Immunological comparisons of integrin alpha IIb beta 3 (GPIIb-IIIa) expressed on platelets and human erythroleukemia cells: evidence for cell specific differences.
Platelet glycoprotein IIb-IIIa (GPIIb-IIIa, alpha IIb beta 3) is expressed on the cell surface of the human erythroleukemia (HEL) cell line. Previous studies have demonstrated differences in GPIIb-IIIa ligand binding properties of HEL cells when compared to platelets. Although the mRNA sequences for GPIIb and GPIIIa are identical in platelets and HEL cells, cell specific differences in the conformation states of the GPIIb-IIIa complex may exist and may explain in part the contrasting functional properties. Two monoclonal antibodies (mAbs), an anti-GPIIb mAb C3 and an anti-GPIIIa mAb D3, were used to determine whether differences in GPIIb-IIIa conformational states could be measured. Initial studies in a purified system showed that the mAbs' binding to isolated GPIIb-IIIa conformers was increased to the active GPIIb-IIIa and to dissociated receptor subunits when compared to the inactive form. Furthermore, soluble active GPIIb-IIIa was a much better inhibitor of D3 binding to the immobilized receptor compared to soluble inactive GPIIb-IIIa. Extending these studies with intact cells, we detected at least two classes of binding sites for each mAb on each cell type. Differences in Bmax and in the relative affinities of the mAbs were identified and may represent subpopulations of GPIIb-IIIa conformations. Total HEL cell and platelet GPIIb-IIIa was determined in our binding assays using a radiolabeled GPIIb-IIIa complex specific mAb, 10E5. HEL cells express approximately five times more GPIIb-IIIa on a per cell basis. The percent of total GPIIb-IIIa that represented each class of mAb binding sites was determined. In summary, the relative differences in GPIIb-IIIa conformation found on platelets and HEL cells may be related to cell-specific ligand binding properties and activation states of the receptor.
Jennings
L K
LK
Department of Medicine, University of Tennessee, Memphis, USA. ljennings@utmem1.utmem.edu
Slack
S M
SM
Wall
C D
CD
Mondoro
T H
TH
D000704:Analysis of Variance; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute / Q000097:blood / Q000235:genetics*; D009842:Oligopeptides / Q000378:metabolism; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured
D009842:Oligopeptides; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; C046617:arginyl-glycyl-aspartyl-serine