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A dataset describing glycolytic inhibitors overcoming the underestimation of maximal mitochondrial oxygen consumption rate in oligomycin-treated cells.

ABSTRACT: Determination of oxygen consumption is one of the most valuable methodologies to evaluate mitochondrial (dys)function. Previous studies demonstrated that a widely used protocol, consisting of adding the ATP synthase inhibitor oligomycin before mitochondrial respiratory uncoupling by sequential addition of a protonophore (e.g., carbonyl cyanide 3-chlorophenyl hydrazone [CCCP]), may lead to underestimation of maximal oxygen consumption rate (OCR_max) and spare respiratory capacity (SRC) parameters in highly glycolytic tumor cell lines. In this dataset, we report the effects of the glycolytic inhibitors 2-deoxy-D-glucose, iodoacetic acid, and lonidamine on overcoming the underestimation of OCR_max and SRC in oligomycin-treated cells. We propose a protocol in which 2-deoxy-D-glucose is added after oligomycin and just before the sequential addition of CCCP to avoid underestimation of OCR_max and SRC parameters in A549, C2C12, and T98G cells. The oxygen consumption rates were determined in intact suspended cell lines using a high-resolution oxygraph device. The data can be used in several fields of research that require characterization of mitochondrial respiratory parameters in intact cells.

SUBMITTER: Ruas JS

PROVIDER: S-EPMC8718714 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

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A dataset describing glycolytic inhibitors overcoming the underestimation of maximal mitochondrial oxygen consumption rate in oligomycin-treated cells.

Ruas Juliana S JS Siqueira-Santos Edilene S ES Navarro Claudia D C CDC Castilho Roger F RF

Data in brief 20211222

Determination of oxygen consumption is one of the most valuable methodologies to evaluate mitochondrial (dys)function. Previous studies demonstrated that a widely used protocol, consisting of adding the ATP synthase inhibitor oligomycin before mitochondrial respiratory uncoupling by sequential addition of a protonophore (e.g., carbonyl cyanide 3-chlorophenyl hydrazone [CCCP]), may lead to underestimation of maximal oxygen consumption rate (OCR<sub>max</sub>) and spare respiratory capacity (SRC) ...[more]

PMID: 35005137

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Project description:The maximal capacity of the mitochondrial electron transport system (ETS) in intact cells is frequently estimated by promoting protonophore-induced maximal oxygen consumption preceded by inhibition of oxidative phosphorylation by oligomycin. In the present study, human glioma (T98G and U-87MG) and prostate cancer (PC-3) cells were titrated with different concentrations of the protonophore CCCP to induce maximal oxygen consumption rate (OCR) within respirometers in a conventional growth medium. The results demonstrate that the presence of oligomycin or its A-isomer leads to underestimation of maximal ETS capacity. In the presence of oligomycin, the spare respiratory capacity (SRC), i.e., the difference between the maximal and basal cellular OCR, was underestimated by 25 to 45%. The inhibitory effect of oligomycin on SRC was more pronounced in T98G cells and was observed in both suspended and attached cells. Underestimation of SRC also occurred when oxidative phosphorylation was fully inhibited by the ATP synthase inhibitor citreoviridin. Further experiments indicated that oligomycin cannot be replaced by the adenine nucleotide translocase inhibitors bongkrekic acid or carboxyatractyloside because, although these compounds have effects in permeabilized cells, they do not inhibit oxidative phosphorylation in intact cells. We replaced CCCP by FCCP, another potent protonophore and similar results were observed. Lower maximal OCR and SRC values were obtained with the weaker protonophore 2,4-dinitrophenol, and these parameters were not affected by the presence of oligomycin. In permeabilized cells or isolated brain mitochondria incubated with respiratory substrates, only a minor inhibitory effect of oligomycin on CCCP-induced maximal OCR was observed. We conclude that unless a previously validated protocol is employed, maximal ETS capacity in intact cells should be estimated without oligomycin. The inhibitory effect of an ATP synthase blocker on potent protonophore-induced maximal OCR may be associated with impaired metabolism of mitochondrial respiratory substrates.

Project description:BackgroundEnhancement in maximal oxygen consumption (VO2max) induced by hypoxic training is important for both athletes and non-athletes. However, the lack of comparison of multiple paradigms and the exploration of related modulating factors leads to the inability to recommend the optimal regimen in different situations. This study aimed to investigate the efficacy of seven common hypoxic training paradigms on VO2max and associated moderators.MethodsElectronic (i.e., five databases) and manual searches were performed, and 42 studies involving 1246 healthy adults were included. Pairwise meta-analyses were conducted to compare different hypoxic training paradigms and hypoxic training and control conditions. The Bayesian network meta-analysis model was applied to calculate the standardised mean differences (SMDs) of pre-post VO2max alteration among hypoxic training paradigms in overall, athlete, and non-athlete populations, while meta-regression analyses were employed to explore the relationships between covariates and SMDs.ResultsAll seven hypoxic training paradigms were effective to varying degrees, with SMDs ranging from 1.45 to 7.10. Intermittent hypoxia interval training (IHIT) had the highest probability of being the most efficient hypoxic training paradigm in the overall population and athlete subgroup (42%, 44%), whereas intermittent hypoxic training (IHT) was the most promising hypoxic training paradigm among non-athletes (66%). Meta-regression analysis revealed that saturation hours (coefficient, 0.004; P = 0.038; 95% CI [0.0002, 0.0085]) accounted for variations of VO2max improvement induced by IHT.ConclusionEfficient hypoxic training paradigms for VO2max gains differed between athletes and non-athletes, with IHIT ranking best for athletes and IHT for non-athletes. The practicability of saturation hours is confirmed with respect to dose-response issues in the future hypoxic training and associated scientific research.RegistrationThis study was registered in the PROSPERO international prospective register of systematic reviews (CRD42022333548).

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A dataset describing glycolytic inhibitors overcoming the underestimation of maximal mitochondrial oxygen consumption rate in oligomycin-treated cells.

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A dataset describing glycolytic inhibitors overcoming the underestimation of maximal mitochondrial oxygen consumption rate in oligomycin-treated cells.

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