Background/Purpose: Human regulatory B cells still need to be characterized. Given the absence of phenotypical definition, a functional definition based on their ability to secrete IL-10 is often used (corresponding to B10⁺ cells). Chemokine receptors (CKR) profiles are useful to characterize some populations of T cells but have never been explored among B10⁺ cells. Moreover, very little is known about B10⁺ cell migration. Chemokines (CK) have also been implicated in the differentiation of naïve T cells towards regulatory T cells. Therefore, the aims of our study were to characterize the profile of CKR on B10⁺ cells and also to investigate CK implicated in their migration and differentiation both in control (CTL) and in rheumatoid arthritis (RA) patients.

Methods: B cells were isolated with Rosette Sep Human B cells enrichment followed by Ficoll separation. B cells were activated 24 hours with CpG and CD40L to generate B10⁺ cells. For RNA sequencing and migration assay, B10⁺ and IL-10^neg B cells (B10^neg) cells were then sorted using IL-10 secretion assay. IL-10 secretion from B cells was assessed by FACS and ELISA. For migration assay, ability to migrate in response to CCL21, CCL22, CXCL11, CXCL12 or CXCL13 or synovial fluid (SF) from RA or osteoarthritis (OA), were evaluated by migration assay in 5μM Transwell chambers and expressed as fold increase compare to basal migration towards control media.

Results: Transcriptomic analysis was done on B10⁺ and B10^neg from 5 CTL and 4 RA patients. 335 genes were found differentially regulated (DEG) in control B10⁺ cells compared to B10^neg in CTL and 188 in RA patients. Of note, only 159 DEG were shared by both groups, while 29 were specific to RA. Gene ontology classification of DEG revealed that genes from CKs pathways were highly enriched in B10⁺ from CTL and to a lesser extend in B10⁺ from RA patients. Among all CK tested, only CXCL13, attracted significantly more B10⁺ than B10^neg from CTL (12.4 ± 10.0-fold increased migration of B10+ vs 6.0 ± 3.4-fold increased migration of B10neg, n=21). This was also true in RA patients (18.7 ± 22.8-fold increased migration for B10⁺ vs 6.4 ± 5.3-fold increased migration for B10^neg, n=13). SF from RA induced a significant migration of B10⁺ cells in CTL (9.6 ± 12.0-fold increased migration, p< 0.0001, n=22) and RA patients (7.1 ± 9.7-fold increased migration, p=0.0002, n=13), higher than those induced by SF from OA. CXCL13 was significantly higher in SF from RA than in SF from OA (685.5 ± 135.1 pg/mL vs 122.8 ± 186.7 pg/mL, p=0.03). Importantly, B10⁺ migration towards SF was significantly correlated with the levels of CXCL13 in these SF in CTL (r=0.62, p=0.007, n=17) and with a trend in RA patients (r=0.45, p=0.07, n=17). Lastly, CXCL13 was also found to increase IL-10 secretion in B cells stimulated with CpG in CTL (1.4 ± 0.3-fold increase, p< 0.0001, n=20) and in RA patients (1.2 ± 0.2-fold increase, p=0.01, n=13).

Conclusion: We showed that CXCL13 is a key driver for migration and differentiation of B10⁺ cells in CTL and in RA patients.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/cxcl13-is-a-key-driver-for-migration-and-differentiation-of-regulatory-b-cells/