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

TY - JOUR

T1 - Sample Preparation as a Critical Aspect of Blood Platelet Mitochondrial Respiration Measurements-The Impact of Platelet Activation on Mitochondrial Respiration

AU - Siewiera, Karolina

AU - Labieniec-Watala, Magdalena

AU - Wolska, Nina

AU - Kassassir, Hassan

AU - Watala, Cezary

PY - 2021/8/28

Y1 - 2021/8/28

N2 - 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.

AB - 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.

KW - Anticoagulants/pharmacology

KW - Blood Platelets/drug effects

KW - Cell Respiration/physiology

KW - Culture Media/pharmacology

KW - Flow Cytometry

KW - Humans

KW - Mitochondria/drug effects

KW - Platelet Activation

U2 - 10.3390/ijms22179332

DO - 10.3390/ijms22179332

M3 - SCORING: Journal article

C2 - 34502240

VL - 22

JO - INT J MOL SCI

JF - INT J MOL SCI

SN - 1661-6596

IS - 17

M1 - 9332

ER -