The mitochondrial membrane potential in human platelets: a sensitive parameter for platelet quality.
BACKGROUND
Deterioration of platelet (PLT) quality during storage is accompanied by an increase in lactate production, indicating a decrease in mitochondrial function. In this study, the optimal conditions under which the fluorescent dye JC-1 can be used to detect changes in mitochondrial function in PLTs were established.


METHODS
PLTs were incubated at 37 degrees C in synthetic medium under various conditions of JC-1 loading. In the presence of a high membrane potential, this dye accumulates in the mitochondria with a concomitant increase in red fluorescence. After JC-1 loading, the ratio of red (FL2) to green (FL1) fluorescence was determined by flow cytometry.


RESULTS
The FL2-to-FL1 ratio of PLTs (3 x 10(7)/mL, loaded with 0.5 micromol/L JC-1) amounted to about 5 in 1-day-old PLTs. At higher dye concentrations, the FL2-to-FL1 ratio was significantly lower, suggesting uncoupling by the dye itself. Plasma concentrations above 3 percent significantly affected the JC-1 signal. The FL2-to-FL1 ratio showed a dose-dependent decrease to an uncoupler of oxidative phosphorylation or to inhibition of the respiratory chain. JC-1-loaded PLTs showed a clear decrease in FL2-to-FL1 ratio after prolonged storage or upon ultraviolet (UV) illumination. Only after UV treatment did changes in JC-1 signal correlate with changes in CD62P expression.


CONCLUSIONS
The FL2-to-F1 ratio of PLTs loaded with JC-1 is a reliable and sensitive indicator of the mitochondrial membrane potential, provided that the proper experimental conditions have been applied.
Verhoeven
Arthur J
AJ
Sanquin Research at CLB, Amsterdam, the Netherlands. a.verhoeven@sanquin.nl
Verhaar
Robin
R
Gouwerok
Eric G W
EG
de Korte
Dirk
D
D001792:Blood Platelets / Q000502:physiology*; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D006801:Humans; D008564:Membrane Potentials; D008928:Mitochondria / Q000502:physiology*; D019007:P-Selectin / Q000097:blood; D014466:Ultraviolet Rays
D005456:Fluorescent Dyes; D019007:P-Selectin