Development of a non-human primate sub-clinical model of heparin-induced thrombocytopenia: platelet responses to human anti-heparin-platelet factor 4 antibodies.
The purpose of this study was to characterize the responses of human and non-human primate (Macaca mulatta) platelets to anti-heparin-platelet factor 4 (AHPF4) antibodies. Due to the variations observed in the functionality and immunoglobulin isotypes in patients with heparin-induced thrombocytopenia (HIT), we used highly characterized human AHPF4 antibodies to study platelet activation responses. Using ELISA and 14C-serotonin release assay (SRA) systems, three patients' plasmapheresis fluid with similar responses to these assays were pooled. This pool was then used to study the platelet activation responses of human and primate platelets in the HIT platelet aggregation assay, a flow cytometry assay, and a variation of the aggregation assay in which glycoprotein IIb/IIIa inhibitors were supplemented. In the plasmapheresis fluid from three patients, the most significant AHPF4 immunoglobulin isotype present (based on optical density readings) was IgG, with less IgM (p < 0.001) and IgA (p < 0.001). The SRA yielded equivalent platelet activation results in all three patients. Using this pool in the platelet aggregation assay, without any heparin present, there was less percent aggregation (p < 0.001) with human platelets (11.8 +/- 2.35, n = 5) compared to the primate platelets (54.3 +/- 10.2, n = 9). In presence of 0.4 U/ml heparin, both platelet types had similar percent aggregations (p > 0.05). Three glycoprotein IIb/IIIa receptor inhibitors were used to evaluate the similarities in platelet activation. Eptifibatide was found to be a strong inhibitor of both species' platelet types at concentrations greater than 0.01 microg/ml. This was not the case with tirofiban which inhibited both human and monkey platelets at concentrations greater than 0.025 microg/ml. Abciximab inhibited aggregation at concentrations greater than 6.25 microg/ml. These data indicate that phylogenetic similarities in platelets of humans and primates may be used to further characterize the pathophysiology of HIT syndrome.
Department of Pathology, Stritch School of Medicine, Loyola University of Chicago, Maywood, IL 60153, USA.
Brian
B
Untch
Sarfraz
S
Ahmad
Harry L
HL
Messmore
Christopher L
CL
Schultz
Qing
Q
Ma
Debra A
DA
Hoppensteadt
Jeanine M
JM
Walenga
Jawed
J
Fareed
D000077284:Abciximab; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000918:Antibody Specificity; D001323:Autoantibodies / Q000276:immunology / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D000077542:Eptifibatide; D006493:Heparin / Q000276:immunology / Q000378:metabolism / Q000633:toxicity*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000494:pharmacology; D008253:Macaca mulatta; D023421:Models, Animal; D010455:Peptides / Q000494:pharmacology; D010802:Phylogeny; D010956:Plasmapheresis; D015539:Platelet Activation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D010978:Platelet Factor 4 / Q000276:immunology* / Q000378:metabolism; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000037:antagonists & inhibitors; D016553:Purpura, Thrombocytopenic, Idiopathic / Q000097:blood / Q000139:chemically induced* / Q000276:immunology / Q000628:therapy; D012701:Serotonin / Q000378:metabolism; D013045:Species Specificity; D000077466:Tirofiban; D014443:Tyrosine / Q000031:analogs & derivatives / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D007140:Immunoglobulin Fab Fragments; D010455:Peptides; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D012701:Serotonin; D010978:Platelet Factor 4; D014443:Tyrosine; D006493:Heparin; D000077466:Tirofiban; D000077542:Eptifibatide; D000077284:Abciximab