Autoantibodies against acidic phospholipids (aPLs) are associated with thrombosis and recurrent pregnancy loss. Although aPLs are believed to directly cause these disorders through inappropriate activation of clotting this assumption is not clearly supported or refuted by any published data. Likewise, there is considerable debate concerning the relevant specificities of aPLs and their mechanisms of action, as well as why individual patients, with apparently identical aPL specificities, present with a large difference in symptoms. Our central hypothesis is that aPLs are directly responsible for pregnancy loss. Our preliminary data would further lead us to propose three testable specific hypotheses. The first is that aPLs are heterologous in specificity, the nature of which determines the severity of the particular patient's clinical symptoms. The second hypothesis is that three cell types may be principal targets of aPLs; the vascular endothelial cell, the platelet, and the trophoblast. Which of these cells is primarily affected depends upon the collective specificities of that particular patient's aPLs. The third hypothesis is that aPLs will directly induce reproductive failure in experimental animals. In order to test these hypotheses five specific aims have been designed. First, using monoclonal antibodies and purified antibodies from patients we will evaluate the heterogeneity of aPLs through blocking studies in ELISAs, adsorption with known phospholipids, reactivity with artificial membranes, elution profiles on phospholipid affinity columns, and comparison of represented idiotypes. Second, we will investigate the effects of aPLs on platelet activation. Using flow cytometry and monoclonal antibodies against platelet specific antigens and platelet activation antigens, as well as in vitro measurement of thromboxane production, we will measure the reactivity of aPLs on resting and activated platelets, determine whether aPLs directly activate platelets or enhance the response of platelets to other activators, and evaluate the level of platelet activation in situ. Third, we will evaluate the reactivity and effects of aPLs on cultured bovine and human endothelial cells, including their binding to resting and activated endothelial cells, their effect on endothelial cell metabolism, and the characterization of endothelial cell antigens reactive with aPLs. Fourth, using placental preparations, isolated cytotrophoblast, and a choriocarcinoma line, BeWo, we will investigate the binding and effect of aPLs, including effects on in vitro cytotrophoblast intercellular fusion and overall metabolic activity. Fifth, using pregnant mice and guinea pigs we will test the direct biological effects of aPLs on pregnancy loss. The product of this study will be an in vitro and in vivo analysis of aPL specificity and function in relationship to pregnancy loss.
Showing the most recent 10 out of 23 publications
