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Measurement and Significance of 3‐Nitrotyrosine in Systemic Lupus Erythematosus
Author(s) -
Khan F.,
Siddiqui A. A.,
Ali R.
Publication year - 2006
Publication title -
scandinavian journal of immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.934
H-Index - 88
eISSN - 1365-3083
pISSN - 0300-9475
DOI - 10.1111/j.1365-3083.2006.01794.x
Subject(s) - nitrotyrosine , chemistry , tyrosine , nitration , antibody , bovine serum albumin , immune complex , epitope , biochemistry , microbiology and biotechnology , enzyme , immunology , medicine , biology , nitric oxide synthase , organic chemistry
Nitration of free and protein associated tyrosine represents, in vivo , a mechanism that can severely compromise the cell function. The detection of 3‐nitrotyrosine (3‐NT) in pathological tissues is suggestive of the occurrence of nitrating pathways and has been identified as a marker of inflammation and a stable end product of increased reactive nitrogen intermediate production. Protein nitration occurs in many disease conditions including systemic lupus erythematosus (SLE). In this study we show that the level of both free and protein bound 3‐NT, which is produced by reactive nitrogen species (RNS)‐dependent oxidative damage, is elevated in patients with SLE and that there is a possible role of RNS‐modified epitopes in the aetiology of the disease. Commercially available poly L ‐tyrosine was exposed to nitrating species, inducing nitration in tyrosine residues. Immunoglobulin‐G (IgG) purified on Protein‐A‐Sepharose matrix from 24 SLE patients was studied for their recognition of native and nitrated poly L ‐tyrosine by direct binding and competition enzyme‐linked immunosorbent assay (ELISA). The formation of immune complex between SLE IgG and nitrated poly L ‐tyrosine was visualized by gel retardation assay. Free 3‐NT in patients’ sera was detected and quantitated by high performance liquid chromatography whereas protein‐bound 3‐NT was analysed by Western blotting and the concentration was calculated by sandwich ELISA. The concentration of free 3‐NT was found to be 1.4 ± 0.09 μ m whereas the concentration of protein bound 3‐NT was 96.52 ± 21.12 μ m nitrated bovine serum albumin equivalents/mg protein, which was significantly higher when compared with healthy controls. Elevated level of 3‐NT was observed in SLE patients using two different techniques, when compared with healthy subjects confirms the overproduction of RNS in the pathogenesis of human SLE.