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 (pH_i) was evaluated. STUDY DESIGN AND 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. pH_i 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 pH _i 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 V _max values (ΔpH/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. CONCLUSION: The results show that the pH _i regulation system is largely preserved during freeze-drying and rehydration of PLTs.