Date of Award
Doctor of Philosophy
Autoimmune thrombocytopenia occurs in a number of clinical settings including idiopathic or autoimmune thrombocytopenic purpura (ITP), systemic lupus erythematosus (SLE), and the lupus anticoagulant (LAC) syndrome. The etiology of the disease, the scope of the autoantigens targeted, and the epitopes involved in autoimmune thrombocytopenia are largely unknown. ITP has the hallmark of the presence of 'pathogenic' autoantibodies targeting platelet glycoproteins (GP) IIb/IIIa and GP Ib/IX. Associated with the thrombocytopenia in the LAC syndrome is the presence of anti-phospholipid antibodies (approximately 95% of cases). The detection of autoantibodies to a variety of self antigens in non-autoimmune individuals prompted this study to determine if normal lymphocytes have the potential to produce antiplatelet autoantibodies.;IgM antibodies to platelets were produced after somatic cell hybridization of tonsil lymphocytes from a non-thrombocytopenic male child to the lymphoblastoid fusion partner GM 4672. These autoantibodies appeared to segregate into two groups; those reactive to platelet proteins (8/13) as demonstrated by radioimmunoprecipitation and immunoblotting and those negative using these assays but reactive with DNA and anionic phospholipids (5/13). One hybridoma, termed STO 171, reacted to platelet GP IIb.;The immunoglobulin variable (Ig V) regions of two of these hybridoma monoclonal antiplatelet antibodies, representative of the two groups of autoreactivity and similar in specificity to autoantibodies from the two disease states, were sequenced at the nucleotide level. Both of these antibodies were generally comprised of unmutated VH, JH, VK and JK germline genes. STO 103 reactive to anionic phospholipids uses the VH4 V71-2 germline gene and a truncated JH4 germline gene. The light chain showed closest homology with the VK4 K18 (99.6% homologous) and JK2 germline genes. STO 171 showed closest homology with the VH4.18 germline gene (98.5% homologous) and a JH6 germline gene. The light chain of STO 171 uses the VK3 Humvk325 germline and a JK4 gene. However, both antibodies had unique diversity (D) regions with evidence of N-nucleotide additions and showed somatic mutation at the VK-JK junctional regions.;Sequence comparisons of these two antiplatelet antibodies to other autoantibodies have revealed that these antibodies are encoded by germline elements that have been used for autoantibodies derived from autoimmune patients and for allotypic antibodies to blood cell antigens. The relationship between the antiplatelet autoantibodies and other pathogenic antibodies and antibodies to foreign antigens remains to be determined.
Denomme, Gregory Andrew, "The Molecular Characterization Of Human Monoclonal Antiplatelet Autoantibodies" (1993). Digitized Theses. 2219.