Interactions Of Platelets With Proteins Of Plasma And Matrices

We have been analyzing the interactions of human blood platelets with plasma proteins arid with proteins of extracellular matrices (collagen, fibronectin etc.) using a variety of techniques. Using a modified enzyme-linked immunosorbent assay (ELISA), as few as 5.10³ bound platelets could be determined quantitatively and proteins bound from solutions could be detected. Using this method the interaction of the plasma proteins fibronectin, FVIII-VWF and fibrinogen with one another, with components of the basement membrane and with the blood platelet were studied. Antibody against carefully washed human platelets recognizes fibronectin, FVIII-VWF, and fibrinogen as well as platelet surfaces. However, specific antisera to these three proteins fail to bind to the surface of gel-filtered, unactivated platelets. When gel-filtered platelets adhere to plastic in the absence of plasma proteins, they spread. Such platelets do react with antibodies to fibronectin, FVIII-VWF, and fibrinogen. These results suggest that these three plasma proteins are found inside platelets but not on their surfaces prior to activation and that they become exposed upon spreading. Furthermore, these proteins are undetectable on the surfaces of unactivated platelets by surface iodination followed by immune precipitation. We have also studied the interactions of platelets with various collagen substrata by Immunofluorescence microscopy which shows recruitment of plasma fibronectin by platelets. These findings confirm and extend recent suggestions concerning the associations of fibronectin, FVIII-VWF and fibrinogen with human platelets. Finally, we are using chemical crosslinking reagents to analyze the same protein-protein and protein-platelet interactions.