TLR9 recognizes unmethylated CpG-rich, pathogen-derived DNA sequences and represents the component of the innate immune system that heavily influences adaptive immunity and may contribute to the immunological disturbances in rheumatoid arthritis (RA). Accumulating data indicate that BM of RA patients participates in the pathogenesis of this disease as a site of proinflammatory cytokines overproduction and lymphocytes activation. Here, we investigated the functionality of TLR9 and its role in the modulation of RA BM B-cell functions. We report that BM B cells isolated from RA patients express TLR9 at the mRNA and protein levels acquired at the stage of preB/immature B-cell maturation. Stimulation of BM CD20 1 B cells by CpG-containing oligodeoxynucleotide-enhanced expression of activation markers (CD86 and CD54) triggered IL-6 and TNF-a secretion and cell proliferation. Significantly higher levels of eubacterial DNA encoding 16S-rRNA were found in BM samples from RA than osteoarthritis patients. Moreover, RA BM B cells exerted higher expression of CD86 than their osteoarthritis counterparts, suggesting their in situ activation via TLR9. Thus, our data indicate that TLR9 may participate in direct activation and proliferation of B cells in BM, and therefore could play a role in the pathogenesis of RA.Key words: B cells . Inflammation . Rheumatoid arthritis . TLR9 Supporting Information available online Introduction TLR9, a member of the family of pathogen recognition receptors, recognizes unmethylated CpG-rich, pathogen-derived DNA sequences [1]. Stimulation of TLR9 by these ligands trigger signaling pathway common for all TLR that results in NF-kB transcription factor activation and production of proinflammatory cytokines, including TNF-a, 3], whose overexpression plays an important role in the development and perpetuation of rheumatoid arthritis (RA) [4]. Furthermore, via activation of antigen presenting cells, TLR9 exerts potent effect on the development of acquired immunity [5]. It was shown that unmethylated DNA sequences containing CpG motifs enhance capacity of antigen presentation, induce cell proliferation, immunoglobulin class switch recombination and production of inflammatory mediators by human peripheral blood (PB) and mouse B lymphocytes [5][6][7][8][9]. Interestingly, TLR9-transduced signaling pathway contributes to the activation of autoreactive, rheumatoidfactor-producing B cells [10]. Moreover, experimental data suggest that TLR9 and its ligands participate in the pathogenesis of RA [11,12]. Intra-articularly injected bacterial DNA containing CpG motifs induces transient arthritis [11] and aggravates collagen-induced [14][15][16]. Furthermore, BM may serve as a secondary lymphoid organ, where an effective inflammatory response with strong antigen presentation, exceeding that observed in lymph nodes, can develop resulting in functional effector and memory T-cell generation [17,18]. Interestingly, the development of pathological processes in RA such as synovitis, joint cartilage and bone destructio...