Freeze-dried rehydrated human blood platelets regulate intracellular pH.
BACKGROUND
Long-term storage of platelets (PLTs) in the dry state would greatly improve options for PLT storage. Whether trehalose-loaded freeze-dried and rehydrated PLTs could regulate intracellular pH (pHi) was evaluated.


METHODS
Previously it was shown that human PLTs can be successfully preserved by freeze-drying with trehalose. Trehalose-loaded freeze-dried rehydrated PLTs and fresh control PLTs were labeled with the pH dye BCECF-AM. pHi was measured in resting cells, cells acidified with nigericin, and cells treated with thrombin. The sodium-proton pump was blocked by treatment with 5-(N-methyl-N-isobutyl)amiloride (MIA).


RESULTS
The pHi of rehydrated PLTs is the same as that of fresh control PLTs, 7.27+/-0.03 (SD; n=5) and 7.27+/-0.02 (n=5), respectively. Nigericin treatment of cells showed that the recovery in pHi was Na+-dependent and followed Michaelis-Menten kinetics. The Vmax values (DeltapH/9 sec) were 0.21+/-0.039 (n=3) and 0.22+/-0.025 (n=3) for rehydrated and control PLTs, respectively. The exchange constants were 17.7+/-2.3 mmol per L (n=3) and 17.0+/-1.9 mmol per L (n=3) for rehydrated and control PLTs, respectively. Treatment of cells with MIA showed that NHE1 remained sensitive to the inhibitor after freeze-drying and rehydration.


CONCLUSIONS
The results show that the pHi regulation system is largely preserved during freeze-drying and rehydration of PLTs.
Tang
Minke
M
Center for Biostabilization, the Section of Molecular and Cellular Biology, and the Department of Anatomy, Physiology, and Cell Biology, the University of California, Davis, California 95616, USA.
Wolkers
Willem F
WF
Crowe
John H
JH
Tablin
Fern
F
D001792:Blood Platelets / Q000166:cytology*; D001793:Blood Preservation / Q000379:methods*; D027682:Cation Transport Proteins / Q000378:metabolism; D005440:Fluid Therapy; D005452:Fluoresceins; D005612:Freeze Drying; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism; D009550:Nigericin / Q000494:pharmacology; D017713:Platelet Transfusion / Q000592:standards; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism; D014199:Trehalose
D027682:Cation Transport Proteins; D005452:Fluoresceins; D008565:Membrane Proteins; C490998:SLC9A1 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; C057433:2',7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester; D014199:Trehalose; D009550:Nigericin