GDKV-induced antiphospholipid antibodies enhance thrombosis and activate endothelial cells in vivo and in vitro

Antiphospholipid (aPL) Abs are associated with thrombosis, pregnancy loss, and thrombocytopenia in patients with systemic lupus erythematosus or prima ry antiphospholipid syndrome (APS), beta(2)-Glycoprotein I (beta(2)GPI), a phospholipid-binding serum protein, is involved in aPL binding to phospholi pids. aPL can be generated in mice by immunization with beta(2)GPI, and the se Abs are thrombogenic and cause pregnancy loss in mice, The objective of this study is to determine whether aPL induced by immunization with the pho spholipid-binding site of beta(2)GPI are thrombogenic and whether they acti vate endothelial cells (EC) in vivo and in vitro. Murine monoclonal aPL wer e generated from spleen cells of a mouse immunized with GDKV, a synthetic 1 5-aa peptide spanning Gly(274)-Cys(288) in the fifth domain of human beta(2 )GPI, which represents the phospholipid-binding site of beta(2)GPI, The Abs generated had aPL and anti-beta(2)GPI activities. The effect of these Abs on thrombus formation and on EC activation in vivo was determined using a m ouse model of thrombosis and microcirculation that enables examination of t he adhesion of leukocyte to EC as an indication of EC activation as wen as adhesion molecule expression using in vitro ELISA analysis, Mice injected w ith this monoclonal aPL showed a significant increase in leukocyte sticking and also produced larger thrombi that persisted longer, Exposure to GDKV-i nduced aPL fur 4 h significantly increased surface Ag expression of E-selec tin, ICAM-1, and VCAM-1, These data indicate that aPL induced by immunizati on with the phospholipid binding site of beta(2)GPI are thrombogenic and ac tivate endothelial cells.