Rheumatoid Arthritis: Alsaleh G

Alsaleh G, Suffert G, Semaan N, Juncker T, Frenzel L, Gottenberg JE, Sibilia J, Pfeffer S, Wachsmann D. · Laboratoire Physiopathologie des Arthrites, EA3432, Université Louis Pasteur de Strasbourg, Unité de Formation et de Recherche Sciences Pharmaceutiques, Illkirch, France. · J Immunol. · Pubmed #19342689 No free full text.

Abstract: MicroRNAs (miRNAs) have emerged as key players in the regulation of expression of target mRNAs expression. They have been associated with diverse biological processes, and recent studies have demonstrated that miRNAs play a role in inflammatory responses. We reported previously that LPS-activated fibroblast-like synoviocytes (FLS) isolated from rheumatoid arthritis (RA) patients express IL-18 mRNA but they do not release IL-18. Based on the observation that this inhibition was due to a rapid degradation of IL-18 mRNA, our group has conducted a study to identify miRNAs that could play a role in the "antiinflammatory" response of LPS-activated RA FLS. LPS challenge modulated the expression of 63 miRNAs as assessed by microarray analysis. Fifteen miRNAs were up-regulated, including miR-346, for which overexpression upon LPS treatment was validated by quantitative RT-PCR. We then transfected FLS with an antisense oligonucleotide targeting miR-346 and found that, in these conditions, IL-18 release could be measured upon LPS activation of FLS. Moreover, we also demonstrated that miR-346 indirectly regulated IL-18 release by indirectly inhibiting LPS-induced Bruton's tyrosine kinase expression in LPS-activated RA FLS. These findings suggest that miRNAs function as regulators that help to fine-tune the inflammatory response in RA.

Messer L, Alsaleh G, Freyssinet JM, Zobairi F, Leray I, Gottenberg JE, Sibilia J, Toti-Orfanoudakis F, Wachsmann D. · Laboratoire Physiopathologie des Arthrites, Université de Strasbourg, UFR Sciences Pharmaceutiques, Illkirch, France. · Arthritis Res Ther. · Pubmed #19291304 links to  free full text

Abstract: INTRODUCTION: In the present study, we investigated the ability of microparticles isolated from synovial fluids from patients with rheumatoid arthritis or osteoarthritis to induce the synthesis and release of key cytokines of B-lymphocyte modulation such as B cell-activating factor, thymic stroma lymphopoietin, and secretory leukocyte protease inhibitor by rheumatoid fibroblast-like synoviocytes. METHODS: Microparticles were analyzed in synovial fluids from patients with rheumatoid arthritis, osteoarthritis, microcristalline arthritis, and reactive arthritis. In addition, microparticle release after activation from various cell lines (CEM lymphocyte and THP-1 cells) was assessed. Microparticles were isolated by differential centrifugation, and quantitative determinations were carried out by prothrombinase assay after capture on immobilized annexin V. B cell-activating factor, thymic stroma lymphopoietin, and secretory leukocyte protease inhibitor release was evaluated by enzyme-linked immunosorbent assay. RESULTS: Microparticles isolated from synovial fluids obtained from rheumatoid arthritis and osteoarthritis patients or microparticles derived from activated THP-1 cells were able to induce B cell-activating factor, thymic stroma lymphopoietin, and secretory leukocyte protease inhibitor release by rheumatoid arthritis fibroblast-like synoviocytes. Conversely, CEM-lymphocytes-derived microparticles generated by treatment with a combination of PHA, PMA and Adt-D did not promote the release of B cell-activating factor but favored the secretion of thymic stroma lymphopoietin and secretory leukocyte protease inhibitor by rheumatoid arthritis fibrobast-like synoviocytes. However, microparticles isolated from actinomycin D-treated CEM lymphocytes were not able to induce B cell-activating factor, thymic stroma lymphopoietin, or secretory leukocyte protease inhibitor release, indicating that microparticles derived from apoptotic T cells do not function as effectors in B-cell activation. CONCLUSIONS: These results demonstrate that microparticles are signalling structures that may act as specific conveyors in the triggered induction and amplification of autoimmunity. This study also indicates that microparticles have differential effects in the crosstalk between B lymphocytes and target cells of autoimmunity regarding the parental cells from which they derive.

