Accumulation of Self-Reactive Naïve and Memory B Cell Reveals Sequential Defects in B Cell Tolerance Checkpoints in Sjögren’s Syndrome

Abstract

Sjögren’s syndrome (SS) is an autoimmune disease characterised by breach of self-tolerance towards nuclear antigens resulting in high affinity circulating autoantibodies. Although peripheral B cell disturbances have been described in SS, with predominance of naïve and reduction of memory B cells, the stage at which errors in B cell tolerance checkpoints accumulate in SS is unknown. Here we determined the frequency of self- and poly-reactive B cells in the circulating naïve and memory compartment of SS patients. Single CD27−IgD+ naïve, CD27+IgD+ memory unswitched and CD27+IgD− memory switched B cells were sorted by FACS from the peripheral blood of 7 SS patients. To detect the frequency of polyreactive and autoreactive clones, paired Ig VH and VL genes were amplified, cloned and expressed as recombinant monoclonal antibodies (rmAbs) displaying identical specificity of the original B cells. IgVH and VL gene usage and immunoreactivity of SS rmAbs were compared with those obtained from healthy donors (HD). From a total of 353 VH and 293 VL individual sequences, we obtained 114 rmAbs from circulating naïve (n = 66) and memory (n = 48) B cells of SS patients. Analysis of the Ig V gene repertoire did not show significant differences in SS vs. HD B cells. In SS patients, circulating naïve B cells (with germline VH and VL genes) displayed a significant accumulation of clones autoreactive against Hep-2 cells compared to HD (43.1% vs. 25%). Moreover, we demonstrated a progressive increase in the frequency of circulating anti-nuclear naïve (9.3%), memory unswitched (22.2%) and memory switched (27.3%) B cells in SS patients. Overall, these data provide novel evidence supporting the existence of both early and late defects in B cell tolerance checkpoints in patients with SS resulting in the accumulation of autoreactive naïve and memory B cells.

A comprehensive investigation on the distribution of circulating follicular T helper cells and B cell subsets in primary Sjögren’s syndrome and systemic lupus erythematosus

“…In our study, we found a significant enrichment of CD19 1 IgD 1 CD27 2 naive B cells in the peripheral blood of both pSS and SLE patients compared to healthy individuals. This observation is consistent with previous reports [12][13][14] and indicates that early B cell tolerance checkpoints are impaired significantly in these autoimmune diseases; moreover, the break of tolerogenic mechanism at this stage probably accelerates the mobilization of autoreactive naive B cells from the bone marrow to the periphery [15,16].…”

“…This observation is consistent with previous reports [12][13][14] and indicates that early B cell tolerance checkpoints are impaired significantly in these autoimmune diseases; moreover, the break of tolerogenic mechanism at this stage probably accelerates the mobilization of autoreactive naive B cells from the bone marrow to the periphery [15,16]. There is another major tolerance checkpoint during the maturation stage of immature B cells when transitional B cells overcome a negative selection.…”

Single cell cloning and recombinant monoclonal antibodies generation from RA synovial B cells reveal frequent targeting of citrullinated histones of NETs

“…Single-cell real time-PCR reactions and IgV gene amplification were performed as described in refs. 21 and 22 . Briefly, cDNA from CD3-CD19+B cells was amplified using reverse primers that bind the Cμ/Cγ or Cα constant region in three independent nested-PCR.…”

“…Immunoglobulin Analysis Tool (IgAT) software was used to calculate the probability of antigen-driven selection within the Ig repertoire of the RA-rmAbs, 23 as previously described. 22 …”

Brief Report: Defective Early B Cell Tolerance Checkpoints in Sjögren's Syndrome Patients

“…Rheumatoid factors are also often detected in these patients and are associated with greater disease activity. A recent study suggests defective autoreactive B cell counterselection in SS patients, as illustrated by the elevated frequency of polyreactive clones in total CD3–CD19+CD27–IgD+ B cells, which contain CD21 –/low clones previously reported to express autoreactive antibodies . However, the functionality of the central and peripheral B cell tolerance checkpoints, which are responsible for the elimination of developing autoreactive clones in the bone marrow and the periphery, respectively, remains to be analyzed in SS patients.…”