DICER and miRNA in rheumatic diseases

Our group has characterized LPS-induced miRNA expression in Fibroblast-like Synoviocytes (FLS) by microarray analysis. Having previously described the role of miR-19 on TLR2 expression, we then investigated the functions of miR-20 which belongs to the same cluster (miR-17-92). We reported (Philippe et al, ARD, 2013) that miR-20 targets the kinase Ask1, which regulates inflammatory responses (IL-6 expression) in these cells. This observation, combined to our description of miR-19 led us to hypothesize a coordinated action of the miRs encoded in the 17-92 cluster on TLR signaling (Philippe et al, Front. Immunol, 2013). Next, our expertise in miRNA-dependent regulation of gene expression (Alsaleh, Methods Mol Biol, 2014), prompted us to study epigenetic regulation of Baff (Alsaleh et al, PLoS One 2014), a cytokine of major importance in autoimmunity and the focus of intense interest in our lab (Francois et al, ART, 2013; Roescher, Mol Ther, 2014; Francois et al, J Autoimmun, 2015). We also analyzed the dialog between resident cells (salivary gland epithelial cells) and follicular helper T cells in primary Sjögren’s syndrome (Gong et al, J Autoimmun, 2014). Our interest in miRNA-dependant control of gene expression led us to collaborate with the group of S. Bluml (Medical University, Vienna, Austria) to analyze the role of miR-146 in RA (Saferding at al, J Autoimmun. 2017). Finally, we collaborate with S. Pfeffer (IBMC CNRS, Strasbourg) to gain more insights into the role of miR-346, discovered by our group (Alsaleh, J Immunol 2009; Seeman, PLoS One 2011), which potentially carries very potent anti-inflammatory properties. We are presently searching its target mRNAs in FLS by PAR-CLIP (Photoactivatable-Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation) and we expect that some of these will be further investigated in the next 5 years.

More recently, the work of our group further concentrated on the analysis of the murine hypomorphic Dicerd/d mutation and the consequences of low Dicer expression on inflammatory response. We showed that Dicer-deficient mice do not spontaneously develop rheumatoid arthritis symptoms, but exhibit increased responses when triggered by the injection of an arthritogenic serum (Alsaleh et al, A&R, 2016). This observation suggests that Dicer mutant mice could serve as a powerful animal model to investigate gene/environment interactions in the initiation of (auto)inflammatory diseases. Furthermore, the moderate impact of low Dicer expression on miRNA biogenesis observed in these mice indicates that reduced miRNA might not be the exclusive reason for the increased severity of the joint symptoms seen in Dicer d/d mice. Indeed, Dicer has also been recently involved in the metabolism of RNA produced upon Alu sequence transcription and we are currently collaborating actively with the group Prof J. Ambati (Kentucky Univ, USA) to explore the relationships between low Dicer expression in our mutant mice and augmented Alu-dependant inflammatory responses (Gelfand et al, Cell Rep, 2015). This recent development in our group will certainly be further investigated in the next 5 years (see below). An important axis being investigated in Dicerd/d mutants concerns the relationships between inflammation and ageing (inflammaging). Our Dicerd/d mutants exhibit striking features of premature ageing, which we are precisely describing now.

Plasmacytoid dendritic cells (pDCs) in rheumatic diseases.

