Session Type: Abstract Submissions (ACR)
Background/Purpose: Altered B cell signaling has been proposed to play an important role in susceptibility to SLE. In mice, genetic manipulations or polymorphisms that attenuate BCR signaling have been shown to lead to altered selection or survival of self-reactive lymphocytes whereas those that lead to enhanced BCR signaling can breach B cell anergy, with both types of defects reported in various murine models of SLE. Most previous reports indicate that SLE B cells are hyper-responsive to BCR crosslinking. However, these findings contrast with preliminary evidence suggesting that some of the genetic risk variants associated with SLE may lead to attenuated BCR signaling. In this study we have examined the association between these risk variants and the altered B cell function observed in SLE.
Methods: Patients (N=44) satisfying at least 4 ACR criteria and (N=30) healthy age-matched controls without a family history of systemic autoimmune disease were recruited. PBMC were isolated over a Ficoll gradient, rested for 1 hr at 37oC, and then stimulated for 2’ with media alone, or 2’ and 10’ with media containing 20 μg/ml of F(ab’)2 goat anti-human IgM. Cells were stained with anti-CD19, -CD27, -IgD, -IgM, and -CD38, to permit gating on naïve mature B cells (CD19+CD27–IgD+CD38+) stratified based on their IgM cell surface expression and transitional B cells (CD19+CD27–IgD+CD38++IgMhi). B cell signaling was examined by Phosflow using anti-pSyk (pY348), –pPLCγ2 (pY759) or -pERK1/2 (pT202/Y204) Ab following fixation and permeabilization. SNP genotyping of subject DNA was performed using TaqMan assays specific for rs2618476 (BLK), rs10516487 (BANK-1), rs7829816 (LYN), and rs2476601 (PTPN22).
Results: There was an increased proportion of naïve B cells in SLE patients that had elevated basal levels of pSYK and pERK, suggesting prior activation in-vivo. Following IgM crosslinking, the naïve B cells of SLE patients demonstrated significantly increased levels of p-SYK and trends to increased levels of p-PLCγ2 and p-ERK above basal levels as compared to controls. This increased signaling was most marked in the IgMhi mature naïve and transitional B cell compartments of SLE patients. There was no correlation between the basal levels of phosphorylated signaling molecules and any of the SNPs examined. No association was seen between BLK or LYN SNPs and any of the phosphorylated signaling molecules following IgM crosslinking. Consistent with attenuated BCR function there was a trend to decreased pPLCγ2 and significantly decreased pERK with the lupus-associated PTPN22 variant at 10’ following IgM crosslinking. The lupus associated BANK-1 variant was associated with decreased pPLCγ2 at 2’. To further explore the role of non-genetic factors in the B cell hyper-responsive phenotype, serial measurements of pSYK were performed. Significant variations between individual visits were seen in a subset of patients.
Conclusion: No correlation was observed between increased B cell signaling and several of the genetic risk variants predicted to alter B cell receptor function in SLE. It is likely that this phenotype arises from other genetic or non-genetic factors.
N. H. Chang,
P. R. Fortin,
D. D. Gladman,
M. B. Urowitz,
J. E. Wither,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/altered-response-to-b-cell-receptor-bcr-crosslinking-in-sle-correlation-with-genetic-risk-variants-predicted-to-impact-bcr-signaling/