Rapid kinetics of changes in oxygen consumption rate in thrombin-stimulated platelets measured by high-resolution respirometry

Sowton, Alice P.; Millington‐Burgess, Sarah L.; Murray, Andrew J.; Harper, Matthew T.

doi:10.1016/j.bbrc.2018.08.031

Cited by 14 publications

(12 citation statements)

References 33 publications

(38 reference statements)

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“…In response to the stimulation of blood platelets with ADP, coupled physiological respiration increased to meet the requirements of elevated energy demands. A similar observation was made by Sowton et al [35] after platelet stimulation with thrombin. Moreover, we noted a significant association between platelet activation and the mitochondrial parameters related to the elevated coupled respiration in response to ADP stimulation, especially when two higher concentrations were used.…”

Section: The Effect Of Platelet Activation With Adp On Mitochondrial Respirationsupporting

confidence: 83%

Sample Preparation as a Critical Aspect of Blood Platelet Mitochondrial Respiration Measurements—The Impact of Platelet Activation on Mitochondrial Respiration

Siewiera

Łabieniec-Watała

Wolska

et al. 2021

IJMS

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Blood platelets are considered as promising candidates as easily-accessible biomarkers of mitochondrial functioning. However, their high sensitivity to various stimulus types may potentially affect mitochondrial respiration and lead to artefactual outcomes. Therefore, it is crucial to identify the factors associated with platelet preparation that may lead to changes in mitochondrial respiration. A combination of flow cytometry and advanced respirometry was used to examine the effect of blood anticoagulants, the media used to suspend isolated platelets, respiration buffers, storage time and ADP stimulation on platelet activation and platelet mitochondria respiration. Our results clearly show that all the mentioned factors can affect platelet mitochondrial respiration. Briefly, (i) the use of EDTA as anticoagulant led to a significant increase in the dissipative component of respiration (LEAK), (ii) the use of plasma for the suspension of isolated platelets with MiR05 as a respiration buffer allows high electron transfer capacity and low platelet activation, and (iii) ADP stimulation increases physiological coupling respiration (ROUTINE). Significant associations were observed between platelet activation markers and mitochondrial respiration at different preparation steps; however, the fact that these relationships were not always apparent suggests that the method of platelet preparation may have a greater impact on mitochondrial respiration than the platelet activation itself.

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Section: The Effect Of Platelet Activation With Adp On Mitochondrial Respirationsupporting

confidence: 83%

Sample Preparation as a Critical Aspect of Blood Platelet Mitochondrial Respiration Measurements—The Impact of Platelet Activation on Mitochondrial Respiration

Siewiera

Łabieniec-Watała

Wolska

et al. 2021

IJMS

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“…These systems are the high-resolution respirometry Oxygraph-2k (O2k, Oroboros Instruments, Austria) and the sensitive high-throughput Seahorse XF Extracellular Flux Analyzer (Seahorse XF, Seahorse Bioscience Inc.). Nowadays, both these methods are widely used for the evaluation of peripheral blood cells bioenergetics, including platelets [ 135 , 199 , 200 ]. The O2k measures, in real time, the rapid changes in oxygen concentration (nmol/mL) and oxygen consumption (pmol/sec/mL), both markers of OXPHOS, using a polarographic sensors.…”

Section: Methods To Assess Platelets Redox Biologymentioning

confidence: 99%

“…Mitochondrial O•2- can be then converted in H 2 O 2 by SOD2 [ 29 ]. Different research groups have demonstrated that platelet activation by collagen and thrombin induces a rapid and transient increase in the mitochondrial membrane potential (ΔΨm) and OXPHOS, likely via Ca 2+ mobilization [ 135 , 136 ]. The increase in ΔΨm is associated to the increased production of ROS in mitochondria, and hyperpolarization of the membrane reduces the electron transport chain, resulting in a leakage of electrons from the chain followed by promotion of the production of O•2- [ 137 ].…”