Another axis of research regarding the control of inflammation in rheumatoid arthritis concerns the involvement of plasmacytoid dendritic cells (pDCs). Here we take advantage of a mutant mouse carrying a hypomorphic allele of the Ikaros gene. Interestingly, these mice are deficient in pDCs in the periphery, while the proportion of other immune cells remains unaffected. We performed a phenotypic analysis of these mice regarding the effect of the mutation on joint inflammation because the role of pDCs in RA is only poorly understood and the current literature on the subject shows incompatible observations. Briefly, we have now evidence that the lack of pDCs is associated with worsening of joint inflammation in two experimental mouse models: the serum transfer and the collagen-induced arthritis. Importantly, severe bone erosion is observed in Ikaros mutants following arthritis induction. Next, we mobilized pDCs in the joints to evaluate the potential anti-inflammatory properties of these cells. For that, we realized topical application of Aldara, an FDA-approved treatment of genital warts and small superficial basal skin carcinoma containing 5% imiquimod, a well-known TLR7 agonist. Interestingly, few (2 to 4) applications of Aldara on inflamed joints led to a major reduction of bone destruction (Nehmar et al; A&R, 2017). We reviewed the roles of Interferons and pDCs recently (Nehmar, Mariotte et al, Trends Mol. Med, 2018) and consider that the analysis of cellular interactions (FLS / pDCs / osteoclasts) important in joint homeostasis will be an important part of our projects for the next 5 years. In addition, other inflammatory joint diseases (such as microcrystalline arthritis) are also investigated in our lab, with a specific focus on the role of inflammasomes. For this project, we collaborate with R. Ricci (IGBMC, Strasbourg, see Zhang et al, J. Exp. Med., 2017).

Inflammatory responses to monosodium urate (MSU) crystals.

We recently clarified the molecular and cellular mechanisms that are engaged to trigger the violent inflammatory response that characterizes an acute crisis of gout (Mariotte et al., Theranostics, 2019). For this, we developed a mouse model in which the subcutaneous injection of preformed MonoSodium Urate (MSU) crystal induced a severe and transient inflammatory response that, according to pharmacological, clinical and molecular criteria, accurately mimics human gout. However, using several genetically modified mouse lines, our laboratory has shown that, while the inflammatory symptoms that appear upon MSU crystal injection strictly require Il-1b gene expression, the upstream molecular mechanisms driving mature IL-1b production are less well characterized. In fact, the laboratory has reported that, opposite to what was shown in vitro, nor the NLRP3 or AIM2 inflammasomes, neither Caspase 1 or 11, are absolutely required to mount an MSU-dependent inflammatory response in vivo. These observations raise several important issues:

            – In vitro, LPS is classically used to initiate the priming signal (also named signal I) that activates the NF-kB pathway leading to Il-1b gene expression. MSU crystals then trigger the activation of the NLRP3 inflammasome that is capable of Caspase 1-dependant cleavage of pro IL-1b to its mature and bioactive form. The scenario appears more complex in vivo, where the injection of MSU crystals only (free of LPS as demonstrated) triggers a massive production of IL-1b. The upstream receptors that, once activated, induce NF-kB signaling are elusive. Nevertheless, high throughput transcriptomic analyses (RNAseq) performed in P. Georgel’s lab point to a potential involvement of TLR 1, 2 and/or 6 in this process.

            – If caspase 1 and 11 are not critical in the MSU-dependent production of mature IL-1b, is it possible that other caspases or proteases might participate in this process? Their identification will be a major focus of our lab in the future.

Finally, we also used to acute uratic inflammation model in mice to confirm that the topical application of Aldara reduces the cellular infiltrate which is mainly composed of neutrophils. This work provides an exciting novel option to treat gout patients in which colchicine, the standard of care, might be inappropriate and reveal important anti inflammatory effects of type I interferons. The elucidation of the pathways involved in the connection between type I or II interferons and Il-1b expression will require extensive efforts from our lab in the future.

COLLABORATIONS

Our group has established multiple collaborations with national and international scientists: Stephan Bluml and Gernot Schabbauer (Medical University, Vienna, Austria), Bradley Gelfand and Jayakrishna Ambati (University of Virginia, VA, US), Florence Apparailly (INSERM U1183, Montpellier), David Moulin and Laurent Peyrin-Biroulet (Université de Lorraine, Vandœuvre-Lès-Nancy), Bernard Ryffel and Valérie Quesniaux (CNRS, UMR7355, Orleans), Roméo Ricci (IGBMC, Strasbourg).

