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MiR-20a regulates ASK1 expression and TLR4-dependent cytokine release in rheumatoid fibroblast-like synoviocytes
  1. Lucas Philippe1,
  2. Ghada Alsaleh1,
  3. Angélique Pichot1,
  4. Eleonore Ostermann1,
  5. Guy Zuber2,
  6. Benoit Frisch2,
  7. Jean Sibilia1,
  8. Sébastien Pfeffer3,
  9. Seiamak Bahram1,
  10. Dominique Wachsmann1,
  11. Philippe Georgel1
  1. 1ImmunoRhumatologie Moléculaire, INSERM UMR_S 1109, Université de Strasbourg, Strasbourg, Cedex, France
  2. 2Laboratoire de Conception et Application de Molécules Bioactives, CNRS—UMR 7199, Université de Strasbourg, Illkirch, France
  3. 3Université de Strasbourg, Architecture et Réactivité de l'ARN UPR 9002, Institut de Biologie Moléculaire et Cellulaire du CNRS, Strasbourg, France
  1. Correspondence to Professor Philippe Georgel, ImmunoRhumatologie Moléculaire, INSERM UMR_S 1109, Université de Strasbourg; Centre de Recherche d'Immunologie et d'Hématologie, 4 rue Kirschleger, Strasbourg, Cedex 67085, France; pgeorgel{at}


Objective To evaluate whether miR-20a belonging to the cluster miR-17–92 is a negative regulator of inflammation in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) by modulating expression of apoptosis signal-regulating kinase (ASK) 1, a key component of the toll-like receptors 4 pathway, upstream of p38 mitogen-activated protein kinase.

Methods Evaluation of miR-20a and ASK1 mRNA was performed by RT-qPCR. ASK1 protein expression was assessed by western blotting. Overexpression of miR-20a was performed by transfection of RA FLS and THP-1 cells with miR-20a mimics. Interleukin (IL)-6, CXCL-10, IL-1β and TNF-α release were measured by ELISA. The role of miR-20a in vivo was assessed by IL-6 release from macrophages obtained from mice injected intraperitoneally with vectorised miR-20a mimics.

Results We showed that stimulation of RA FLS with lipopolysacharide (LPS) and bacterial lipoproteins (BLP) induces a drop in expression of miR-20a and that this decrease is associated with an upregulation of ASK1 expression. Using transfection of Ask1 3′UTR reporters, we demonstrate that Ask1 is a direct target of miR-20a. Overexpression of miR-20a led to a global decrease in ASK1 protein in BLP- and LPS-activated cells indicating that miR-20a regulates the expression of ASK1 at the translational level. Transfection of miR-20a mimics decreases IL-6 and CXCL10 release by RA FLS and IL-1β and TNF-α by activated THP-1 cells but only in response to LPS. Last, injection of vectorised miR-20a mimics to mice led to a global decrease in ASK1 protein expression and IL-6 secretion in LPS-activated macrophages.

Conclusions Our data point toward an important role for miR-20a in the regulation of pro-inflammatory cytokines release, by controlling ASK1 expression in RA FLS.

  • Fibroblasts
  • Rheumatoid Arthritis
  • Inflammation
  • Cytokines

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