B. G. Firkin

Summary 1. The antibotic, Ristocetin, causes precipitation of fibrinogen from platelet poor plasma, as well as platelet aggregation. 2. The precise mechanism of Ristocetin’s effect on platelets has not been elucidated, but it has been shown to initiate ADP release, which may contribute in part to its aggregating ability. 3. Ristocetin has been shown to produce aggregation in platelet rich plasma from thrombasthenic patients. 4. Three patients with Von Willebrand’s disease were examined. In 2, Ristocetin caused no platelet aggregation whatsoever, whilst the third aggregated normally. It is suggested, on this basis, that Von Willebrand’s disease may be subdivided into two types and that Ristocetin could prove to be a valuable technique for further study of this group of disorders.

Abstract Earlier studies have shown that patients with von Willebrand’s disease are considered to have normal platelets, but they lack at least one plasma protein. Results are presented indicating that ristocetin, known to aggregate normal platelets, fails to cause platelet aggregation in the group of von Willebrand’s disease patients exhibiting no platelet adhesiveness. It is postulated that there may be two groups of patients within von Willebrand’s disease and, further, that ristocetin will provide a useful approach to study the plasma deficiency or abnormality in von Willebrand’s disease.

Abstract Surgically removed spleens from 14 subjects were perfused to determine their platelet content. Some patients had Cr51 labeled platelets prior to surgery, and radioactive platelet recoveries were also obtained. A sizable pool of splenic platelets has been demonstrated, which seems dependent on splenic size only. Increase in size of this pool in splenomegaly (and not suppression of thrombopoiesis) may be the sole reason for thrombocytopenia. A dynamic exchange between splenic and circulating platelets appears to be present. Awareness of the existence of a splenic platelet pool is important in considering variation in platelet survival and other platelet dynamic studies, especially in the presence of splenomegaly.

von Willebrand factor (vWF) is a multimeric glycoprotein that mediates the adhesion of platelets to the subendothelium by binding to platelet glycoprotein Ib. For human vWF, this interaction can be induced in vitro by the antibiotic ristocetin or the snake venom protein botrocetin. A missense mutation, Gly-561-->Ser, was identified within the proposed glycoprotein Ib binding domain of vWF in the proband with von Willebrand disease type B, a unique variant characterized by no ristocetin-induced, but normal botrocetin-induced, binding to glycoprotein Ib. The corresponding mutant recombinant protein, rvWF(G561S), formed normal multimers and exhibited the same functional defect as the patient's plasma vWF, confirming that this mutation causes von Willebrand disease type B. These data show that botrocetin and ristocetin cofactor activities of vWF can be dissociated by a point mutation and confirm that these mediators promote vWF binding to platelets by different mechanisms. The normal botrocetin-induced binding and the defective ristocetin-induced binding of rvWF(G561S) suggest that the primary defect in von Willebrand disease type B may be a failure of normal allosteric regulation of the glycoprotein Ib binding function of vWF.