FUNDING

  • ANR-PRTS
  • SPIRALE Project
  • PHC Hubert Curien
  • Program Amadeus

Relevant publications

  1. Philippe L, Alsaleh G, Suffert G, Meyer A, Georgel P, Sibilia J, Wachsmann D and Pfeffer S. TLR2 Expression Is Regulated by MicroRNA miR-19 in Rheumatoid Fibroblast-like Synoviocytes. J Immunol. (2012) Jan 1;188(1):454-61.
  2. Philippe L, Alsaleh G, Pichot A, Ostermann E, Zuber G, Frisch B, Sibilia J, Pfeffer S, Bahram S, Wachsmann D, Georgel P. MiR-20a regulates ASK1 expression and TLR4-dependent cytokine release in rheumatoid fibroblast-like synoviocytes. Ann Rheum Dis (2013);72:1071–1079.
  3. Philippe L, Alsaleh G, Bahram S, Pfeffer S and Georgel P. The miR-17~92 cluster: a key player in the control of inflammation during rheumatoid arthritis. (2013) Front. Immunol. 4:70.
  4. Ostermann E, Ruer J, Pfeffer S, Bahram S, Soulas-Sprauel P and Georgel P. MicroRNAs: Fine-tuners of Type I Interferon-dependent inflammatory responses. Current Trends in Immunol. (2013). Vol 14; pp 35-44.
  5. Alsaleh G, François A, Philippe L, Gong YZ, Bahram S, Cetin S, Pfeffer S, Gottenberg JE, Wachsmann D, Georgel P and Sibilia J. miR-30a-3p negatively regulates BAFF synthesis in Systemic Sclerosis and Rheumatoid Arthritis fibroblasts. PLoS One. (2014) Oct 31;9(10):e111266.
  6. Gelfand BD, Wright CB, Kim Y, Yasuma T, Yasuma R, Li S, Fowler BJ, Bastos-Carvalho A, Kerur N, Uittenbogaard A, Han YS, Lou D, Kleinman ME, McDonald WH, Núñez G, Georgel P, Dunaief JL, Ambati J. Iron Toxicity in the Retina Requires Alu RNA and the NLRP3 Inflammasome. Cell Rep. (2015) Jun 9. pii: S2211-1247(15)00549-5.
  7. Alsaleh G, Nehmar R, Blüml S, Schleiss C, Ostermann E, Dillenseger JP, Sayeh A, Choquet P, Dembele D, Francois A, Salmon JH, Paul N, Schabbauer G, Bierry G, Meyer A, Gottenberg JE, Haas G, Pfeffer S, Vallat L, Sibilia J, Bahram S, Georgel P. Reduced DICER1 expression bestows rheumatoid arthritis synoviocytes proinflammatory properties and resistance to apoptotic stimuli. Arthritis Rheumatol. (2016) Arthritis Rheumatol. 1839-48. doi: 10.1002/art.39641.
  8. Messer L, Alsaleh G, Georgel P, Carapito R, Waterham HR, Dali-Youcef N, Bahram S Sibilia J. Homozygosity for the V377I mutation in mevalonate kinase causes distinct clinical phenotypes in two sibs with hyperimmunoglobulinaemia D and periodic fever syndrome (HIDS). RMD Open. (2016) Mar 7;2(1):e000196.
  9. Meyer A., Alsaleh G., Heuschling C., Gottenberg JE., Georgel P., Geny B., Bahram S. and Sibilia J. Dermatomyositis flare on imiquimod therapy highlights a crucial role of aberrant TLR7 signalling. RMD Open. (2016). Oct 31;2(2):e000294.
  10. Saferding V, Puchner A, Goncalves-Alves E, Hofmann M, Bonelli M, Brunner JS, Sahin E, Niederreiter B, Hayer S, Kiener HP, Einwallner E, Nehmar R, Carapito R, Georgel P, Koenders MI, Boldin M, Schabbauer G, Kurowska-Stolarska M, Steiner G, Smolen JS, Redlich K, Blüml S. MicroRNA-146a governs fibroblast activation and joint pathology in arthritis. J Autoimmun. (2017) Aug;82:74-84.
