Background/Purpose:  Rheumatoid arthritis (RA) is frequently associated with bone loss due to imbalanced bone resorption and formation. B cell depletion therapy (BCDT) attenuates bone erosion in a subset of RA patients. B cells promote osteoclast formation, but their role in osteoblasts (OB) in RA has not been well studied. Here, we used B cells from TNF-transgenic (TNF-Tg) mice and mice with Collagen-Induced Arthritis (CIA), mouse models of RA, and from RA patients to investigate if RA B cells inhibit OB differentiation and the mechanisms involved.

Methods:  B cells were isolated using CD19 magnetic beads from bone marrow (BM) and subchondral BM (SB) of TNF-Tg and CIA mice, and from peripheral blood of RA patients with active disease, but not on biologics. Immunofluorescent (IF) staining, RNA sequencing, co-cultures of mesenchymal precursor cells (MPCs) and B cells, and BCDT were performed.

Results:  To study the distribution of B cells and their relationship with OBs, we performed IF on frozen bone sections from TNF-Tg and CIA mice with anti-B220 Ab for B cells and anti-osteocalcin Ab for OBs. B cells were enriched in the SB and endosteal BM areas of RA mice and were adjacent to OBs. To investigate if RA B cells produce OB inhibitors, we performed RNA sequencing of BM and SB B cells from TNF-Tg mice. Compared to WT BM B cells, TNF-Tg BM B cells expressed about 2-fold higher levels of OB inhibitors, including CCL3, TNF, and Dkk3. SB B cells expressed much higher levels of these inhibitors compared to their BM counterparts, and this was confirmed by qPCR (fold increase in TNF-Tg SB/TNF-Tg BM: CCL3 x12, TNF x4, Dkk3 x12; in CIA SB/CIA BM: CCL3 x20, TNF x10, Dkk3 x38). To determine if RA B cells directly affect OBs, we co-cultured B cells with WT MPCs in OB-inducing medium. RA B cells significantly inhibited OB differentiation (ALP+ area: 7±1 in TNF-Tg vs. 16±2 mm² in WT; 7±2 in CIA vs. 20±2mm² in non-CIA). To investigate potential signal pathways responsible for OB inhibition, expression of NF-kB, ERK and b-catenin in MPCs after co-culture with B cells was examined. RA B cells increased expression of NF-kB and pERK in MPCs.The effect of RA B cells on OB inhibition and NF-kB/ERK activation in MPCs was partially blocked by a CCL3 or TNF neutralizing Ab. To examine the overall effect of BCDT on bone mass and OB function, we treated TNF-Tg mice with murine anti-CD20. BCDT increased SB bone volume (BV/TV: 40±2 vs. 31±1% in IgG) and OBs (#/mm bone surface: 26±5 vs. 16±3 in IgG). BM cells from BCDT-treated mice had increased OB differentiation (CFU-ALP#: 53±9 vs. 21±5 in IgG). To determine if B cells from RA patients have a similar OB inhibitory effect, we co-cultured peripheral blood B cells from RA patients with MPCs. RA B cells inhibited OB differentiation and this was partially blocked by CCL3 or TNF neutralization. Furthermore, IF of synovial specimens from RA patients showed expansion of CCL3 and TNF-producing B cells.

Conclusion: B cells are enriched in the SB area of RA joints and are adjacent to OBs. RA B cells produce multiple OB inhibitors, including CCL3 and TNF, which inhibit OB differentiation. BCDT reduces bone loss and stimulates OB function. Thus, B cells may directly target OBs, and thus contribute to bone loss and joint erosion in RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/b-cells-inhibit-osteoblast-differentiation-in-inflammatory-arthritis/