Section: Role Of Platelet Mitochondria In Redox Balancementioning

confidence: 99%

ROS in Platelet Biology: Functional Aspects and Methodological Insights

Masselli

Pozzi

Vaccarezza

et al. 2020

IJMS

114

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Reactive oxygen species (ROS) and mitochondria play a pivotal role in regulating platelet functions. Platelet activation determines a drastic change in redox balance and in platelet metabolism. Indeed, several signaling pathways have been demonstrated to induce ROS production by NAPDH oxidase (NOX) and mitochondria, upon platelet activation. Platelet-derived ROS, in turn, boost further ROS production and consequent platelet activation, adhesion and recruitment in an auto-amplifying loop. This vicious circle results in a platelet procoagulant phenotype and apoptosis, both accounting for the high thrombotic risk in oxidative stress-related diseases. This review sought to elucidate molecular mechanisms underlying ROS production upon platelet activation and the effects of an altered redox balance on platelet function, focusing on the main advances that have been made in platelet redox biology. Furthermore, given the increasing interest in this field, we also describe the up-to-date methods for detecting platelets, ROS and the platelet bioenergetic profile, which have been proposed as potential disease biomarkers.

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“…[1][2][3][4][5][6][7] These early investigations used radiometric methods with labeled glucose to study glycolysis and oxidative phosphorylation in platelets. 8,9 Recently, these techniques have been supplanted by more sophisticated investigative tools that allow more rapid analysis of small volume samples, including high-resolution respirometry 10,11 and Seahorse extracellular flux analysis. [12][13][14] Glycolysis and oxidative phosphorylation, as well as glutaminolysis and fatty acid b oxidation, all function to support the metabolic demand in platelets.…”

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confidence: 99%

Platelet respiration

Kholmukhamedov

Jobe

2019

Blood Advances

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Historically, metabolic studies in platelets have primarily investigated events occurring during ex vivo platelet storage, less so the consequences of metabolic alterations on platelet homeostasis and physiologic function. The importance of such studies is emphasized by the recent finding of platelet mitochondrial dysfunction in type 2 diabetes, sickle cell disease, and sepsis. [1][2][3][4][5][6][7] These early investigations used radiometric methods with labeled glucose to study glycolysis and oxidative phosphorylation in platelets. 8,9 Recently, these techniques have been supplanted by more sophisticated investigative tools that allow more rapid analysis of small volume samples, including high-resolution respirometry 10,11 and Seahorse extracellular flux analysis. [12][13][14] Glycolysis and oxidative phosphorylation, as well as glutaminolysis and fatty acid b oxidation, all function to support the metabolic demand in platelets. 10,[14][15][16] The metabolic flexibility of the platelet has been emphasized by investigations demonstrating that activated platelets exhibit a glycolytic phenotype while preserving mitochondrial function and can easily switch between glucose and fatty acid catabolism to support activation. 12 The recently published study by A. K. Chauhan's group further advances our understanding of the understudied field of platelet metabolism. 13 In this paper, the authors demonstrate that dichloroacetate, an inhibitor of pyruvate dehydrogenase kinases, alters platelet metabolism and function. 13 Finding salutary antiaggregatory and antithrombotic effects of dichloroacetate, the authors propose targeting of the platelet metabolic response as a novel antithrombotic approach. However, there are few points of clarification we would like to add, which we believe will be of great benefit for the platelet and mitochondria research community in their exploration of this new therapeutic avenue. 13

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Rapid kinetics of changes in oxygen consumption rate in thrombin-stimulated platelets measured by high-resolution respirometry

Cited by 14 publications

References 33 publications

Sample Preparation as a Critical Aspect of Blood Platelet Mitochondrial Respiration Measurements—The Impact of Platelet Activation on Mitochondrial Respiration

Sample Preparation as a Critical Aspect of Blood Platelet Mitochondrial Respiration Measurements—The Impact of Platelet Activation on Mitochondrial Respiration

ROS in Platelet Biology: Functional Aspects and Methodological Insights

Platelet respiration

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