic consequences of tafazzin deficiency in the mammalian heart.
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Bricker, Daniel K., Eric B. Taylor, John C. Schell, Thomas Orsak, Audrey Boutron, Yu-Chan Chen, James E. Cox, et al. "A Mitochondrial Pyruvate Carrier Required for Pyruvate Uptake in Yeast,Drosophila, and Humans." Science 337, no. 6090 (May 24, 2012): 96–100. http://dx.doi.org/10.1126/science.1218099.
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Pyruvate constitutes a critical branch point in cellular carbon metabolism. We have identified two proteins, Mpc1 and Mpc2, as essential for mitochondrial pyruvate transport in yeast,Drosophila, and humans. Mpc1 and Mpc2 associate to form an ~150-kilodalton complex in the inner mitochondrial membrane. Yeast andDrosophilamutants lackingMPC1display impaired pyruvate metabolism, with an accumulation of upstream metabolites and a depletion of tricarboxylic acid cycle intermediates. Loss of yeast Mpc1 results in defective mitochondrial pyruvate uptake, and silencing ofMPC1orMPC2in mammalian cells impairs pyruvate oxidation. A point mutation inMPC1provides resistance to a known inhibitor of the mitochondrial pyruvate carrier. Human genetic studies of three families with children suffering from lactic acidosis and hyperpyruvatemia revealed a causal locus that mapped toMPC1, changing single amino acids that are conserved throughout eukaryotes. These data demonstrate that Mpc1 and Mpc2 form an essential part of the mitochondrial pyruvate carrier.
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Kümmel, Ladislav. "Mitochondrial pyruvate carrier—A possible link between gluconeogenesis and ketogenesis in the liver." Bioscience Reports 7, no. 7 (July 1, 1987): 593–97. http://dx.doi.org/10.1007/bf01119777.
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Effects of various ketogenic substrates on gluconeogenesis from lactate or alanine were compared. The results suggest that, in intact liver cells, cytoplasmic pyruvate is transported into mitochondria in exchange for intramitochondrially generated acetoacetate. An interrelationship between gluconeogenesis and ketogenesis may thus exist in the liver at the level of mitochondrial pyruvate carrier.
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Wolf, Christina, Rahel Zimmermann, Osamah Thaher, Diones Bueno, Verena Wüllner, Michael K. E. Schäfer, Philipp Albrecht, and Axel Methner. "The Charcot–Marie Tooth Disease Mutation R94Q in MFN2 Decreases ATP Production but Increases Mitochondrial Respiration under Conditions of Mild Oxidative Stress." Cells 8, no. 10 (October 21, 2019): 1289. http://dx.doi.org/10.3390/cells8101289.
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Charcot–Marie tooth disease is a hereditary polyneuropathy caused by mutations in Mitofusin-2 (MFN2), a GTPase in the outer mitochondrial membrane involved in the regulation of mitochondrial fusion and bioenergetics. Autosomal-dominant inheritance of a R94Q mutation in MFN2 causes the axonal subtype 2A2A which is characterized by early onset and progressive atrophy of distal muscles caused by motoneuronal degeneration. Here, we studied mitochondrial shape, respiration, cytosolic, and mitochondrial ATP content as well as mitochondrial quality control in MFN2-deficient fibroblasts stably expressing wildtype or R94Q MFN2. Under normal culture conditions, R94Q cells had slightly more fragmented mitochondria but a similar mitochondrial oxygen consumption, membrane potential, and ATP production as wildtype cells. However, when inducing mild oxidative stress 24 h before analysis using 100 µM hydrogen peroxide, R94Q cells exhibited significantly increased respiration but decreased mitochondrial ATP production. This was accompanied by increased glucose uptake and an up-regulation of hexokinase 1 and pyruvate kinase M2, suggesting increased pyruvate shuttling into mitochondria. Interestingly, these changes coincided with decreased levels of PINK1/Parkin-mediated mitophagy in R94Q cells. We conclude that mitochondria harboring the disease-causing R94Q mutation in MFN2 are more susceptible to oxidative stress, which causes uncoupling of respiration and ATP production possibly by a less efficient mitochondrial quality control.
