(P-CB-7) Contribution of Platelets and Their Subpopulations to the Mechanics of Clot Formation According to Platelet Concentrate Storage Conditions

Background/Case Studies: Platelets form a homogenous population in buffy-coat platelet concentrates (BC-PCs) stored at 22°C for 7 days, unlike at 4°C. Using multicolor flow cytometry analysis of platelet activation markers, we recently highlighted the emergence of procoagulant and apoptotic platelet subpopulations that progressively supplanted native platelets during cold storage of PCs up to 21 days. We evaluated the effects of storage conditions on the contribution of platelets and their subpopulations to clot formation using the Quantra viscoelastic analyzer (HemoSonics LLC, Durham, NC, USA), providing quantification of the Platelet Contribution to Clot Stiffness (PCS), enabling a comprehensive understanding of how platelets contribute to the mechanics of clot formation.

Study

Design/Methods: A pool-and-split strategy was used to obtain double-dose BC-PCs collected into PAS-III/plasma (55/45%) treated with amotosalen-UVA (INTERCEPT™ Blood System) and stored at 22°C with constant agitation or at 4°C without agitation up to 21 days. Platelet samples were reconstituted with washed red blood cells and thawed fresh frozen plasma (Ht 40%; platelet count 300 x 10⁹/L) as a model of whole blood to measure clot viscoelasticity (Clot Time CT, Clot Stiffness CS, Fibrinogen Contribution to clot Stiffness FCS and PCS) with the Quantra. Statistical comparisons were done by Pearson's correlation and two-way ANOVA followed by Tukey’s post-hoc test.

Results/Findings: When stored at 22°C, CT and CS of BC-PCs remained stable from D1 to D7. In contrast, CT of cold-stored BC-PCs was faster at D7 (189±4 s) compared to D1 (277±10 s, p=0.005), while CS was reduced (8±1 vs. 19±4 hPa, p=0.0004). FCS remained stable, highlighting a reduced contribution of platelets to clot stiffness during cold storage, evidenced by reduced PCS at D7 compared to D1 (6±1 vs. 17±4 hPa, p=0.0003). Interestingly, the procoagulant platelet subpopulation inversely correlated with CT and only in cold-stored PCs (r=-0.778, p=0.0041). At 22°C, the platelet count (r=0.8848, p< 0.0001), the proportion of native platelets (r=0.8585, p=0.0004) and the ability of platelets to aggregate (r=0.6901, p=0.0003) highly correlated with PCS, while the proportion of apoptotic platelets showed strong inverse correlation (r=-0.8788, p=0.0002) with PCS. At 4°C, only the proportion of native platelets remained highly correlated with PCS (r=0.8922, p< 0.0001), while procoagulant platelets in addition to apoptotic were strongly inversely correlated with PCS (r=-0.7227 p=0.0079), in line with the loss of their aggregation properties.

Conclusions: At 4°C, platelets display faster clot formation but lower clot stiffness than at 22°C, as evaluated with the Quantra analyzer. These results are consistent with the decline in aggregation and the emergence of procoagulant and apoptotic platelet subpopulations during storage.