  11. Zhang Z, Meszaros G, He WT, Xu Y, de Fatima Magliarelli H, Mailly L, Mihlan M, Liu Y, Puig Gámez M, Goginashvili A, Pasquier A, Bielska O, Neven B, Quartier P, Aebersold R, Baumert TF, Georgel P, Han J and Ricci R. Protein kinase D at the Golgi controls NLRP3 inflammasome activation. J Exp Med. (2017) Sep 4;214(9):2671-2693.
  12. Nehmar R, Alsaleh G, Voisin B, Flacher V, Mariotte A, Saferding V, Puchner A, Niederreiter B, Vandamme T, Schabbauer G, Kastner P, Chan S, Kirstetter P, Holcmann M, Mueller C, Sibilia J, Bahram S, Blüml S and Georgel P. Therapeutical modulation of plasmacytoid dendritic cells in experimental arthritis. Arthritis Rheumatol. (2017) Nov;69(11):2124-2135.
  13. Schwinté P, Mariotte A, Anand P, Keller L, Idoux-Gillet Y, Huck O, Fioretti F, Tenenbaum H, Georgel P, Wenzel W, Irusta S, Benkirane-Jessel N. Anti-inflammatory effect of active nanofibrous polymeric membrane bearing nanocontainers of atorvastatin complexes. Nanomedicine (Lond). (2017) Dec;12(23):2651-2674.
  14. Bonelli M, Puchner A, Goeschl L, Hayer S, Niederreiter B, Steiner G, Tillmann K, Plasenzotti R, Podesser B, Georgel P, Smolen J, Scheinecker C and Blüml S. CCR6 controls autoimmune but not innate immunity driven experimental arthritis. J Cell Mol Med. (2018) Nov;22(11):5278-5285.
  15. Carapito R, Carapito C, Morlon A, Paul N, Jacome ASV, Alsaleh G, Rolli V, Tahar O, Aouadi I,  Rompais M, Delalande F, Pichot A, Georgel P , Messer L, Sibilia J, Cianferani S, Van Dorsselaer A and Bahram S. Multi-OMICS analyses unveils STAT1 as a potential modifier gene in mevalonate kinase deficiency. Ann. Rheum. Dis. (2018) Nov;77(11):1675-1687.
  16. Ammari M, Presumey J, Ponsolles C, Roussignol G, Roubert C, Escriou V, Toupet K, Mausset-Bonnefont AL, Cren M, Robin M, Georgel P, Nehmar R, Taams L, Grün J, Grützkau A, Häupl T, Pers YM, Jorgensen C, Duroux-Richard I, Courties G, Apparailly F. Delivery of miR-146a to Ly6Chigh Monocytes Inhibits Pathogenic Bone Erosion in Inflammatory Arthritis. Theranostics. (2018) Nov 13;8(21):5972-5985.
  17.  Nehmar R, Mariotte A, de Cauwer A, Sibilia J, Bahram S and Georgel P. Therapeutic Perspectives for Interferons and Plasmacytoid Dendritic Cells in Rheumatoid Arthritis. Trends Mol Med. (2018) Apr;24(4):338-347.
  18. De Cauwer A, Mariotte A, Sibilia J, Bahram S and Georgel P. DICER1: a key player in rheumatoid arthritis, at the crossroads of cellular stress, innate immunity and chronic inflammation in ageing. (2018) Front. Immunol. fimmu.2018.01647.
  19. Mariotte A, De Cauwer A, Po C, Abou-Faycal C, Pichot A, Paul N, Aouadi I, Carapito R, Frisch B, Macquin C, Chatelus E, Sibilia J, Armspach JP, Bahram S and Georgel P. NLRP3- and AIM2-autonomy in a mouse model of MSU crystal-induced acute inflammation in vivo highlights imiquimod-dependent targeting of Il-1β expression as relevant therapy for gout patients. bioRxiv 772756; doi: https://doi.org/10.1101/772756. (2019) Theranostics. In Press