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Szibor, Marten, Zemfira Gizatullina, Timur Gainutdinov, Thomas Endres, Grazyna Debska-Vielhaber, Matthias Kunz, Niki Karavasili, et al. "Cytosolic, but not matrix, calcium is essential for adjustment of mitochondrial pyruvate supply." Journal of Biological Chemistry 295, no. 14 (February 24, 2020): 4383–97. http://dx.doi.org/10.1074/jbc.ra119.011902.
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Mitochondrial oxidative phosphorylation (OXPHOS) and cellular workload are tightly balanced by the key cellular regulator, calcium (Ca2+). Current models assume that cytosolic Ca2+ regulates workload and that mitochondrial Ca2+ uptake precedes activation of matrix dehydrogenases, thereby matching OXPHOS substrate supply to ATP demand. Surprisingly, knockout (KO) of the mitochondrial Ca2+ uniporter (MCU) in mice results in only minimal phenotypic changes and does not alter OXPHOS. This implies that adaptive activation of mitochondrial dehydrogenases by intramitochondrial Ca2+ cannot be the exclusive mechanism for OXPHOS control. We hypothesized that cytosolic Ca2+, but not mitochondrial matrix Ca2+, may adapt OXPHOS to workload by adjusting the rate of pyruvate supply from the cytosol to the mitochondria. Here, we studied the role of malate-aspartate shuttle (MAS)-dependent substrate supply in OXPHOS responses to changing Ca2+ concentrations in isolated brain and heart mitochondria, synaptosomes, fibroblasts, and thymocytes from WT and MCU KO mice and the isolated working rat heart. Our results indicate that extramitochondrial Ca2+ controls up to 85% of maximal pyruvate-driven OXPHOS rates, mediated by the activity of the complete MAS, and that intramitochondrial Ca2+ accounts for the remaining 15%. Of note, the complete MAS, as applied here, included besides its classical NADH oxidation reaction the generation of cytosolic pyruvate. Part of this largely neglected mechanism has previously been described as the “mitochondrial gas pedal.” Its implementation into OXPHOS control models integrates seemingly contradictory results and warrants a critical reappraisal of metabolic control mechanisms in health and disease.
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Sharma, Pushpa, Kane T. Walsh, Kimberly A. Kerr-Knott, John E. Karaian, and Paul D. Mongan. "Pyruvate Modulates Hepatic Mitochondrial Functions and Reduces Apoptosis Indicators during Hemorrhagic Shock in Rats." Anesthesiology 103, no. 1 (July 1, 2005): 65–73. http://dx.doi.org/10.1097/00000542-200507000-00013.
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Background Dysfunctional mitochondria have been widely accepted as one of the key targets and a mediator of secondary cell injury and organ failure during hemorrhagic shock (HS). The liver is known to be the first organ to display the signs of injury during HS. This report describes experiments to determine whether modulation of hepatic mitochondrial dysfunctions by pharmacologic agents could prevent liver injury in rats subjected to HS. Methods In this study, Sprague-Dawley rats were either treated as controls or subjected to computer-controlled arterial hemorrhage (40 mmHg) for 60 min followed by resuscitation with hypertonic saline, hypertonic beta-hydroxybutyrate, or hypertonic sodium pyruvate for the next 60 min before death. During the course of the experiment, animals were continuously monitored for hemodynamic and metabolic parameters. At the end of the experiment, the liver was excised and examined for oxidative injury, mitochondrial functions, expression of nitric oxide synthase, and indicators of apoptosis. Results In comparison to hypertonic saline and hypertonic beta-hydroxybutyrate, pyruvate significantly protected the liver from oxidative injury, prevented the up-regulation of nitric oxide synthase, inhibited pyruvate dehydrogenase deactivation, and improved cellular energy charge and mitochondrial functions. In addition, pyruvate also reduced cleavage of poly-adenosine diphosphate ribose polymerase by preventing leakage of mitochondrial cytochrome c in the liver of HS animals. Conclusions These data suggest that modulation of mitochondrial metabolic functions is likely to be one of the important mechanisms by which pyruvate exerts its protective effects on the liver during HS and resuscitation in rats.
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Lai, James C. K. "Oxidative metabolism in neuronal and non-neuronal mitochondria." Canadian Journal of Physiology and Pharmacology 70, S1 (May 15, 1992): S130—S137. http://dx.doi.org/10.1139/y92-254.
