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Vesiculation of platelets during in vitro aging
A P Bode1, S M Orton, M J Frye
1Department of Clinical Pathology and Diagnostic Medicine, East Carolina University School of Medicine, Greenville, NC 27858.
Abstract:
Membranous microparticles (MP) appearing in the supernatant plasma of stored platelet concentrates (PC) were analyzed by flow cytometry. Two populations of MP were arbitrarily delineated by light scatter as larger or smaller than 0.5 micron fluorescent beads. An estimate of MP concentration was obtained by adding a known amount of fluorescent beads to each sample before analysis of a set number of counts on the flow cytometer. The addition of platelet activation inhibitors (prostaglandin E-1, theophylline, and aprotinin) to the anticoagulant during preparation of PC combined with a reduction in surface area of the storage container caused approximately a 40% reduction in the number of MP appearing during storage relative to donor-matched controls. In addition, the inhibited concentrates had 84% less platelet factor 3 (PF3) activity in the supernatant and 61% less released lactic dehydrogenase. A reduction in surface area of the container in the controls partially offset these differences. A significant correlation was found (rs = .748) between PF3 levels and the concentration of larger MP. The inhibitors did not reduce the small number of MP found in stored platelet-poor plasma. Surface antigen analysis showed that the majority of MP in PC were platelet-derived; most were positive for glycoprotein (GP) IIbIIIa (73%) and/or for GPIb (43% to 46%). We conclude that procoagulant MP are released from platelets during storage as a result of platelet activation augmented by interaction of platelets with the bag wall.
Insights
Storage of platelet concentrates releases membranous microparticles (MP). Inhibitors and reduced container surface area significantly decrease MP formation and associated procoagulant activity, improving platelet concentrate quality.
Area of Science:
- Hematology
- Biotechnology
- Materials Science
Background:
- Membranous microparticles (MP) are released into plasma from stored platelet concentrates (PC).
- MP formation is influenced by storage conditions and platelet activation.
Purpose of the Study:
- To analyze MP populations in stored PC.
- To investigate methods for reducing MP generation during PC storage.
- To characterize the nature and origin of MP in PC.
Main Methods:
- Flow cytometry was used to quantify MP populations based on size.
- Platelet activation inhibitors (prostaglandin E-1, theophylline, aprotinin) and reduced container surface area were tested.
- Platelet factor 3 (PF3) activity and lactic dehydrogenase release were measured.
- Surface antigen analysis identified MP origins.
Main Results:
- Inhibitors and reduced container surface area reduced MP by ~40%.
- Inhibited concentrates showed significantly lower PF3 activity (84%) and lactic dehydrogenase release (61%).
- A strong correlation (rs = .748) was found between PF3 levels and larger MP concentration.
- Most MP were platelet-derived, expressing glycoprotein (GP) IIbIIIa and/or GPIb.
Conclusions:
- Procoagulant MP are released from activated platelets during PC storage.
- Platelet activation is exacerbated by interaction with the storage container.
- Storage conditions can be modified to reduce MP formation and improve PC quality.