The Mechanism of Cold-Induced Platelet Hyperfunction.

Refrigerated platelets function better when transfused than do platelets stored at room temperature, although chilled and rewarmed platelets do not survive long in the circulation. Here, we describe the one possible mechanism of this increase in the hemostatic function of platelets upon cold treatment. Cold exposure has been demonstrated to cluster GP Ib-IX complexes on platelets, enabling platelet recognition and phagocytosis by macrophages. We find that, in addition to increasing the binding to the leukocyte integrin Mac-1 (aMb2), clustering of GP Ib-IX complexes also augments von Willebrand factor (VWF)-related functions. Before the functional studies, platelets were chilled to 4°C for 2 hr and rewarmed to 37°C for 30 min. Chilled, rewarmed platelets, whether in platelet-rich plasma (PRP), whole blood, or reconstituted whole blood (all anticoagulated with Cirtate Sodium) perfused through a parallel-plate flow chamber (36 dynes/cm²) attached in greater numbers to VWF-coated surfaces than did platelets from the same donor kept at 37°C or 22°C. The increased function was also reflected in increased thrombus number and size on a surface of type III collagen. The surface coverage, an indicator of thrombus formation, increased from 10% to 25% upon cold. Chilled, rewarmed platelets also responded to lower doses of a VWF-specific agonist, ristocetin Chilled, rewarmed platelets showed 80% aggregation while control platelet only had 10% aggregation at low concentration of ristocetin (0.5mg/ml) treatment, and displayed a decreased threshold for shear-induced aggregation, aggregating at lower shear rates than control platelets at both 22°C and 37°C. Finally, when tested in a platelet function analyzer (PFA-100), chilled, rewarmed platelets demonstrated significantly shorter closure times than control platelets using both epinephrine/collagen and ADP/collagen cartridges. The epinephrine/collagen cartridges clotting time of chilled platelets decreased from normal 180 s to 162 s and ADP/collagen cartridges clotting time decreased from 118s to 99s. To exclude that the observed functional increases were not due to changes in GP Iba expression or prior platelet activation, we evaluated by flow cytometry the surface levels of GP Iba, P-selectin, and activated α_IIbβ₃.(antibody PAC-1 binding). We found no differences in any of these parameters between chilled, rewarmed platelets and control platelets. These results suggest that, if their rapid clearance can be prevented, cold-stored platelets may have enhanced hemostatic function of potential benefit in the bleeding patient. The results also caution, however, that trials will be necessary to exclude potential thrombotic consequences of infusing hyperfunctional, refrigerated platelets.