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Methodological advances have allowed the isolation of two populations of synaptic (SM and SM2) and two populations of nonsynaptic (A and B) mitochondria from rat forebrain. All four populations of brain mitochondria are metabolically active and essentially free from nonmitochondrial contaminants. They (SM, SM2, A, and B) can oxidize a variety of substrates; the best substrate is pyruvate. With pyruvate as the substrate, the respiratory control ratios (i.e., state 3/state 4) in all four populations are routinely >6. Results from numerous enzyme activity measurements provide strong support for the hypothesis that brain mitochondria are very heterogeneous with respect to their enzyme contents and that the enzymatic activities in a particular population of mitochondria, be they synaptic or nonsynaptic, differ from those in another population of mitochondria derived from either the same or another brain region. The major methodological advances in brain mitochondrial isolation greatly facilitate metabolic studies. For example, we have demonstrated that the K+ stimulation of brain mitochondrial pyruvate oxidation is mediated through a K+-induced elevation of the activation state of the pyruvate dehydrogenase complex and the K+ stimulation of the flux through the pyruvate dehydrogenase complex. Our previous and ongoing studies using primary cultures of hypothalamic neurons and astrocytes are consistent with the proposal that brain cells are heterogeneous with respect to their capabilities in energy metabolism. I can envisage that in the not-so-distant future, one could adapt these preparations of cells as the starting material for the isolation of mitochondria of known cellular origin for metabolic studies.Key words: heterogeneity of brain mitochondria, regulation of intermediary metabolism.
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Wilson, Leanne, Qing Yang, Joseph D. Szustakowski, P. Scott Gullicksen, and Reza Halse. "Pyruvate induces mitochondrial biogenesis by a PGC-1 α-independent mechanism." American Journal of Physiology-Cell Physiology 292, no. 5 (May 2007): C1599—C1605. http://dx.doi.org/10.1152/ajpcell.00428.2006.
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Oxidative cells increase mitochondrial mass in response to stimuli such as changes in energy demand or cellular differentiation. This plasticity enables the cell to adapt dynamically to achieve the necessary oxidative capacity. However, the pathways involved in triggering mitochondrial biogenesis are poorly defined. The present study examines the impact of altering energy provision on mitochondrial biogenesis in muscle cells. C2C12 myoblasts were chronically treated with supraphysiological levels of sodium pyruvate for 72 h. Treated cells exhibited increased mitochondrial protein expression, basal respiratory rate, and maximal oxidative capacity. The increase in mitochondrial biogenesis was independent of increases in peroxisomal proliferator activator receptor-γ coactivator-1α (PGC-1α) and PGC-1β mRNA expression. To further assess whether PGC-1α expression was necessary for pyruvate action, cells were infected with adenovirus containing shRNA for PGC-1α before treatment with pyruvate. Despite a 70% reduction in PGC-1α mRNA, the effect of pyruvate was preserved. Furthermore, pyruvate induced mitochondrial biogenesis in primary myoblasts from PGC-1α null mice. These data suggest that regulation of mitochondrial biogenesis by pyruvate in myoblasts is independent of PGC-1α, suggesting the existence of a novel energy-sensing pathway regulating oxidative capacity.
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Bilonoha, O., B. O. Manko, and V. Manko. "Effects of insulin on adaptive capacity of rat pancreatic acinar cells mitochondria." Visnyk of Lviv University. Biological series, no. 83 (December 25, 2020): 24–30. http://dx.doi.org/10.30970/vlubs.2020.83.03.
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Insulin increases the basal and agonist-stimulated secretion of pancreatic acinar cells, which leads to increase of energy demand and requires sufficient oxidative substrates supply. Cholecystokinin substantially increases the respiration rate of pancreatic acinar cells upon pyruvate oxidation. However, it is not clear how insulin affects mitochondrial oxidative processes at rest and upon secretory stimulation. Experiments were carried out on male Wistar rats (250–300 g) kept on standard diet. Animals were fasted 12 h before the experiment. Pancreatic acini were isolated with collagenase. Basal and FCCP-stimulated respiration of rat pancreatic acini was measured with Clark electrode. Adaptive capacity of mitochondria was assessed by the maximal rate of uncoupled respiration. Statistical significance (P) of differenced between the means was assessed either with a paired t-test or with repeated measures two-way ANOVA and post-hoc Turkey test. Adaptive capacity of pancreatic acinar mitochondria was significantly higher when pyruvate (2 mM) was used as oxidative substrate comparing with glucose (10 mM). Incubation with insulin (100 nM) for 20 minutes elevated the basal respiration and adaptive capacity of pancreatic acinar mitochondria upon glucose, but not pyruvate, oxidation. Cholecystokinin (0.1 nM, 30 min) stimulated the rate of basal and maximal uncoupled respiration of acinar cells upon pyruvate oxidation, but insulin completely negated this increase of mitochondrial adaptive capacity. Thus, insulin increases the glucose oxidation in pancreatic acinar cells at resting state, but suppresses pyruvate oxidation upon secretory stimulation with cholecystokinin. The mechanisms of insulin action of pyruvate metabolism in pancreatic acinar cells require further elucidation.
