Recognition of new citrulline‐containing peptide epitopes by autoantibodies produced in vivo and in vitro by B cells of rheumatoid arthritis patients

Abstract

SummaryAnti-citrullinated peptide/protein antibodies (ACPAs) are highly sensitive and specific markers of rheumatoid arthritis (RA). Identification of peptide epitopes that may detect different subgroups of RA patients might have diagnostic and prognostic significance. We have investigated citrulline-and arginine-containing peptide pairs derived from filaggrin, collagen or vimentin, and compared this citrulline-peptide panel with the serological assays conventionally used to detect ACPAs. Furthermore, we studied if the same citrulline-peptides identify antibody-secreting cells in in vitro cultures of RA B cells. Recognition of citrulline-and arginine-containing filaggrin, vimentin and collagen peptide epitopes were tested by Multipin ELISA system, by indirect ELISA and by a peptide-specific microarray. B cells were purified from blood by negative selection; antibody-producing cells were enumerated by ELISPOT assay. The panel composed of citrulline-peptide epitopes of filaggrin, collagen and vimentin was recognized by RA sera with a sensitivity and specificity comparable with the currently used tests. Moreover, the combined citrulline-peptide panel including the new short epitope peptide of filaggrin, fil311-315, also identified nearly one-third of RA cases that were negative for antibodies against cyclic citrullinated peptides, mutated citrullinated vimentin or for rheumatoid factor. The results with the peptide-specific microarray have shown that although most ACPAs recognizing the four citrulline peptides are IgG, some of them specifically recognizing citrulline-containing filaggrin peptides (fil311-315 and fil306-326) are IgM, and so may be produced either by newly formed activated B cells or by unswitched B memory cells. Furthermore, the citrulline-peptides of filaggrin and vimentin detect ACPA-producing cells, and so could also be applied to study the B cells of RA patients.

Biokonjugátumok – összefogni érdemes

“…20,21 Ezen epitóprégión belül -az egészséges és RA betegségben szenvedõ személyek szérummintái-nak összehasonlításával -azonosítottuk azt a legkisebb peptidszakaszt ( 311 TXGRS 315 ), amely ellenanyag (B-sejt) epitópként viselkedik és alkalmas lehet a betegség korai és specifikus kimutatására. 123. évfolyam, 3. szám, 2017.…”

Citrullination: A Specific Target for the Autoimmune Response in Rheumatoid Arthritis

“…ACPAs can react with a variety of citrullinated proteins, such as vimentin, fibrinogen, a enolase, fillagrin, and type II collagen (3,4). Most clinical assays currently in use employ cyclic citrullinated peptides as the target Ag, which may lead to a small percentage of false-negative results (5,6).…”

Suppression of innate and adaptive B cell activation pathways by antibody coengagement of FcγRIIb and CD19

“…[26][27][28] We and others have previously shown that in vitro-activated B cells from ACPA-positive RA patients produce citrullinated peptide-specific IgG. 36,37 To see if XmAb5871 has an effect on citrulline peptide-specific antibody production, purified B cells from selected RA patients were cultured in the presence of IL-2 and R848 to stimulate memory B cells. The number of antibody-producing cells was estimated on citrulline-containing filaggrin-peptide coated plates.…”

“…37 Briefly, purified B cells from RA patient blood were isolated using MACS. 10 6 cells/ml were cultured in RPMI 1640 containing 10% FCS in the presence of 10 ng/ml rhIL-2 and 1 μg/ml R848 (Sigma-Aldrich, SML0196) polyclonal activator with or without XmAb5871.…”

“…54 Cells were harvested on the third day, washed and counted, and then were transferred into the wells of ELISPOT plates precoated with neutravidin and the biotinylated citrulline-containing 19mer filaggrin peptide (SHQESTXGXSXGRSGRSGS, X stands for citrulline). 37 The spots were developed after 20 h.…”