Quantitative analysis of immunoglobulin Vκ CDR3 lengths in rheumatoid arthritis: Evidence of antigenic selection or dysregulation of gene rearrangement

Academic Article

Abstract

  • The CDR3 domain of a κ light chain is generated by Vκ-Jκ joining and lies at the center of the antigen binding site; its sequence can thus influence antigenic specificity of the antibody. Despite the presence of N region addition, CDR3s > 10 amino acids (aa) long are expressed in < 5% of κ chains in B cells from normal adults. We sought to extend our sequence analysis of 108 Vκ clones amplified from 3 RA synovia and 2 matched PBLs, 3 control PBLs, and 1 control spleen, which demonstrated enrichment for 11 aa CDR3s in κ light chains of 2 of 3 RA patients (pts). Using leader/FR1 and Cκ primers, we PCR-amplified VκIII transcripts from first strand cDNA of 6 RA synovia, PBLs of 3 of these RA pts, PBLs of 7 other RA pts and PBLs of 3 normals. A VκIII FR3 primer and a 32P-labeled internal Cκ primer were used to perform nested PCR, followed by polyacrylamide gel electrophoresis (PAGE) and autoradiography. Intensities of bands representing CDR3s of 8, 9, 10, and 11 aa codons were quantitated by phosphoimaging. Proportions of 11 aa CDR3s were > 10% in 2/6 synovia and 8/10 RA PBL, but in none of 3 controls. PBLs of an RA pt found by sequence analysis and PCR-PAGE to have enrichment for κ CDR3s of 11 aa were FACS sorted into IgM+/IgD+ (naive) and IgM-/IgD- (class-switched) B cells. PCR-PAGE analysis of these two B cell subsets was performed. Similar proportions of 11 aa CDR3s were found in the naive and class-switched populations, suggesting abnormal κ gene rearrangement, rather than antigenic selection, as the underlying mechanism. We conclude that Vκ repertoires of some RA patients are enriched for unusually long CDR3s. Preliminary data from one RA pt suggest this enrichment results from dysregulation of κ gene rearrangement during B cell ontogeny. Alternatively, there may be antigenic selection of B cells bearing unusually long κ light chain CDR3s in some RA pts, either in germinal centers (GCs) of normal lymphoid organs or in GCs in inflamed synovial tissue.
  • Published In

    Author List

  • Bridges SL; Lavelle JC; Byer S; Koopman WJ; Schroeder HW
  • Volume

  • 44
  • Issue

  • 1