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Kim, Yong Kyung, Lori Sussel, and Howard W. Davidson. "Inherent Beta Cell Dysfunction Contributes to Autoimmune Susceptibility." Biomolecules 11, no. 4 (March 30, 2021): 512. http://dx.doi.org/10.3390/biom11040512.
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The pancreatic beta cell is a highly specialized cell type whose primary function is to secrete insulin in response to nutrients to maintain glucose homeostasis in the body. As such, the beta cell has developed unique metabolic characteristics to achieve functionality; in healthy beta cells, the majority of glucose-derived carbons are oxidized and enter the mitochondria in the form of pyruvate. The pyruvate is subsequently metabolized to induce mitochondrial ATP and trigger the downstream insulin secretion response. Thus, in beta cells, mitochondria play a pivotal role in regulating glucose stimulated insulin secretion (GSIS). In type 2 diabetes (T2D), mitochondrial impairment has been shown to play an important role in beta cell dysfunction and loss. In type 1 diabetes (T1D), autoimmunity is the primary trigger of beta cell loss; however, there is accumulating evidence that intrinsic mitochondrial defects could contribute to beta cell susceptibility during proinflammatory conditions. Furthermore, there is speculation that dysfunctional mitochondrial responses could contribute to the formation of autoantigens. In this review, we provide an overview of mitochondrial function in the beta cells, and discuss potential mechanisms by which mitochondrial dysfunction may contribute to T1D pathogenesis.
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Padua, Rodolfo A., Kyle T. Baron, Bhaskar Thyagarajan, Colin Campbell, and Stanley A. Thayer. "Reduced Ca2+ uptake by mitochondria in pyruvate dehydrogenase-deficient human diploid fibroblasts." American Journal of Physiology-Cell Physiology 274, no. 3 (March 1, 1998): C615—C622. http://dx.doi.org/10.1152/ajpcell.1998.274.3.c615.
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Physiological and pathological Ca2+ loads are thought to be taken up by mitochondria via a process dependent on aerobic metabolism. We sought to determine whether human diploid fibroblasts from a patient with an inherited defect in pyruvate dehydrogenase (PDH) exhibit a decreased ability to sequester cytosolic Ca2+ into mitochondria. Mobilization of Ca2+ stores with bradykinin (BK) increased the cytosolic Ca2+ concentration ([Ca2+]c) to comparable levels in control and PDH-deficient fibroblasts. In normal fibroblasts transfected with plasmid DNA encoding mitochondrion-targeted apoaequorin, BK elicited an increase in Ca2+-dependent aequorin luminescence corresponding to an increase in the mitochondrial Ca2+ concentration ([Ca2+]mt) of 2.0 ± 0.2 μM. The mitochondrial uncoupling agent carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone blocked the BK-induced [Ca2+]mtincrease, although it did not affect the [Ca2+]ctransient. Basal [Ca2+]cand [Ca2+]mtin control and PDH-deficient cells were similar. However, confocal imaging of the potential-sensitive dye JC-1 indicated that the percentage of highly polarized mitochondria was reduced from 30 ± 1% in normal cells to 19 ± 2% in the PDH-deficient fibroblasts. BK-elicited [Ca2+]mttransients in PDH-deficient cells were reduced to 4% of control, indicating that PDH-deficient mitochondria have a decreased ability to take up cytosolic Ca2+. Thus cells with compromised aerobic metabolism have a reduced capacity to sequester Ca2+.
