Induction of anti-Ro60/anti-La by immunisation with spectrin and induction of anti-spectrin by immunisation with Ro60 and 4-hydroxy-2-nonenal-modified Ro60 immunisation

• Published in: Clinical and experimental rheumatology 2012-11, Vol.30 (6), p.886-893
• Main Authors: KURIEN, B. T, DORRI, Y, BACHMANN, M, SCOFIELD, R. H
• Format: Article
• Language: English
• Subjects: Aldehydes - administration & dosage; Aldehydes - immunology; Animals; Antibodies, Antinuclear - blood; Autoantigens - immunology; Autoimmunity; Binding, Competitive; Biological and medical sciences; Diseases of the osteoarticular system; DNA - metabolism; Enzyme-Linked Immunosorbent Assay; Epitope Mapping; Epitopes; Immunization; Medical sciences; Protein Binding; Rabbits; Ribonucleoproteins - administration & dosage; Ribonucleoproteins - immunology; Sarcoidosis. Granulomatous diseases of unproved etiology. Connective tissue diseases. Elastic tissue diseases. Vasculitis; Spectrin - administration & dosage; Spectrin - immunology; SS-B Antigen
• ISSN: 0392-856X; 1593-098X

The Ro ribonucleoprotein particle, targeted in systemic lupus erythematosus (SLE) and Sjögren's syndrome (SS), includes Ro60 (SSA) and La (SSA) autoantigens. Anti-Ro60 occurs in SLE and SS. The importance of α-fodrin and spectrin as well as anti-Ro and anti-fodrin/spectrin antibodies in SS and SLE, led us to hypothesise that rabbit immunisation with Ro60 or 4-hydroxy-2-nonenal-modified Ro60 would induce anti-spectrin. In addition, we hypothesised that antibodies to Ro60 and La will develop in animals immunised with spectrin. Two NZW rabbits each were immunised with 4-hydroxy-2-nonenal-modified Ro60 or unmodified Ro60. Methods used included ELISA, including an inside-out RBC membrane ELISA, and Crithidia lucilae assays. Commercial anti-spectrin sera bound significantly to Ro60 (OD 2.6 ± 0.1), Ro60 multiple antigenic peptides (MAPs) (3 out of 21 Ro60 MAPs), La (OD 4.4±0.5), and La fragments as well as to double stranded DNA but not to BSA (OD 0.6±0.1). Anti-spectrin binding to purified spectrin could be inhibited by spectrin (>95%), and Ro60 or La (70%). When the binding of anti-spectrin was tested against a nested set of La fragments we found that a N4 fragment representing the C-terminal 250 aa (aa 159 to 408) bound the strongest (OD=4.12) followed by a N9 fragment (the C-terminal 36aa; aa373 to 408 (OD=1.36). Also, significant anti-spectrin antibody levels were induced by Ro60 and HNE-modified Ro60 immunisation. We found intermolecular epitope spreading from Ro60/La to spectrin and vice versa, and this may have pathological significance in these animal models of autoimmunity.