Elsevier

Life Sciences

Volume 75, Issue 22, 15 October 2004, Pages 2633-2642
Life Sciences

Immunogenicity of superoxide radical modified-DNA: studies on induced antibodies and SLE anti-DNA autoantibodies

https://doi.org/10.1016/j.lfs.2004.04.034Get rights and content

Abstract

Superoxide anion radical (SAR) is formed in almost all aerobic cells and it is the most abundant species generated by several enzymatic and non-enzymatic pathways in mammalian tissues, leading to unfavorable alteration of biomolecules including DNA. The SAR-modified macromolecules have been implicated in several disease states including disorders of inflammation. The SAR-induced damage to DNA showed hyperchromicity, single strand breaks, decrease in melting temperature, and modification of bases. Superoxide modified-DNA in rabbits elicited high titer antibodies and showed diverse antigens binding characteristics. The induced antibodies recognized native DNA and other nucleic acid polymers. Anti-DNA IgG from SLE sera, purified on Protein-A-Sepharose matrix, exhibited increased recognition of superoxide anion radical modified-DNA than native DNA in competitive immunoassay. The visual formation of immune complex between induced antibodies and native DNA, and between SLE anti-DNA IgG and superoxide modified-DNA, is a clear indication of property sharing between SLE autoantibodies and experimentally induced antibodies against superoxide modified-DNA.

Section snippets

Materials and methods

Calf thymus DNA, nuclease S1, riboflavin, anti-human and anti-rabbit IgG alkaline phosphatase conjugates were purchased from Sigma Chemical Company, U.S.A. Nitrobluetetrazolium (NBT) was from Loba-Chemie, India. Polystyrene flat-bottom ELISA plates (96 wells) were obtained from NUNC, Denmark. All other chemiclas used were of highest purity available in the country.

Superoxide radical induced modification of DNA

Exposure of riboflavin to white fluorescent light generates highly reactive superoxide anion radical (SAR). Its formation was confirmed by the reduction of NBT as observed by increase in absorbance at 560 nm (data not shown). The optimum generation of superoxide radical was observed at 20 min of illumination (1.65 absorbance unit at 560 nm). In presence of 10 μg/ml of superoxide dismutase the formation of superoxide radical was significantly dropped as observed by decrease in absorbance from

Discussion

The aim of this study was to compare the antigen binding characteristics of SLE anti-DNA autoantibodies with experimentally induced antibodies against superoxide radical modified-DNA. Production of superoxide anion radical in mammalian tissues by enzymatic and non-enzymatic pathways is well documented (Min et al., 2002). These radicals have been implicated in several disease states and often produce undesirable changes in DNA and proteins.

In present communication, DNA was exposed to superoxide

Acknowledgements

Authors are grateful to Professor R. Ali for critical reading of the manuscript. This study was supported in part by a research grant [37(980)/98/EMR-II] to KA from the Council of Scientific and Industrial Research, New Delhi.

References (31)

  • H. Ahsan et al.

    Oxygen free radicals and systemic autoimmunity

    Clinical and Experimental Immunology

    (2003)
  • C. Aitcheson et al.

    Antinuclear antibodies

  • K. Alam et al.

    Human anti-DNA autoantibodies and induced antibodies against ROS-modified DNA show similar antigenic binding characteristics

    Biochemistry and Molecular Biology International

    (1999)
  • R. Ali et al.

    Evaluation of antibodies against oxygen free radical-modified DNA by ELISA

  • R.W. Burlingame et al.

    The central role of chromatin in autoimmune response to histones and DNA in systemic lupus erythematosus

    Journal of Clinical Investigations

    (1994)
  • Cited by (0)

    1

    Present address. Department of Microbiology, Program in Molecular Biology, University of Colorado Health Sciences Center, 4200 E, 9th Avenue, Denver, Colorado 80262, USA.

    View full text