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Warren, Blair E., Phing-How Lou, Eliana Lucchinetti, Liyan Zhang, Alexander S. Clanachan, Andreas Affolter, Martin Hersberger, Michael Zaugg, and Hélène Lemieux. "Early mitochondrial dysfunction in glycolytic muscle, but not oxidative muscle, of the fructose-fed insulin-resistant rat." American Journal of Physiology-Endocrinology and Metabolism 306, no. 6 (March 15, 2014): E658—E667. http://dx.doi.org/10.1152/ajpendo.00511.2013.
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Although evidence that type 2 diabetes mellitus (T2DM) is accompanied by mitochondrial dysfunction in skeletal muscle has been accumulating, a causal link between mitochondrial dysfunction and the pathogenesis of the disease remains unclear. Our study focuses on an early stage of the disease to determine whether mitochondrial dysfunction contributes to the development of T2DM. The fructose-fed (FF) rat was used as an animal model of early T2DM. Mitochondrial respiration and acylcarnitine species were measured in oxidative (soleus) and glycolytic [extensor digitorum longus (EDL)] muscle. Although FF rats displayed characteristic signs of T2DM, including hyperglycemia, hyperinsulinemia, and hypertriglyceridemia, mitochondrial content was preserved in both muscles from FF rats. The EDL muscle had reduced complex I and complex I and II respiration in the presence of pyruvate but not glutamate. The decrease in pyruvate-supported respiration was due to a decrease in pyruvate dehydrogenase activity. Accumulation of C14:1 and C14:2 acylcarnitine species and a decrease in respiration supported by long-chain acylcarnitines but not acetylcarnitine indicated dysfunctional β-oxidation in the EDL muscle. In contrast, the soleus muscle showed preserved mitochondrial respiration, pyruvate dehydrogenase activity, and increased fatty acid oxidation, as evidenced by overall reduced acylcarnitine levels. Aconitase activity, a sensitive index of reactive oxygen species production in mitochondria, was reduced exclusively in EDL muscle, which showed lower levels of the antioxidant enzymes thioredoxin reductase and glutathione peroxidase. Here, we show that the glycolytic EDL muscle is more prone to an imbalance between energy supply and oxidation caused by insulin resistance than the oxidative soleus muscle.
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Li, Aiyun, Qun Liu, Qiang Li, Baolin Liu, Yang Yang, and Ning Zhang. "Berberine Reduces Pyruvate-driven Hepatic Glucose Production by Limiting Mitochondrial Import of Pyruvate through Mitochondrial Pyruvate Carrier 1." EBioMedicine 34 (August 2018): 243–55. http://dx.doi.org/10.1016/j.ebiom.2018.07.039.
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Hagve, Martin, Petter Fosse Gjessing, Ole Martin Fuskevåg, Terje S. Larsen, and Øivind Irtun. "Skeletal muscle mitochondria exhibit decreased pyruvate oxidation capacity and increased ROS emission during surgery-induced acute insulin resistance." American Journal of Physiology-Endocrinology and Metabolism 308, no. 8 (April 15, 2015): E613—E620. http://dx.doi.org/10.1152/ajpendo.00459.2014.
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Development of acute insulin resistance represents a negative factor after surgery, but the underlying mechanisms are not fully understood. We investigated the postoperative changes in insulin sensitivity, mitochondrial function, enzyme activities, and release of reactive oxygen species (ROS) in skeletal muscle and liver in pigs on the 2nd postoperative day after major abdominal surgery. Peripheral and hepatic insulin sensitivity were assessed by d-[6,6-2H2]glucose infusion and hyperinsulinemic euglycemic step clamping. Surgical trauma elicited a decline in peripheral insulin sensitivity (∼34%, P < 0.01), whereas hepatic insulin sensitivity remained unchanged. Intramyofibrillar (IFM) and subsarcolemma mitochondria (SSM) isolated from skeletal muscle showed a postoperative decline in ADP-stimulated respiration (VADP) for pyruvate (∼61%, P < 0.05, and ∼40%, P < 0.001, respectively), whereas VADP for glutamate and palmitoyl-l-carnitine (PC) was unchanged. Mitochondrial leak respiration with PC was increased in SSM (1.9-fold, P < 0.05) and IFM (2.5-fold, P < 0.05), indicating FFA-induced uncoupling. The activity of the pyruvate dehydrogenase complex (PDC) was reduced (∼32%, P < 0.01) and positively correlated to the decline in peripheral insulin sensitivity ( r = 0.748, P < 0.05). All other mitochondrial enzyme activities were unchanged. No changes in mitochondrial function in liver were observed. Mitochondrial H2O2 and O2·− emission was measured spectrofluorometrically, and H2O2 was increased in SSM, IFM, and liver mitochondria (∼2.3-, ∼2.5-, and ∼2.3-fold, respectively, all P < 0.05). We conclude that an impairment in skeletal muscle mitochondrial PDC activity and pyruvate oxidation capacity arises in the postoperative phase along with increased ROS emission, suggesting a link between mitochondrial function and development of acute postoperative insulin resistance.
