Apoptosis, or programmed cell death, is an active process by which unwanted cells are eliminated. The swift removal of apoptotic cells by phagocytes occurs in the absence of inflammation and without provide an autoimmune response. An important mechanism of apoptotic cell clearance involves the cell's loss of membrane phospholipid asymmetry and recognition of phosphatidylserine (PS) but putative macrophage PS receptors. Beta/2-glycoprotein I (beta2GPI), a phospholipid-binding plasma protein, has recently been identified as a major autoantigen in patients with SLE and the anti-phospholipid antibody syndrome. Although its physiological function is not known, there is growing evidence that Beta2GPI, a member of the complement control protein family, may be involved in the opsonization and phagocytosis of apoptotic cells. Preliminary studies demonstrate specific binding of beta2GPI to apoptotic murine splenocytes leading to the hypothesis the beta2GPI may function as a physiological, non-inflammatory opsonin for apoptotic cells, and that anti-beta2GPI antibodies may contribute to the pathogenesis of SLE by altering the normal clearance of apoptotic cells, and that anti-beta2GPI antibodies may contribute to the pathogenesis of SLE by altering the normal clearance of apoptotic cells. The goals of this proposal are to investigate the role of beta2GPI and the effect of anti-beta2GPI autoantibodies, on the phagocytosis of apoptotic cells.
The first aim i s to determine the effect of beta2GPI on the phagocytosis of apoptotic cells. In vitro phagocytosis assays will be performed in serum-containing medium and in serum-free medium in the presence and absence of purified beta2GPI.
The second aim i s to identify and characterize phagocyte cell surface receptors for beta2GPI. The hypothesis that beta2GPI acts as an opsonin for apoptotic cells require recognition for beta2GPI by receptors on phagocytes. Receptor-mediated binding of beta2GPI to certain phagocytes is suggested by a number of recent reports and our preliminary data. Phage display cloning will be used to isolate and identify beta2GPI binding molecules.
The third aim i s to investigate the effect of autoantibodies of beta2GPI apoptotic cell clearance. The effects of polyclonal and monoclonal autoantibodies to beta2GPI will be investigated in in vitro phagocytosis assays and in the in vivo clearance of apoptotic cells. Additional in vivo experiments will investigate the possibility that disease-related anti-beta2GPI autoantibodies could stimulate the development of an autoimmune response to apoptotic cell antigens.
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