Monoclonal antibodies bound to subunits of the integrin GPIIb-IIIa are internalized and interfere with filopodia formation and platelet aggregation.
The monoclonal antibodies Tab and AP3 are directed, respectively, against GPIIb and GPIIIa, the subunits of the platelet fibrinogen receptor. When added together to platelets, these antibodies prevent adenosine diphosphate (ADP)-induced platelet aggregation, despite normal fibrinogen binding (Newman et al, Blood 69:668, 1987). To explore the cellular requirements of aggregation after fibrinogen binding, we used several techniques to study platelets treated with Tab and AP3, then stimulated with ADP. We used scanning and transmission electron microscopy to evaluate platelet morphology, immunolabel-surface replication to determine whether individual GPIIb-IIIa complexes clustered, immunocytochemistry on frozen thin sections to study the subcellular distribution of the integrin GPIIb-IIIa and fibrinogen, and biochemical methods to assess the activation of the platelet cytoskeleton. We found that the treated cells had short, blunted projections instead of normal filopodia. Other morphologic abnormalities, apparent in thin section, were aberrantly placed alpha-granules and microtubules, and a prominent, worm-like, fibrinogen-filled surface-connected canalicular system. Biochemical analysis suggested that such platelets undergo massive actomyosin-controlled membrane flow, which serves to sequester GPIIb-IIIa and makes the platelets refractory to aggregation. We conclude that aggregation requires the formation of long, slender filopodia, probably directed by cytoskeletal rearrangements after activation, and that the transmembrane GPIIb-IIIa complex may play a role in signaling these events.
Isenberg
W M
WM
Department of Pathology, University of California, San Francisco 94143-0506.
Bainton
D F
DF
Newman
P J
PJ
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D003599:Cytoskeleton / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D010980:Platelet Membrane Glycoproteins / Q000276:immunology / Q000502:physiology*; D013917:Thrombin / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D010980:Platelet Membrane Glycoproteins; D000244:Adenosine Diphosphate; D013917:Thrombin