Bub1 negatively regulates osteoclast activation through NF-κB suppression in inflammatory arthritis

Rheumatoid arthritis (RA) is an inflammatory autoimmune disease characterized by synovitis and, bone and cartilage destruction. Although disease-modifying anti-rheumatic drugs (DMARDs) have dramatically improved clinical outcomes, these therapies are not universally effective in all patients due to the heterogeneity of RA pathogenesis. Therefore, there is still a need to elucidate molecular mechanisms underlying RA pathogenesis to identify novel therapeutic targets. In this study, we identified Budding uninhibited by benzimidazoles 1 (BUB1) was highly expressed in RA patients’ synovium and murine ankle tissue with arthritis. Since we revealed CD45+CD11b+ myeloid cells as Bub1 highly expressing population among synovial cells, myeloid cell-specific Bub1 conditional knockout (cKO) mice were generated. Obtained cKO mice showed no significant phenotypes in healthy conditions while they exhibited reduced bone mineral density under K/BxN serum-transfer arthritis. Moreover, histomorphometric analysis suggested that the reduced bone mass was caused by increased osteoclast differentiation and activation. RNA-seq and subsequent Gene set enrichment analysis (GSEA) demonstrated significantly enriched NF-κB pathway among up-regulated genes in RANKL-stimulated bone marrow macrophages obtained from Bub1 cKO, suggesting Bub1 KO macrophages are sensitive to inflammatory conditions. Indeed, osteoclastogenesis in Bub1 KO macrophages was enhanced by RANKL and TNFα treatment and localization of NF-κB component p65 was observed. Finally, osteoclastogenesis was increased by Bub1 inhibitor BAY1816032 treatment. These data suggested that BUB1 in myeloid cells plays a protective role against bone loss under inflammatory arthritis. To investigate the functions of Bub1 in myeloid cells, bone marrow-derived macrophages (BMMs) were isolated from Ctrl or myeloid specific Bub1cKO mice. We then induced osteoclast differentiation and performed gene expression profiling analysis using RNA-seq data of BMMs.