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Vary, T. C. "Increased pyruvate dehydrogenase kinase activity in response to sepsis." American Journal of Physiology-Endocrinology and Metabolism 260, no. 5 (May 1, 1991): E669—E674. http://dx.doi.org/10.1152/ajpendo.1991.260.5.e669.
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The effect of sterile inflammation and sepsis on the proportion of active pyruvate dehydrogenase complex (PDH) in mitochondria isolated from skeletal muscle has been investigated. The proportion of active PDH in mitochondria isolated from septic animals was significantly reduced compared with control under all incubation conditions examined, even in the presence of inhibitors of the PDH kinase. There was no significant difference between control and sterile inflammation in any of the incubations examined. The rate constant for ATP-dependent inactivation of the PDH complex in mitochondrial extracts from control animals was -0.42 min-1 (r = 0.993; P less than 0.001) and was not altered in mitochondrial extracts from sterile inflammatory animals (-0.43 min-1; r = 0.999; P less than 0.001). However, rate constants for inactivation in septic animals was significantly increased over twofold to -1.08 min-1 (r = 0.987; P less than 0.001) (P less than 0.001 vs. control or sterile inflammation). In the presence of inhibitors of the PDH kinase reaction (2.5 mM pyruvate or 1 mM dichloroacetate), inactivation of PDH after addition of ATP was significantly greater in mitochondrial extracts from septic than either control or sterile inflammatory animals. These results suggest that sepsis, but not sterile inflammation, induces a stable factor in skeletal muscle mitochondria that increased PDH kinase activity.
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Zhao, Weicheng, Amy C. Kelly, Rosa I. Luna-Ramirez, Christopher A. Bidwell, Miranda J. Anderson, and Sean W. Limesand. "Decreased Pyruvate but Not Fatty Acid Driven Mitochondrial Respiration in Skeletal Muscle of Growth Restricted Fetal Sheep." International Journal of Molecular Sciences 24, no. 21 (October 30, 2023): 15760. http://dx.doi.org/10.3390/ijms242115760.
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Fetuses with intrauterine growth restriction (FGR) have impaired oxidative and energy metabolism, with persistent consequences on their postnatal development. In this study, we test the hypothesis that FGR skeletal muscle has lower mitochondrial respiration rate and alters the transcriptomic profiles associated with energy metabolism in an ovine model. At late gestation, mitochondrial oxygen consumption rates (OCRs) and transcriptome profiles were evaluated in the skeletal muscle collected from FGR and control fetuses. The ex vivo mitochondrial OCRs were reduced (p < 0.01) in permeabilized FGR soleus muscle compared to the control muscle but only with pyruvate as the metabolic substrate. Mitochondrial OCRs were similar between the FGR and control groups for palmitoyl-carnitine (fatty acid-driven) or pyruvate plus palmitoyl-carnitine metabolic substrates. A total of 2284 genes were differentially expressed in the semitendinosus muscle from growth restricted fetuses (false discovery rate (FDR) ≤ 0.05). A pathway analysis showed that the upregulated genes (FGR compared to control) were overrepresented for autophagy, HIF-1, AMPK, and FOXO signaling pathways (all with an FDR < 0.05). In addition, the expression of genes modulating pyruvate’s entry into the TCA cycle was downregulated, whereas the genes encoding key fatty acid oxidation enzymes were upregulated in the FGR muscle. These findings show that FGR skeletal muscle had attenuated mitochondrial pyruvate oxidation, possibly associated with the inability of pyruvate to enter into the TCA cycle, and that fatty acid oxidation might compensate for the attenuated energy metabolism. The current study provided phenotypic and molecular evidence for adaptive deficiencies in FGR skeletal muscle.
