FEINSTEIN INSTITUTE FOR MEDICAL RESEARCH, THE
The purpose of this proposal is to determine the mechanisms for the loss of B cell tolerance to phospholipids and for tissue damage by anti-phospholipid
antibodies in NZW/BXSB F1 (W/B) mice. W/B mice develop both proliferative glomerulonephritis and spontaneous anti-phospholipid syndrome (APS).
Male W/B mice expresses two copies of TLR7 and it is our hypothesis that this results in increased stringency of naïve B cell selection but loss of
tolerance in the antigen activated B cell compartments. T cell help is therefore required to break tolerance in this model but once autoantibodies are
present they may propagate disease by T cell independent mechanisms. We are using a transgenic system in which the 3H9 anti-cardiolipin/anti-DNA
heavy chain is used to analyze the repertoires of B cell subsets; this allows us to determine checkpoints for regulation of anti-cardiolipin antibodies.
There are three specific aims in the parent proposal. In this one year ARRA proposal we will mainly work on Aim 1 and perform some initial
experiments related to Aims 2 and 3. (1) How do TLR7 overexpression and activation alter B cell selection? We are addressing this question using
bone marrow chimeras bearing the 3H9 autoreactive heavy chain gene and by analyzing the B cell repertoire and B cell subset selection of TLR7+/+
and TLR7-/+ cells. (2) How do T cells, BAFF and IFN? interact with TLR7 in loss of tolerance to phospholipids? This year we will mostly study
the effects of T cell depletion in W/B mice. (3) How do anti-phospholipid antibodies mediate their pathogenic effects? We plan to analyze the
pathogenicity of autoantibodies derived from control and treated W/B mice with respect to a) thrombocytopenia, fetal demise and hypercoagulability
and b) ability to induce endothelial cell death via the ceramide pathway. This year we plan to generate and purify the panel of antibodies needed
for this experiment and perform some initial testing in the pathogenicity systems. Our experiments should define the interactions of crucial
pathways that contribute to induction, propagation and clinical outcomes of APS and may suggest new therapeutic strategies for this devastating syndrome.