All-trans retinoic acid suppresses interleukin-6 expression in interleukin-1-stimulated synovial fibroblasts by inhibition of ERK1/2 pathway independently of RAR activation.
BACKGROUND
Interleukin-6 (IL-6) is thought to play a pathogenic role in rheumatoid arthritis and synovium is a major source of IL-6 release. We investigated the ability of retinoids to suppress IL-6 expression in IL-1-stimulated synovial fibroblasts, with special care to the contribution of retinoic acid receptor (RAR) and retinoid X receptor (RXR) subtypes, and the implication of the mitogen-activated protein kinase (MAPK) pathway.


METHODS
RAR-alpha, -beta, and -gamma and RXR-alpha, -beta, and -gamma levels were determined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) or Western blot in rat synovial fibroblasts stimulated with 10 ng/mL of IL-1beta. Stimulated levels of IL-6 were assessed by RT-qPCR or immunoassays in the presence or absence of 1 microM all-trans retinoic acid (ATRA) (RAR agonist) or 0.3 microM BMS-649 (RXR agonist). The contribution of RAR subtypes was checked with selective agonists or small interfering RNAs. The effect of ATRA on upstream MAPK (p38 MAPK, c-Jun N-terminal kinase [JNK], and extracellularly regulated kinase 1/2 [ERK1/2]) was assessed by Western blot, and the contribution of the ERK1/2 pathway to the activation of pro-inflammatory transcription factors was studied by TransAm assays.


RESULTS
Synovial fibroblasts expressed all RAR and RXR subtypes except RXR-gamma. In IL-1-stimulated cells, ATRA, but not BMS-649, reduced IL-6 expression whereas selective RAR agonists were inactive. The inhibitory effect of ATRA on IL-6 was not affected by the silencing of RAR subtypes. ATRA also reduced the phosphorylation of ERK1/2, but not of p38 MAPK or of JNK. The suppressive effect of ATRA on the activation of activator protein-1 (AP-1) and nuclear factor-IL-6 (NF-IL-6) was reproduced by the MEK1 (mitogen-activated protein extracellularly regulated kinase kinase 1) inhibitor PD-98059, whereas ATRA and PD-98059 had no effect on NF-kappaB activation.


CONCLUSIONS
Among RAR and RXR agonists, only ATRA inhibited IL-1-induced IL-6 expression in rat synovial fibroblasts by inhibiting ERK1/2 pathway and subsequent activation of AP-1 and NF-IL-6 independently of RAR.
Kirchmeyer
Mélanie
M
Laboratoire de Physiopathologie et Pharmacologie Articulaires, UMR 7561 CNRS-Nancy Université, Vandoeuvre-lès-Nancy, France. melkirch2002@yahoo.fr
Koufany
Meriem
M
Sebillaud
Sylvie
S
Netter
Patrick
P
Jouzeau
Jean-Yves
JY
Bianchi
Arnaud
A
D000818:Animals; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D007375:Interleukin-1 / Q000494:pharmacology*; D015850:Interleukin-6 / Q000096:biosynthesis*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D018168:Receptors, Retinoic Acid / Q000819:agonists / Q000378:metabolism*; D013583:Synovial Membrane / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014212:Tretinoin / Q000494:pharmacology*; D000093242:Interleukin-6 Inhibitors
D007375:Interleukin-1; D015850:Interleukin-6; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D018168:Receptors, Retinoic Acid; D014212:Tretinoin; D000093242:Interleukin-6 Inhibitors