ANTICOAGULANT EFFECT OF THE HUMAN SECRETORY PHOSPHOLIPASE A(2) ON BLOOD-PLASMA AND ON A CELL-FREE SYSTEM IS DUE TO A PHOSPHOLIPID-INDEPENDENT MECHANISM OF ACTION INVOLVING THE INHIBITION OF FACTOR VA

Blood platelets play a central role in haemostasis by leading to plug formation and by increasing the efficiency of blood coagulation. We ha ve previously shown that blood platelets contain a group II secretory phospholipase A(2) (sPLA(2) grII) which is released into the extracell ular medium upon activation but is unable to stimulate blood platelets . We presently reported an investigation of the putative involvement o f the human sPLA(2) grII (hsPLA(2) grII) in the coagulation process, b oth in the absence and in the presence of activated platelets. We show that this enzyme prolongs the recalcification time of blood plasma ev en in the presence of activated platelets. The positive action of bloo d platelets on coagulation is correlated, at least in part, with the a ppearence at the cellular surface of procoagulant phospholipids which constitute a potential target for hsPLA(2) grII. We therefore investig ated the involvement of its enzymatic activity in the anticoagulant ef fect of this enzyme. We observed that the replacement of CaCl2 by SrCl 2 to initiate the coagulation cascade did not suppress, but rather inc reased, the inhibitory action of hsPLA(2) grII. Moreover, hsPLA(2) grI I hydrolyzed only a minor proportion of platelet phospholipids, and it did not affect plasma phospholipids. Taken together, these observatio ns strongly suggest that the major action of hsPLA(2) grII on blood co agulation does not involve the hydrolysis of phospholipids, in contras t with the strong anticoagulant effect of the group II venom phospholi pase A(2) from Crotalus durrissus terrificus. We next studied which st ep of the coagulation cascade was affected by hsPLA(2) grII. Using pur ified coagulation factors, we demonstrated that hsPLA(2) grII strongly inhibited the prothrombinase activity. This inhibitory effect was ind ependent of the presence of phospholipids but required factor Va, lead ing to the hypothesis that hsPLA(2) grII inhibited this factor. Furthe r, the anticoagulant effect of hsPLA(2) grII was observed on normal an d factor-X-deficient plasma, but not on factor-V-deficient plasma. In conclusion, the anticoagulant action of hsPLA(2) grII is based on a no nenzymatic mechanism of action involving the inhibition of factor Va.