RESUMEN
In quinine- and quinidine-induced thrombocytopenic purpura IgG antibodies are known to react in a drug-dependent manner with different combinations of surface glycoproteins (GP) Ib, IIb, IIIa and IX. Because endothelial cells share a number of properties of platelets, including the presence of GP IIIa and GPIb-like proteins, we have compared these two cell types for their quinine-dependent IgG binding abilities. By immunoblotting of endothelial cells, quinine-dependent IgG binding from four patient sera was observed only to a 93 kDa component corresponding to GP IIIa. Strong IgG binding independent of the drug was found at 170-180 kDa. Thus endothelial cells express the GP IIIa quinine-dependent epitope on platelet GP IIIa, but not those on other platelet glycoproteins.
Asunto(s)
Autoanticuerpos/inmunología , Autoantígenos/biosíntesis , Plaquetas/inmunología , Endotelio Vascular/inmunología , Glicoproteínas de Membrana Plaquetaria/inmunología , Quinina/farmacología , Plaquetas/efectos de los fármacos , Humanos , Immunoblotting , Inmunoglobulina G/inmunología , Glicoproteínas de Membrana Plaquetaria/efectos de los fármacosRESUMEN
Previously described platelet-aggregating antibodies associated with thrombosis and thrombocytopenia required heparin for their in vivo and in vitro expression. We have observed a patient with thrombosis who became thrombocytopenic during heparin treatment, but who suffered further thrombotic events and continued thrombocytopenia for 3 months after heparin withdrawal. The patient's plasma contained a potent platelet aggregating factor reactive with both his own and normal platelets in the absence of heparin. It also caused [14C]serotonin secretion from labelled platelets from normal donors and patients with either Glanzmann's thrombasthenia or Bernard-Soulier syndrome. This factor was an IgG and was neutralized by antibody specific for IgG lambda light chains. While the patient was thrombocytopenic an IgG paraprotein with lambda light chains was detected by isoelectrofocussing. After corticosteroid treatment it disappeared and the patient recovered. The active, but not the recovery serum contained IgG which immunoprecipitated a glycoprotein with characteristics of Glycoprotein IV from platelets labelled with Na[3H]BH4/periodate. Thus platelet-aggregating IgG antibodies with direct specificity for platelet surface glycoproteins may be associated with thrombosis/thrombocytopenia.
Asunto(s)
Inmunoglobulina G/fisiología , Embolia y Trombosis Intracraneal/inmunología , Agregación Plaquetaria/inmunología , Glicoproteínas de Membrana Plaquetaria/inmunología , Trombocitopenia/inmunología , Humanos , Embolia y Trombosis Intracraneal/complicaciones , Masculino , Persona de Mediana Edad , Trombocitopenia/complicacionesRESUMEN
Immunoblotting of platelets that have been subjected to SDS-PAGE has revealed that sera from normal individuals contain IgG which binds to many platelet components. This binding was seen with autologous and heterologous platelets using serum of males and of nulliparous females who had not received blood transfusions. Although binding patterns of different sera were not identical, almost all sera caused IgG binding to platelet components of 87-90 kD, 140 kD (identified as vinculin) and 220-240 kD (tentatively identified as talin and actin-binding protein). Purified IgG showed the same binding pattern as whole serum and F(ab')2 fragments retained their ability to bind to many components. The titre of IgG binding in serum was 1:50-1600 while that of alloantibodies to the PlA1 antigen was 1:3200. IgG binding components were not secreted when platelets were stimulated and were rarely associated with isolated membranes, but were located either in platelet cytoplasm or cytoskeletons. IgG binding was decreased by absorbing sera with lysed platelets or isolated cytoskeletons, but only slightly with intact platelets. Microaffinity purification of IgG which formed a major band on immunoblots showed that it was antibody with specificity for vinculin or its degradation products. These findings suggest that normal sera contain naturally occurring IgG antibodies with specificity for intracellular platelet antigens and that in some cases their titre approaches that of antibodies of pathological significance.
