alpha-chymotrypsin and Purpura--Thrombocytopenic

Quinidine-induced thrombocytopenia has been associated with both immune complex and autoantibody binding to platelets. In the present study, serum antibody from six of six patients with quindine purpura was shown by immunoblotting to bind to a single platelet membrane protein of mol wt 80,000. This target protein was absent from Bernard-Soulier (BSS) platelets. F(ab)2 prepared from one patient's serum also bound to this protein, indicating autoantibody rather than immune complex binding to the target antigen. Antibody binding to the 80-kd protein was preserved after treatment of platelets with concentrations of trypsin or chymotrypsin that completely removed glycoprotein Ib (GPIb). Preincubation of platelet proteins with one patient's serum blocked binding of a polyclonal rabbit antibody against glycoprotein V (GPV), indicating that these antibodies recognize the same antigen. By wheat germ affinity chromatography, GPV was shown to copurify with GPIb. Quinidine-induced antibody bound to the wheat germ-purified GPV but not to GPIb. We conclude that quinidine purpura is associated with autoantibody directed against platelet GPV.

Topics: Animals; Autoantibodies; Blood Platelets; Chromatography, Affinity; Chymotrypsin; Glycoproteins; Humans; Immunoglobulin Fab Fragments; Immunosorbent Techniques; Lectins; Membrane Proteins; Molecular Weight; Platelet Membrane Glycoproteins; Purpura, Thrombocytopenic; Quinidine; Rabbits; Trypsin; Wheat Germ Agglutinins

Platelet glycoprotein I (GPI) is known to be required for the interaction of platelets with ristocetin and factor VIII:von Willebrand factor (VIII:vWf). However, its role as Fc receptor is not clear. Some studies have shown that enzymatic removal of GPI destroys the ability of platelets to react with VIII:vWf but not their ability to bind Ig G (IgG). Others have shown that IgG immune complexes which block the Fc receptor also inhibit VIII:vWf interaction with platelets. This subject has been re-examined by testing the ability of platelets with reduced amounts of GPI to aggregate and undergo the release reaction in response to stimuli which act at the platelet Fc receptor. Platelets from two patients with Bernard-Soulier syndrome, congenitally deficient in GPI, both aggregated and released 14C-serotonin normally when exposed to latex particles coated with IgG. Levels of GPI were decreased experimentally in normal platelets by treating them with chymotrypsin. Platelets treated in this manner did not aggregate or release [14C]serotonin in response to ristocetin-VIII:vWf. They did, however, both aggregate and release when incubated with heat-aggregated IgG, antigen-antibody complexes or latex particles coated with IgG. Thus the presence of GPI is not a prerequisite for platelet stimulation via the Fc receptor.

Topics: Blood Platelets; Chymotrypsin; Electrophoresis, Polyacrylamide Gel; Glycoproteins; Humans; Membrane Proteins; Platelet Aggregation; Platelet Membrane Glycoproteins; Purpura, Thrombocytopenic; Receptors, Fc; Serotonin; Syndrome

In summary: Incubation of platelets with ADP or proteolytic enzymes (chymotrypsin or pronase) results in an exposure of two classes of specific binding sites on platelet surface: low and high affinity fibrinogen receptors. Fibrinogen interaction with these receptors results in platelet aggregation. High affinity fibrinogen receptors are not exposed on thrombasthenic platelets stimulated by ADP but are rendered available on chymotrypsin-treated thrombasthenic platelets; low affinity receptors cannot be exposed by ADP or chymotrypsin on these platelets. Availability of high affinity fibrinogen receptors on thrombasthenic platelets may depend on the residual glycoprotein IIIa. Fibrinogen receptors appear to be associated with glycoproteins IIb, IIIa and a 66,000 Mr platelet membrane component that is exposed during proteolysis of platelet membranes. Some of the platelet-binding sites on the fibrinogen molecule appear to be associated with the COOH-terminal portion of the gamma chain (gamma 374-411). Additional binding sites may also be located in the COOH-terminal portion of the A alpha chain. The conformation of the fibrinogen molecule may be important in its interaction with platelets. Platelet aggregation may result from bridging platelets by fibrinogen molecule in the presence of bivalent cations. In conclusion, platelet interaction with fibrinogen is a complex process involving different binding sites of the fibrinogen molecule. Our own data and review of literature suggest that platelet-interaction with fibrinogen is of major significance in hemostasis.

Topics: Adenosine Diphosphate; Blood Platelets; Cell Membrane; Chymotrypsin; Fibrinogen; Glycoproteins; Humans; Platelet Aggregation; Platelet Membrane Glycoproteins; Purpura, Thrombocytopenic; Receptors, Cell Surface; Structure-Activity Relationship; Syndrome