Decline in platelet microparticles contributes to reduced hemostatic potential of stored plasma

doi:10.1016/j.thromres.2011.02.011

Thrombosis Research

Volume 128, Issue 1, July 2011, Pages 35-41

https://doi.org/10.1016/j.thromres.2011.02.011 Get rights and content

Abstract

Introduction

In an effort to administer life-saving transfusions quickly, some trauma centers maintain thawed plasma (TP). According to AABB, TP is approved for transfusion for up to five days when stored at 1 - 6 °C. However, the alterations in microparticles (MP) contained in the plasma, which are an integral component of plasma's hemostatic capacity, are not well characterized. We report on MP changes in TP between its initial thaw (FFP-0) and five days (FFP-5) of storage.

Materials and Methods

FFP units (n = 30) were thawed at 37 °C and kept refrigerated for five days. Phenotypes of residual cells, which include platelets, erythrocytes, leukocytes, monocytes, endothelial cells, and MP counterparts of each cell type, were analyzed by flow cytometry. Functional assays were used for MP procoagulant activity, plasma thrombin generation, and clotting properties (thromboelastography).

Results

In FFP-0 the majority (94%) of residual cells were platelets, along with significant levels of platelet MPs (4408 × 10³/L). FFP-5 showed a decline in MP count by 50% (p < 0.0001), and procoagulant activity by 29% (p < 0.0001). FFP-5 exhibited only 54% (p < 0.0001) of the potential for thrombin generation as FFP-0, while thromboelastography indicated a slower clotting response (p < 0.0001) and a longer delay in reaching maximum clot (p < 0.01). Removal of MP by filtration resulted in reduced thrombin generation, while the MP replacement restored it.

Conclusions

Decline in MP with storage contributes to FFP-5's reduced ability to provide the hemostatic potential exhibited by FFP-0, suggesting the presence of platelet MPs in freshly TP may be beneficial and protective in the initial treatment of hemorrhage.

Section snippets

Plasma

FFP units from 30 blood donors were obtained from the Gulf Coast Regional Blood Center (Houston, TX), as previously reported [11]. Blood was collected in citrate-phosphate-dextrose anticoagulant and plasma was prepared by centrifugation and separation from cellular components [6]. The mean donor age was 42.5 years (range 17 – 66 years), included different blood groups (13 O, 11 A, 4 B and 2 AB) and 47% were male blood donors. FFP was thawed in a water bath at 37 °C according to standard AABB

Residual cells

Flow cytometry results were recorded and analyzed for residual cells. The median concentrations (IQR) and percentages of residual cells are presented in Table 1. The majority of residual cells (94%) were platelets, and less of other cell types. Residual platelets were activated as evidenced by their light scatter properties and increased expression of the glycoprotein IIb (presented as median fluorescent intensity): 198 (IQR 184 – 302) vs. 116 (IQR 103 - 186), p < 0.01), compared to non-activated

Discussion

Our study confirms that TP units from a commercial blood bank vendor contain residual cells, predominately platelets, and high levels of platelet MPs. Although quality control requirements for blood components differ between US and Europe, and AABB does not require testing for residual cells, our results indicate that FFP meets the Council of Europe standards [20]. Residual platelets were highly activated during the freeze-thaw cycle and storage, which was evidenced by light scatter

Conflict of interest statement

None.

Acknowledgments

Authors thank Ms. Robin Fuller from the Gulf Coast Regional Blood Center for providing assistance in procurement of plasma products, and Dr. R. Michelle Sauer for her editorial support. Supported by the Department of Defense via the PRospective Observational Multicenter Massive Transfusion sTudy (PROMMTT) (W81XWH-08-C-0712).

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Submicron-sized extra-cellular vesicles generated by budding from the external cell membranes, microparticles (MPs) are important actors in transfusion as well as in other medical specialties. After briefly positioning their role in the characterization of labile blood products, this technically oriented chapter aims to review practical points that need to be considered when trying to use flow cytometry for the analysis, characterization and absolute counting of MP subsets. Subjects of active discussions relative to instrumentation will include the choice of the trigger parameter, possible standardization approaches requiring instrument quality-control, origin and control of non-specific background and of coincidence artifacts, choice of the type of electronic signals, optimal sheath fluid and sample speed. Questions related to reagents will cover target antigens and receptors, multi-color reagents, negative controls, enumeration of MPs and limiting artifacts due to unexpected (micro-) coagulation of plasma samples. Newly detected problems are generating innovative solutions and flow cytometry will continue to remain the technology of choice for the analysis of MPs, in the domain of transfusion as well as in many diverse specialties.
An overview of the role of microparticles/microvesicles in blood components: Are they clinically beneficial or harmful?
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One study identified that the MP content was reduced by leucofiltration of whole blood on a negatively charged filter, and such reduction had an impact by lowering the procoagulant and haemostatic potential of FFP [97]. The plasma's haemostatic activity involves several parameters such as its content of coagulant factors and coagulation inhibitors, as well as procoagulant MPs [93]. Although MPs contribute to clot formation in plasma and to a haemostatic effect [4,93,96], their excess in plasma for transfusion may possibly trigger thrombotic events in susceptible patients.
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Plasma continues to be commonly transfused, both prophylactically to prevent bleeding in critically ill patients perceived to be at higher risk of bleeding and therapeutically to treat bleeding in patients with uncontrolled bleeding. For both indications, the main rationale is to replace or supplement deficiencies or depleted levels of prohemostatic coagulation factors. What is often not appreciated by clinicians is the variability between plasma collections for transfusion, nor the impact of collection and processing and storage on levels of coagulant factors.

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Regular ArticleDecline in platelet microparticles contributes to reduced hemostatic potential of stored plasma

Abstract

Introduction

Materials and Methods

Results

Conclusions

Section snippets

Plasma

Residual cells

Discussion

Conflict of interest statement

Acknowledgments

Blood Rev

Transfus Sci

J Thromb Haemost

J Thromb Haemost

J Thromb Haemost

Thromb Res

Cell

Blood Cells Mol Dis

Blood

Bench-to-bedside review: latest results in hemorrhagic shock

Crit Care

The coagulopathy of trauma: a review of mechanisms

J Trauma

Increased plasma and platelet to red blood cell ratios improves outcome in 466 massively transfused civilian trauma patients

Ann Surg

AABB Guidelines and standards for blood banks and transfusion services

AABB Technical Manual

Stability of coagulation factors in thawed, solvent/detergent-treated plasma during storage at 4 degrees C for 6 days

Vox Sang

Coagulation parameters of thawed fresh-frozen plasma during storage at different temperatures

Transfus Med

Protective effects of fresh frozen plasma on vascular endothelial permeability, coagulation, and resuscitation after hemorrhagic shock are time dependent and diminish between days 0 and 5 after thaw

J Trauma

Study of coagulation factor activities in apheresed thawed fresh frozen plasma at 1–6 degrees C for five days

J Clin Apher

Regular Article
Decline in platelet microparticles contributes to reduced hemostatic potential of stored plasma