Intramolecular epitope spreading among anti-caspase-8 autoantibodies in patients with silicosis, systemic sclerosis and systemic lupus erythematosus, as well as in healthy individuals.

Abstract

Dysregulation of apoptosis through the Fas-Fas ligand pathway is relevant in autoimmune disease onset. We recently reported elevated serum levels of sFas in patients with silicosis, systemic sclerosis (SSC) and systemic lupus erythematosus (SLE), and proposed a block of apoptosis in the pathogenesis. The disturbance of apoptosis in lymphocytes including autoreactive clones could induce autoantibody production. Since autoantibodies directed against unknown antigens are present in the sera of these patients, the sera samples were examined for the presence of autoantibodies directed to caspase-8. Using Western blotting, autoantibodies against caspase-8 were detected in healthy individuals and in over 60% of patients. Using epitope mapping employing 12 amino acid polypeptides with SPOTs system, a minimum of 4 epitopes and a maximum of 13 were found, which implied that epitope spreading was in progress. It is noteworthy that two important catalytic cystein residues were included within the epitopes; firstly the active site cystein Cys287, and secondly Cys360 located in the unique pentapeptide motif QACQG. Using recombinant human caspase-8 linked protein chip array, autoantibodies were identified and molecular weight determined. The antibodies were mainly IgG; 80% were subclass IgG1(lambda); 20% were IgG4(kappa). Despite the ratio of human light chain kappa:lambda = 2:1, the predominance of IgG1(lambda) is noticeable. Anti-caspase-8 autoantibodies are detectable in healthy individuals and in patients suffering silicosis, SSc or SLE. A few epitopes were detected in healthy individuals compared to those suffering autoimmune diseases, indicating the intramolecular epitope spreading. Relationship of autoantibodies and the clinical background of the patients requires clarification.