Comparison of pathogenic and non-pathogenic murine antibodies to DNA: antigen binding and structural characteristics

Abstract

Three pathogenic and two non-pathogenic NZB/NZW F1 mAbs to DNA were compared. Pathogenicity was defined as the ability to induce nephritis in BALB/c mice. All mAbs were IgG2a or 2b, had high avidity for double-stranded DNA and fixed complement well. All three pathogens expressed idiotype IdGN2. Mice receiving pathogenic mAbs (compared with non-pathogenic) had more glomerular IgG deposits. The unique properties of two of the pathogens were: strong homogeneous staining of Hep-2 nuclei and the ability to bind (i) nucleosomes, (ii) histone (after mAb complexed with DNA), (iii) heparan sulfate in renal basement membranes (after complexing with DNA/histone) and (iv) nuclei in vivo. Comparison of nucleotide and amino acid sequences of the V regions of heavy and light Ig chains showed use of multiple VHDJH and V kappa J kappa gene families, with representation of several anti-DNA 'families' described by others. Arginine (R) occurred in the CDR2 or CDR3 of VH chains in all pathogens; R was absent in the CDRs of VH chains of non-pathogens. Positively and negatively charged AA were more frequent in VH CDR of pathogens than of non-pathogens. We hypothesize that the tertiary structure of mAbs determined by VH CDR regions permits stronger binding to negatively charged antigens (DNA and heparan sulfate) and to positively charged molecules (histone) in pathogens compared with non-pathogens.

Induction of Autoantibody Production Is Limited in Nonautoimmune Mice

“…Injection of a control, nonautoreactive mAb, MOPC-21, did not induce IgG anti-DNA Ab or proteinuria (Ref. 3 and data not shown). This suggests that potentially autoreactive B cells in nonautoimmune mice can be activated to secrete pathogenic IgG anti-DNA Ab by appropriate Th cells.…”

“…Plates were washed again with Tris/Tween 20, and incubated with a mixture of 5-bromo-4-chloro-3-indolyl phosphate with agarose gel at a final concentration of 0.6%. AntidsDNA Ab-forming cells (AFC) 3 were enumerated as blue spots using an inverted microscope (Leitz Canada, Midland, Ontario, Canada) by two of us (F.M.E. and R.R.S.)…”

“…Briefly, spleen cells were cultured with or without peptides for 5 days; supernatants were tested for IgG anti-dsDNA Ab, as described previously (3,4). Briefly, 96-well plates (Costar) were coated with dsDNA (100 g/ml; Sigma-Aldrich) overnight at 4°C.…”

“…Four B cell hybridomas, including a dsDNA-specific (A6.1) and three non-DNA binding (1GE6, 375-57, and 375-100), were established from BWF 1 mice by fusion of their spleen cells with a nonsecreting murine myeloma cell, as described previously (3,18).…”

Anti‐DNA Ig peptides promote Treg cell activity in systemic lupus erythematosus patients

“…In humans, similar amino acid sequences (matching 12-14 of the 15 amino acids) occur in anti-DNA and antiphospholipid antibodies, particularly those derived from the V3-23 V H family (45,46). In addition, a recent search of the National Institutes of Health BLAST protein database (www.ncbi.nlm.nih.gov/blast) indicated identity in 8-9 amino acids of the 12-mer wild peptide p34 from the anti-DNA antibody 6.1 (47) in murine antibodies to phosphorylcholine and galactan, and human antibodies to flavin, and 2 human myelomas. Thus, cross-reactivity secondary to sequence homology could explain those findings.…”