Summary Currently, platelets for transfusion are stored at room temperature (RT) for 5–7 days with gentle agitation, but this is less than optimal because of loss of function and risk of bacterial contamination. We have previously demonstrated that cold (4°C) storage is an attractive alternative because it preserves platelet metabolic reserves, in vitro responses to agonists of activation, aggregation and physiological inhibitors, as well as adhesion to thrombogenic surfaces better than RT storage. Recently, the US Food and Drug Administration clarified that apheresis platelets stored at 4°C for up to 72 h may be used for treating active haemorrhage. In this work, we tested the hypothesis that cold-stored platelets contribute to generating clots with superior mechanical properties compared to RT-stored platelets. Rheological studies demonstrate that the clots formed from platelets stored at 4°C for 5 days are significantly stiffer (higher elastic modulus) and stronger (higher critical stress) than those formed from RT-stored platelets. Morphological analysis shows that clot fibres from cold-stored platelets were denser, thinner, straighter and with more branch points or crosslinks than those from RT-stored platelets. Our results also show that the enhanced clot strength and packed structure is due to cold-induced plasma factor XIII binding to platelet surfaces, and the consequent increase in crosslinking.

Currently, platelets (PLTs) stored at room temperature (RT) for 5-7 days with gentle agitation are exclusively used for transfusion although FDA recently clarified that apheresis PLTs stored at 4°C for up to 72 hours may be used for treating active hemorrhage. We have demonstrated that cold (4C) storage of PLT is an attractive alternative to RT storage since it better preserves the PLT metabolic reserves, in vitro responses to agonists of activation, aggregation and physiologic inhibitors, as well as adhesion to thrombogenic surfaces. In this study, we tested the hypothesis that 4C-stored PLT will form clots with mechanical strength superior to those from RT-stored PLT due to higher hemostatic potential. From rheological measurements, we observed that the clots formed from 5 day 4C-stored PLTs are significantly stiffer (elastic modulus) and stronger (critical stress) than those formed from RT-stored PLT but comparable to fresh PLT (Fig. A). We also observed from ultrastructural microscopy that the fibrin fibers in clots from cold-stored PLT were thinner with more branch points than those from RT-stored PLTs, indicating the presence of increased crosslinks (Fig. B, C). Finally, molecular analysis revealed an increase FXIII transglutaminase activity due to the binding of plasma FXIII/fibrinogen to the surface of 4C-stored PLTs (Fig D).In conclusion, we have shown that cold-induced plasma FXIII binding to PLT surface result in increased fibrin crosslinking and enhanced clot strength. Our data, together with the benefit of reduced risk of late thrombosis due to their rapid clearance in vivo , underscores the consideration of 4C-stored PLT for acute response to hemorrhage. Figure 1 (A) RT-stored platelets form clots with less stiffness (n=5); (B) Representative SEM images of clots (n=3) (C) Clots from 4C-stored platelets have higher cross-linking density (n=5) (D) In FXIII-deficient plasma, 4C-stored platelets form clots with higher stiffness than fresh platelets (n=4)