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Loss of thrombin-induced Ca2+ mobilization in a subpopulation of platelets during storage
R Fijnheer1, C H Homburg, B Hooibrink
1Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, Amsterdam.
Insights
Stored human platelets show reduced calcium signaling upon thrombin stimulation. This decline is due to a growing subpopulation of unresponsive platelets and diminished responsiveness in others, impacting platelet function.
Area of Science:
- Hematology
- Cellular Physiology
Background:
- Platelet function is critical for hemostasis.
- Storage of platelets can lead to functional impairments.
- Understanding these changes is vital for transfusion medicine.
Purpose of the Study:
- To investigate the impact of storage duration on thrombin-induced calcium signaling in human platelets.
- To elucidate the mechanisms behind the functional decline of stored platelets.
Main Methods:
- Human platelets stored for up to 6 days were used.
- Cytosolic free calcium ([Ca2+]i) changes were measured using Indo-1 fluorescence.
- Flow cytometry was employed to quantify responses in individual platelets.
Main Results:
- Maximal [Ca2+]i concentration upon thrombin stimulation decreased significantly with storage duration.
- The percentage of responsive platelets declined from 90% on day 0 to 47% on day 6.
- Both a non-responsive subpopulation and reduced responsiveness in remaining platelets contributed to the overall decline.
Conclusions:
- Platelet storage leads to impaired thrombin-induced calcium signaling.
- The functional defect arises from both platelet unresponsiveness and diminished signaling capacity.
- This provides insight into the functional deterioration of stored platelets.
Abstract:
Thrombin-induced changes in cytosolic free Ca2+ ([Ca2+]i) were studied in human platelets that had been stored for up to 6 days. Changes in [Ca2+]i were measured with Indo-1-loaded platelets and quantitated with two different methods: (i) measurement of the changes in total fluorescence; (ii) measurement of the [Ca2+]i changes in individual platelets in a flow cytometer, allowing the detection of non-responding platelets. The maximal concentration of [Ca2+]i after stimulation with 0.5 U of thrombin/ml decreased from 544 +/- 58 nM (mean +/- SEM, n = 6) on day 0, to 276 +/- 9 nM on day 3 and to 203 +/- 23 nM on day 6. The percentage of platelets responding to 0.5 U of thrombin/ml declined from 90 +/- 2% on day 0 to 72 +/- 4% on day 3, and to 47 +/- 8% on day 6. Nevertheless, also the responding platelets showed a decreased rise in [Ca2+]i. The study shows that during platelet storage a decrease in the rise in [Ca2+]i upon thrombin stimulation occurs. This decrease is partly due to the formation of a subpopulation of platelets that is completely unresponsive and partly due to a decreased responsiveness in the remainder of the platelets; it is not due to a gradual decline in [Ca2+]i rise in all platelets. This phenomenon provides new insight in the functional defect of stored platelets.