Prothrombin binds to the surface of apoptotic, but not viable, cells and serves as a target of lupus anticoagulant autoantibodies.
Anti-phospholipid Ab (aPL) are a heterogeneous group of autoantibodies directed against various combinations of phospholipids (PL) and PL-binding proteins. Lupus anticoagulant (LA) Ab, a subset of aPL, exhibit anticoagulant properties in vitro, but are procoagulant in vivo. Most LA Ab are specific for either beta(2)-glycoprotein I (beta(2)GPI) or prothrombin (PT), two PL-binding proteins. We have previously shown that beta(2)GPI and beta(2)GPI-dependent aPL bind specifically to apoptotic, but not viable, thymocytes. In this study, we demonstrate that PT, like beta(2)GPI, binds selectively to the surface of apoptotic, but not viable, Jurkat cells. Furthermore, PT supports the binding of systemic lupus erythematosus-derived polyclonal and murine monoclonal LA Ab to apoptotic cells. Two LA mAb, which differed dramatically in their relative affinities for PT, were studied. Although one mAb (29J3-62) had a high affinity for PT alone, the other (29I4-24) showed minimal reactivity with PT alone and required PL for elevated binding. Monovalent fragments of 29I4-24 reacted with PL-bound PT with high affinity, suggesting that this mAb recognizes a PL-dependent epitope. Despite these differences, PT-dependent binding of both mAb to apoptotic cells was 30-fold greater than that to viable cells. Moreover, binding of PT to apoptotic cells was, itself, increased in the presence of bivalent, but not monovalent, forms of either mAb. In summary, our data demonstrate the following: 1) specific binding of PT to apoptotic cells, an effect enhanced by PT-dependent LA Ab; 2) heterogeneity of PT-dependent LA Ab; and 3) potential pathogenicity of Ab of either low or high affinity for PT.
D'Agnillo
Paolo
P
Division of Rheumatology, Department of Medicine, Research Institute of the McGill University Health Center, McGill University, Montreal, Quebec, Canada.
Levine
Jerrold S
JS
Subang
Rebecca
R
Rauch
Joyce
J
D000911:Antibodies, Monoclonal / Q000378:metabolism; D000915:Antibody Affinity; D017209:Apoptosis / Q000187:drug effects / Q000276:immunology*; D001666:Binding Sites, Antibody; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G / Q000378:metabolism; D019169:Jurkat Cells; D016682:Lupus Coagulation Inhibitor / Q000378:metabolism*; D008180:Lupus Erythematosus, Systemic / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D008565:Membrane Proteins / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D008856:Microscopy, Fluorescence; D010743:Phospholipids / Q000494:pharmacology; D011485:Protein Binding / Q000276:immunology; D011516:Prothrombin / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D019311:Staurosporine / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D016682:Lupus Coagulation Inhibitor; D008565:Membrane Proteins; D010743:Phospholipids; D011516:Prothrombin; D019311:Staurosporine