Platelet spreading on fibrinogen matrix, a reliable and sensitive marker of platelet functional activity during storage

Upon platelet activation, inside-out signals synergistically induced by a variety of agonists and adhesion molecules can enhance the affinity of platelet main integrin, alpha(IIb)beta(3) to its ligands. Integrin ligation with fibrinogen induces potent signals which develop platelet function including aggregation, release and spreading of which platelet spreading is considered as a major early consequence of alpha(IIb)beta(3) outside-in signaling. Study presented here evaluated platelet spreading on fibrinogen matrix as a marker of platelet activation during storage. PRP-platelet concentrates were subjected to flowcytometry analysis and the expression levels of P-selectin, CD61, GPIb alpha and active conformation of the alpha(IIb)beta(3) (PAC-1 binding) were examined on day 0, 1, 3 and 5 post-storage. Concurrently platelet adhesion and spreading on fibrinogen matrix, glucose concentration and LDH activity were also determined at the same intervals. Results showed significant decreases in platelet spreading on fibrinogen matrix during storage. Spreading was dominant pattern of adhesion till the first day of storage. In 3 day-stored platelets, filopodial or lamellipodial formation was dominant event whereas 5 day-stored platelets simply adhered to fibrinogen with less protrusion formation and partially failed to spread. Compared to simple adhesion, reduction of platelet spreading was also more significantly correlated with the usual markers of platelet storage lesion including P-selectin (r = - 0.88; p < 0.0001) and GPIb alpha expression (r = 0.76; p = 0.0001), PAC-1 binding (r = 0.66; p = 0.001), glucose concentration and LDH activity. Taken together, we introduced platelet spreading on fibrinogen matrix as a reliable and sensitive marker of platelets functional activity during storage. As a valid marker which is directly and obviously relevant to the platelet functional capacities, the rapid reduction of platelet spreading during storage overshadows other markers of platelet storage lesion while raising serious question about the quality of 5 day-stored platelets.