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Lerchundi, Rodrigo, Ignacio Fernández-Moncada, Yasna Contreras-Baeza, Tamara Sotelo-Hitschfeld, Philipp Mächler, Matthias T. Wyss, Jillian Stobart, et al. "NH4+ triggers the release of astrocytic lactate via mitochondrial pyruvate shunting." Proceedings of the National Academy of Sciences 112, no. 35 (August 18, 2015): 11090–95. http://dx.doi.org/10.1073/pnas.1508259112.
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Neural activity is accompanied by a transient mismatch between local glucose and oxygen metabolism, a phenomenon of physiological and pathophysiological importance termed aerobic glycolysis. Previous studies have proposed glutamate and K+ as the neuronal signals that trigger aerobic glycolysis in astrocytes. Here we used a panel of genetically encoded FRET sensors in vitro and in vivo to investigate the participation of NH4+, a by-product of catabolism that is also released by active neurons. Astrocytes in mixed cortical cultures responded to physiological levels of NH4+ with an acute rise in cytosolic lactate followed by lactate release into the extracellular space, as detected by a lactate-sniffer. An acute increase in astrocytic lactate was also observed in acute hippocampal slices exposed to NH4+ and in the somatosensory cortex of anesthetized mice in response to i.v. NH4+. Unexpectedly, NH4+ had no effect on astrocytic glucose consumption. Parallel measurements showed simultaneous cytosolic pyruvate accumulation and NADH depletion, suggesting the involvement of mitochondria. An inhibitor-stop technique confirmed a strong inhibition of mitochondrial pyruvate uptake that can be explained by mitochondrial matrix acidification. These results show that physiological NH4+ diverts the flux of pyruvate from mitochondria to lactate production and release. Considering that NH4+ is produced stoichiometrically with glutamate during excitatory neurotransmission, we propose that NH4+ behaves as an intercellular signal and that pyruvate shunting contributes to aerobic lactate production by astrocytes.
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Herbst, Eric A. F., Mitchell A. J. George, Karen Brebner, Graham P. Holloway, and Daniel A. Kane. "Lactate is oxidized outside of the mitochondrial matrix in rodent brain." Applied Physiology, Nutrition, and Metabolism 43, no. 5 (May 2018): 467–74. http://dx.doi.org/10.1139/apnm-2017-0450.
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The nature and existence of mitochondrial lactate oxidation is debated in the literature. Obscuring the issue are disparate findings in isolated mitochondria, as well as relatively low rates of lactate oxidation observed in permeabilized muscle fibres. However, respiration with lactate has yet to be directly assessed in brain tissue with the mitochondrial reticulum intact. To determine if lactate is oxidized in the matrix of brain mitochondria, oxygen consumption was measured in saponin-permeabilized mouse brain cortex samples, and rat prefrontal cortex and hippocampus (dorsal) subregions. While respiration in the presence of ADP and malate increased with the addition of lactate, respiration was maximized following the addition of exogenous NAD+, suggesting maximal lactate metabolism involves extra-matrix lactate dehydrogenase. This was further supported when NAD+-dependent lactate oxidation was significantly decreased with the addition of either low-concentration α-cyano-4-hydroxycinnamate or UK-5099, inhibitors of mitochondrial pyruvate transport. Mitochondrial respiration was comparable between glutamate, pyruvate, and NAD+-dependent lactate oxidation. Results from the current study demonstrate that permeabilized brain is a feasible model for assessing lactate oxidation, and support the interpretation that lactate oxidation occurs outside the mitochondrial matrix in rodent brain.
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O'Reilly, Ian, and Michael P. Murphy. "Studies on the rapid stimulation of mitochondrial respiration by thyroid hormones." Acta Endocrinologica 127, no. 6 (December 1992): 542–46. http://dx.doi.org/10.1530/acta.0.1270542.