Semaan N, Alsaleh G, Gottenberg JE, Wachsmann D, Sibilia J. · Laboratoire Physiopathologie des Arthrites, Université Louis Pasteur de Strasbourg, Unité de Formation et de Recherche Sciences Pharmaceutiques, Illkirch, France. · J Immunol. · Pubmed #18292575 links to  free full text

Abstract: MyD88 and focal adhesion kinase (FAK) are key adaptors involved in signaling downstream of TLR2, TLR4, and integrin alpha5beta1, linking pathogen-associated molecule detection to the initiation of proinflammatory response. The MyD88 and integrin pathways are interlinked, but the mechanism of this cross-talk is not yet understood. In this study we addressed the involvement of Etk, which belongs to the Tec family of tyrosine kinases, in the cross-talk between the integrin/FAK and the MyD88 pathways in fibroblast-like synoviocytes (FLS) and in IL-6 synthesis. Using small interfering RNA blockade, we report that Etk plays a major role in LPS- and protein I/II (a model activator of FAK)-dependent IL-6 release by activated FLS. Etk is associated with MyD88, FAK, and Mal as shown by coimmunoprecipitation. Interestingly, knockdown of Mal appreciably inhibited IL-6 synthesis in response to LPS and protein I/II. Our results also indicate that LPS and protein I/II induced phosphorylation of Etk and Mal in rheumatoid arthritis FLS via a FAK-dependent pathway. In conclusion, our data provide support that, in FLS, Etk and Mal are implicated in the cross-talk between FAK and MyD88 and that their being brought into play is clearly dependent on FAK.

Alsaleh G, Messer L, Semaan N, Boulanger N, Gottenberg JE, Sibilia J, Wachsmann D. · EA 3432, Université Louis Pasteur de Strasbourg, Illkirch, France. · Arthritis Rheum. · Pubmed #17907168 links to  free full text

Abstract: OBJECTIVE: It was recently demonstrated that synoviocytes (FLS) from rheumatoid arthritis (RA) patients express BAFF transcripts that are up-regulated by tumor necrosis factor alpha (TNFalpha) and interferon-gamma (IFNgamma). Thus, BAFF increases in RA target cells might be related to activation of the receptors of innate immunity. The purpose of this study was to determine whether ligands of Toll-like receptor 2 (TLR-2), TLR-4, TLR-9, and alpha5beta1 integrin are able to induce BAFF synthesis by RA FLS. METHODS: Quantitative reverse transcription-polymerase chain reaction analyses and enzyme-linked immunosorbent assays were performed to evaluate BAFF messenger RNA induction and BAFF release from FLS after stimulation by ligands for TLR-2, TLR-4, TLR-9, alpha5beta1 integrin (bacterial lipopeptide [BLP] palmitoyl-3-cysteine-serine-lysine-4, lipopolysaccharide [LPS], CpG, and protein I/II, respectively), TNFalpha, and IFNgamma. RESULTS: In contrast to IFNgamma, neither TNFalpha, LPS, BLP, nor CpG induced the de novo synthesis and release of BAFF by FLS. Priming of cells with IFNgamma did not have a synergistic effect on BAFF synthesis by FLS stimulated with bacterial products known as pathogen-associated molecular patterns. Moreover, we found that IFNgamma-induced BAFF synthesis is inhibited by simultaneous stimulation with either TLR ligands or TNFalpha. We also showed that interplay between TLRs, TNF receptors, and IFNgamma signaling induces the expression of suppressor of cytokine signaling 1 (SOCS-1) and SOCS-3 and reduces IFNgamma-dependent STAT-1 phosphorylation, which might explain this inhibition. In contrast, we demonstrated that stimulation of alpha5beta1 integrin can induce BAFF synthesis and release per se and that stimulation of this pathway has no inhibitory effect on IFNgamma-induced BAFF synthesis. CONCLUSION: Our findings indicate that BAFF secretion by resident cells in target organs of autoimmunity is tightly regulated by innate immunity, with positive and negative controls, depending on the receptors and the pathways triggered.