Platelet surface thiols and disulphides play an important role in platelet
responses. Agents that reduce disulphide bonds expose the fibrinogen recept
or in platelets and activate the purified glycoprotein (GP) IIbIIIa recepto
r. Protein disulphide isomerase (PDI), an enzyme that rearranges disulphide
s bonds, is found on the platelet surface where it is catalytically active.
We investigated the role of PDI in platelet responses using (1) rabbit ant
i-PDI IgG specific for PDI, (2) a competing substrate (scrambled ribonuclea
se A), and (3) the PDI inhibitor, bacitracin. Fab fragments of the rabbit a
nti-PDI IgG inhibited platelet responses to the agonists tested (ADP and co
llagen), whereas Fab fragments prepared identically from normal rabbit IgG
had no inhibitory effect. Scrambled ribonuclease A blocked platelet aggrega
tion and secretion, but native ribonuclease A did not. When biphasic platel
et aggregation was examined using platelets in citrated plasma, the princip
le effect of bacitracin was on second phase or irreversible aggregation res
ponses and the accompanying secretion. Using flow cytometry and an antibody
specific for activated GPIIbIIIa (PAC-1), the rabbit anti-PDI Fab fragment
s substantially inhibited activation of GPIIbIIIa when added before, but no
t after, platelet activation. In summary, we have demonstrated that protein
disulphide isomerase mediates platelet aggregation and secretion, and that
it activates GPIIbIIIa, suggesting this receptor as the target of the enzy
me.