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Injection of L-3,5-diiodothyronine (T2) into rats made hypothyroid by 6-n-propyl-2-thiouracil (PTU) increased the respiration rates of subsequently isolated liver mitochondria; this stimulation of respiration by T2 occurred in the presence of cycloheximide and is therefore independent of protein synthesis on cytoplasmic ribosomes. Injection of T3 into PTU-treated rats had a lesser effect than T2 on the respiration rates of subsequently isolated mitochondria; as PTU is an inhibitor of 5′-iodothyronine deiodinases, which convert T3 into T2 in vivo, the rapid stimulation of mitochondrial respiration by T3, which has been shown in a range of systems, may not be due directly to T3 itself, but may be mediated by its deiodination product T2. Injection of T2, or T3, into hypothyroid or euthyroid rats had no effect on the percentage activity of mitochondrial pyruvate hydrogenase assayed 30 min later. The amount of active pyruvate dehydrogenase is regulated by changes in mitochondrial calcium concentration and matrix ATP/ADP ratio; therefore these parameters are not persistently affected by treatment with T3 or T2. In addition, the total amount of pyruvate dehydrogenase present was the same in euthyroid and hypothyroid rats, indicating that the expression of this enzyme is not stringently controlled by thyroid hormone status.
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John, Scott, Guillaume Calmettes, Shili Xu, and Bernard Ribalet. "Real-time resolution studies of the regulation of pyruvate-dependent lactate metabolism by hexokinases in single cells." PLOS ONE 18, no. 11 (November 2, 2023): e0286660. http://dx.doi.org/10.1371/journal.pone.0286660.
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Lactate is a mitochondrial substrate for many tissues including neuron, muscle, skeletal and cardiac, as well as many cancer cells, however little is known about the processes that regulate its utilization in mitochondria. Based on the close association of Hexokinases (HK) with mitochondria, and the known cardio-protective role of HK in cardiac muscle, we have investigated the regulation of lactate and pyruvate metabolism by hexokinases (HKs), utilizing wild-type HEK293 cells and HEK293 cells in which the endogenous HKI and/or HKII have been knocked down to enable overexpression of wild type and mutant HKs. To assess the real-time changes in intracellular lactate levels the cells were transfected with a lactate specific FRET probe. In the HKI/HKII double knockdown cells, addition of extracellular pyruvate caused a large and sustained decrease in lactate. This decrease was rapidly reversed upon inhibition of the malate aspartate shuttle by aminooxyacetate, or inhibition of mitochondrial oxidative respiration by NaCN. These results suggest that in the absence of HKs, pyruvate-dependent activation of the TCA cycle together with the malate aspartate shuttle facilitates lactate transformation into pyruvate and its utilization by mitochondria. With replacement by overexpression of HKI or HKII the cellular response to pyruvate and NaCN was modified. With either hexokinase present, both the decrease in lactate due to the addition of pyruvate and the increase following addition of NaCN were either transient or suppressed altogether. Blockage of the pentose phosphate pathway with the inhibitor 6-aminonicotinamide (6-AN), abolished the effects of HK replacement. These results suggest that blocking of the malate aspartate shuttle by HK may involve activation of the pentose phosphate pathway and increased NADPH production.
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Štáfková, Jitka, Jan Mach, Marc Biran, Zdeněk Verner, Frédéric Bringaud, and Jan Tachezy. "Mitochondrial pyruvate carrier inTrypanosoma brucei." Molecular Microbiology 100, no. 3 (February 10, 2016): 442–56. http://dx.doi.org/10.1111/mmi.13325.
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Toleikis, Adolfas, Sonata Trumbeckaite, and Daiva Majiene. "Cytochrome c Effect on Respiration of Heart Mitochondria: Influence of Various Factors." Bioscience Reports 25, no. 5-6 (October 12, 2005): 387–97. http://dx.doi.org/10.1007/s10540-005-2897-2.
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The effect of exogenous cytochrome c on respiration rate of the rat and human heart mitochondria was assessed in situ, using permeabilized fibers. It was (i) much more pronounced in State 2 and 4 than in State 3 with all the respiratory substrates (pyruvate+malate, succinate, palmitoyl-CoA+carnitine and octanoyl-L-carnitine), (ii) different with different substrates, (iii) much higher after ischemia in both metabolic states, particularly in the case of succinate oxidation compared to pyruvate+malate, (iv) the highest in State 4 with succinate as a substrate. Similar results were obtained with the isolated rat and rabbit heart mitochondria. The differences in the degree of stimulation of mitochondrial respiration by cytochrome c and, thus, sensitivity of cytochrome c test in evaluation of the intactness/injury of outer mitochondrial membrane are probably determined by the differences in the cytochrome c role in the control of mitochondrial respiration in the above-described conditions.