Freeze-dried rehydrated human blood platelets regulate intracellular pH

Minke Tang, Willem F. Wolkers, John H. Crowe, Fern Tablin

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

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. 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. 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 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.

Original languageEnglish (US)
Pages (from-to)1029-1037
Number of pages9
JournalTransfusion
Volume46
Issue number6
DOIs
StatePublished - Jun 2006

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Blood Platelets
Trehalose
Freeze Drying
Nigericin
Fluid Therapy
Proton Pumps
Sodium-Potassium-Exchanging ATPase
Amiloride
Thrombin
Coloring Agents
Therapeutics

ASJC Scopus subject areas

  • Hematology
  • Immunology

Cite this

Freeze-dried rehydrated human blood platelets regulate intracellular pH. / Tang, Minke; Wolkers, Willem F.; Crowe, John H.; Tablin, Fern.

In: Transfusion, Vol. 46, No. 6, 06.2006, p. 1029-1037.

Research output: Contribution to journalArticle

Tang, Minke ; Wolkers, Willem F. ; Crowe, John H. ; Tablin, Fern. / Freeze-dried rehydrated human blood platelets regulate intracellular pH. In: Transfusion. 2006 ; Vol. 46, No. 6. pp. 1029-1037.
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abstract = "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. 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. 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 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.",
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N2 - 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. 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. 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 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.

AB - 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. 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. 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 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.

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