The hallmarks of rheumatoid arthritis (RA) are leukocytic infiltration of the synovium and expansiveness of fibroblast-like synoviocytes (FLS). The abnormal proliferation of FLS and their resistance to apoptosis is mediated, at least in part, by present in RA joints proinflammatory cytokines and growth factors. Because IL-15 exerts properties of antiapoptotic and growth factors, and is produced by RA FLS, we hypothesized that IL-15 participates in RA FLS activation. To test this hypothesis, we first examined whether RA FLS express chains required for high affinity functional IL-15R. Indeed, RA FLS express IL-15Rα at mRNA and protein levels. Moreover, we confirmed the presence of IL-2Rβ and common γ-chains. Interestingly, TNF-α or IL-1β triggered significant elevation of IL-15Rα chain at mRNA and protein levels. Next, we investigated the effects of exogenous or endogenous IL-15 on Bcl-2 and Bcl-xL expression, FLS proliferation, and apoptosis. Exogenous IL-15 enhanced RA FLS proliferation and increased the level of mRNA-encoding Bcl-xL. To test the role of endogenous IL-15 in the activation of RA FLS, an IL-15 mutant/Fcγ2a protein exerting properties of specific antagonist to the IL-15Rα chain was used. We found that blocking IL-15 biological activities using this protein substantially reduced endogenous expression of Bcl-2 and Bcl-xL, and RA FLS proliferation that was reflected by increased apoptosis. Thus, we have demonstrated that a distinctive phenotype of RA FLS, i.e., persistent activation, proliferation, and resistance to apoptosis, is related to the autocrine activation of IL-15Rs by FLS-derived IL-15.

IL-15, a key cytokine linking innate and acquired immunity, is expressed in many cell types and tissues. Recent data indicate constitutive expression of IL-15 in human neural cell lines and tissues. The aim of the present study was to examine the expression patterns of mRNA encoding IL-15 and IL-15 receptor alpha (IL-15Ralpha) isoforms in select structures of human fetal brain. We report that mRNA for IL-15 and IL-15Ralpha isoforms were expressed in all tested brain structures: cerebral cortex, cerebellum, hippocampus, and thalamus. However, the levels of IL-15 and IL-15Ralpha mRNA were higher in the hippocampus and cerebellum in comparison with cortex and thalamus. Moreover, higher levels of cytosol in comparison with membrane-bound IL-15 isoform were present in all brain structures. The constitutive, but distinct, expression of IL-15 and its receptors in select human fetal brain structures suggests that IL-15 plays a role in their development and physiology.

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...

Objective. To investigate the effects of taurine chloramine (Tau-Cl), a chlorinated derivative of the amino acid taurine, on the expression of cyclooxygenase (COX) isoenzymes and prostaglandin E 2 (PGE 2 ) synthesis in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS).Methods. FLS, isolated from the synovial tissue of RA patients, were treated in vitro with either interleukin-1␤ (IL-1␤; 1 ng/ml) alone or together with 200-500 M Tau-Cl. The expression of COX isoenzymes was evaluated at both the protein (Western blotting) and the messenger RNA (mRNA) (reverse transcriptase-polymerase chain reaction) levels. The concentration of PGE 2 was measured by competitive acetylcholinesterase enzyme immunoassay.Results. Resting FLS expressed mRNA encoding both COX-1 and COX-2, but only COX-1 was present at the protein level. These cells produced negligible amounts of PGE 2 . Upon stimulation with IL-1␤, elevation of COX-2, but not COX-1, mRNA and protein preceded the enhancement of PGE 2 synthesis. In the presence of 300-400 M Tau-Cl, significant inhibition of IL-1␤-triggered COX-2 mRNA and protein, and a related decrease in PGE 2 production, was observed. In contrast, no significant changes in COX-1 mRNA and protein levels were noted.Conclusion. Tau-Cl inhibits IL-1␤-triggered elevation of COX-2 and generation of PGE 2 by RA FLS. These results expand the spectrum of known antiinflammatory activities of this compound.

Fibroblast-like synoviocytes (FLS) participate in rheumatoid arthritis (RA) chronic synovitis by producing pro-inflammatory cytokines (IL-6, IL-8), growth factors (VEGF) and other inflammatory mediators (PGE2, NO). We have previously reported that Tau-Cl, generated by neutrophils, inhibits in vitro some of these pathogenic RA FLS functions. Taurine bromamine (Tau-Br) originates from eosinophils and neutrophils, and its immunoregulatory activities are poorly known. Therefore, we investigated the effects of Tau-Br on RA FLS functions and compared it to Tau-Cl anti-inflammatory action. When applied at noncytotoxic concentrations: (i) Tau-Br inhibited IL-6 and PGE2 production with potency similar to Tau-Cl (IC50 approximately 250 microM), (ii) Tau-Br failed to affect VEGF and IL-8 synthesis, while Tau-Cl exerted inhibitory effect (IC50 approximately 400 microM), (iii) none of these compounds affected NO generation and iNOS expression. Thus, Tau-Cl is more effective than Tau-Br in normalization of pro-inflammatory RA FLS functions.