Monoclonal anti-double stranded DNA antibody is a leucocyte-binding protein to up-regulate interleukin-8 gene expression and elicit apoptosis of normal human polymorphonuclear neutrophils

Objectives. To determine whether anti-double stranded DNA (anti-dsDNA) auto antibody could bind and affect the functions of normal human polymorphonucl ear neutrophils (PMN). Methods. Normal human PMN were incubated with different concentrations of a monoclonal mouse anti-dsDNA antibody (12B3) or mouse isotype-matched IgG2a . The binding of anti-dsDNA and PMN was measured by flow cytometry and inte rleukin-8 (IL-8) gene expression in PMN was detected by enzyme-linked immun osorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). PMN apoptosis was justified by morphological changes. The cognate antigen(s) of anti-dsDNA on the PMN surface was identified by membrane bio tinylation, immunoprecipitation and Western blot. Results. The binding of PMN with anti-dsDNA was much higher than with non-s pecific mouse IgG2a (70.8 vs 2.0%). Anti-dsDNA at concentrations higher tha n 12.5 ng/ml significantly enhanced the production and mRNA expression of I L-8 by PMN. However, anti-dsDNA facilitated PMN apoptosis after 3 h incubat ion. Western blot analysis of biotinylated PMN cell lysates demonstrated th at a 50-52 kDa membrane molecule is the cognate antigen of anti-dsDNA. Conclusions. Anti-dsDNA autoantibody up-regulates IL-8 gene expression and elicits activation-induced cell death (AICD) of human PMN via binding to a 50-52 kDa membrane-expressed molecule.