Asunto(s)
Autoanticuerpos/inmunología , Plaquetas/inmunología , Citoesqueleto/inmunología , Inmunoglobulina G/inmunología , Membranas Intracelulares/inmunología , Adulto , Especificidad de Anticuerpos , Plaquetas/ultraestructura , Proteínas del Citoesqueleto/inmunología , Humanos , Immunoblotting , Valores de Referencia , VinculinaRESUMEN
Reactivity of quinine- and quinidine-dependent antiplatelet antibodies has been compared in platelet-rich-plasma (PRP) from normal donors and from patients with von Willebrand's disease (vWd). One quinine-dependent antibody (Q.Ab) caused platelet aggregation and [14C] serotonin release with only 7 of 12 normal donors, while another Q.Ab and a quinidine-dependent antibody (Qd.Ab) caused aggregation and release with all 12. Drug-dependent IgG binding and PF 3 availability induced by the antibodies were, however, comparable in all donors. Differences in responsiveness were associated with platelets and not plasma. vWd platelets showed normal drug-dependent IgG binding, but decreased aggregation and serotonin release to most drug-dependent antibodies. Responsiveness was not restored by purified vWf:Ag, but, in one case, was corrected by normal plasma or cryoprecipitate. Drug-dependent binding of the Q.Ab which caused variable responsiveness in normals was to the same platelet antigens (GPIb and GPIIIa) in both normal and vWd platelets and did not require plasma components. Reduced PF 3 availability was seen with some antibodies in some vWd patients. Plasma from two of these patients inhibited aggregation of normal platelets to Q.Ab and one of these inhibited aggregation to ADP. Antiplatelet antibodies were detected in these two plasmas by ELISA. Thus some Q.Ab produce different responses with platelets from different donors. In vWd, reduced responsiveness to Q.Ab and Qd.Ab may result from production of inhibitory antiplatelet antibodies.
Asunto(s)
Autoanticuerpos/inmunología , Plaquetas/inmunología , Quinidina/farmacología , Quinina/farmacología , Enfermedades de von Willebrand/inmunología , Plaquetas/metabolismo , Antígenos HLA/análisis , Humanos , Immunoblotting , Inmunoglobulina G/metabolismo , Agregación Plaquetaria , Factor Plaquetario 3/metabolismo , Serotonina/sangre , Enfermedades de von Willebrand/sangre , Factor de von Willebrand/aislamiento & purificaciónRESUMEN
The molecular nature of platelet receptors for quinine- and quinidine-dependent antiplatelet antibodies (Q.Ab and Qd.Ab) was studied by immunoblotting. One Q.Ab caused quinine-dependent IgG binding to platelet proteins with molecular weights (mol wts) of 174 Kd and 93 Kd and another to only a 93-Kd protein. A third Q.Ab caused binding to 174-, 140-, 93-, and 57-Kd proteins, while a fourth Q.Ab and a Qd.Ab caused IgG binding to 174- and 18-Kd proteins. Using platelets from patients with Glanzmann's thrombasthenia or Bernard Soulier syndrome and purified GPIIIa, these proteins were shown to be GPIb, GPIIb, GPIIIa, GPIX, and an unidentified 57-Kd protein missing in Bernard Soulier syndrome. Binding to the 93-Kd protein was independent of the PIA1 antigen. Absorption of one Q.Ab with Glanzmann's thrombasthenia platelets revealed different populations of antibodies with different specificities within the one patient. Thus Q.Ab and Qd.Ab are heterogeneous and may be directed toward different epitopes on major platelet glycoproteins.
Asunto(s)
Isoanticuerpos/análisis , Isoantígenos/análisis , Glicoproteínas de Membrana Plaquetaria/metabolismo , Quinidina/efectos adversos , Quinina/efectos adversos , Trombocitopenia/sangre , Antígenos de Diferenciación/análisis , Sitios de Unión de Anticuerpos/efectos de los fármacos , Trastornos de las Plaquetas Sanguíneas/sangre , Trastornos de las Plaquetas Sanguíneas/inmunología , Plaquetas/efectos de los fármacos , Plaquetas/inmunología , Plaquetas/metabolismo , Western Blotting , Humanos , Fragmentos Fab de Inmunoglobulinas/metabolismo , Inmunoglobulina G/metabolismo , Isoantígenos/inmunología , Glicoproteínas de Membrana Plaquetaria/deficiencia , Glicoproteínas de Membrana Plaquetaria/aislamiento & purificación , Receptores Fc/análisis , Receptores de IgG , Trombocitopenia/inducido químicamente , Trombocitopenia/inmunologíaRESUMEN
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.