Background/Purpose:

Systemic lupus erythematosus (SLE) is a chronic, complex disease and autoantibodies to self-antigens are a characteristic feature of the disorder. Our group and others have observed free radical mediated oxidative damage in SLE. Our previous data implicated red cell membrane catalase as a target of 4-hydroxy-2-nonenal (HNE; a by-product of oxidative damage) modification in SLE subjects. Here, we tested the hypothesis that HNE- or malondialdehyde- (MDA) modification occurs in the sera of SLE subjects and that antibodies to HNE would be present in SLE sera, and may precede lupus autoantibodies.

Methods:

Serum proteins from 8 SLE subjects and 8 age and sex-matched normal subjects were analyzed for HNE- or MDA protein-adducts by ELISA. Three additional SLE subjects, who had sera collected longitudinally for 12- 15 years, were tested for the presence of anti-HNE antibodies by ELISA. One SLE subject developed anti-Ro60 under observation and another developed anti-P similarly (after disease diagnosis). The third SLE subject had anti-Sm from time of diagnosis. Ro60, Sm and a multiple antigen peptide synthesized from the P autoantigen were coated on ELISA plates, HNE-modified and used as substrate for anti-HNE ELISA. Unmodified Ro60, Sm and P multiple antigen peptide served as control. To test for HNE or MDA-protein adducts in sera from SLE subjects or normal controls, the sera were coated on ELISA plates as antigen. HNE or MDA adducts in sera was determined with rabbit anti-HNE or anti-MDA antibodies. HNE-modified proteins in sera from SLE or normal controls were also studied by immunoblotting.

Results:

We found significantly increased oxidative damage in the sera of SLE subjects compared to normal controls by ELISA and immunoblotting. There was significantly more HNE-protein adducts in SLE sera compared to controls as determined by ELISA (0.084 ± 0.022 versus 0.055 ± 0.012, p=0.0044; average OD±SD). But, we did not observe any significant difference in MDA-modified proteins between SLE subjects and controls. Analysis of SLE sera by immunoblotting showed HNE modification of two proteins migrating at 24 kD and 38 kD respectively. Anti-HNE P autoantibodies preceded anti-P autoantibodies in the SLE subject that developed anti-P antibodies several months after SLE diagnosis. Anti-HNE P antibodies developed strongly in the anti-P SLE subject in the 68^th month (1.2 OD), while anti-P response developed only by month 112. Anti-P response was strong by 125^th month (1.7 OD). Anti-HNE P response at this time point was 3.0 OD, almost twice as the anti-P response. Anti-HNE Ro60 response preceded anti-Ro60 antibody response by 20 months in the subject developing anti-Ro60 under observation. Anti-HNE Ro60 response correlated with anti-Ro60 response in this subject for the rest of the dates for which samples were available for testing. However, there was no difference in anti-Sm or anti-HNE Sm antibody levels over time for the subject that had anti-Sm autoantibodies right from the time of diagnosis.

Conclusion:

Increased HNE-modified protein occur in sera of SLE subjects. Anti-HNE antibody may precede anti-Ro60 or anti-P autoantibodies in SLE. HNE-products are potential neoantigens and thus could be involved in the pathogenesis of SLE.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/4-hydroxy-2-nonenal-serum-protein-adducts-and-anti-4-hydroxy-2-nonenal-protein-adduct-antibodies-in-sle/