Interleukin 17 (IL-17) induces collagenase-3 production in human osteoarthritic chondrocytes via AP-1 dependent activation: differential activation of AP-1 members by IL-17 and IL-1beta.
OBJECTIVE
In osteoarthritic (OA) synovial fluid, many proinflammatory cytokines coexist and stimulate chondrocytes. As interleukin 17 (IL-17) is a catabolic cytokine, we explored its effects on collagenase-3 production. In a comparative manner we identified IL-17 and IL-1beta induced transcription factors mediating upregulation of this enzyme's production.


METHODS
Collagenase-3 levels were determined by ELISA. Transfection experiments of human OA chondrocytes were performed, with the plasmids -1599CAT and -133CAT consisting of 1.6 kb and the first proximal 133 bp containing polyomavirus enhancer A-3 (PEA-3), activating protein-1 (AP-1), and TATA box of the human collagenase-3 promoter, respectively. Electrophoretic mobility shift assays were done with the AP-1 and PEA-3 oligonucleotides derived from the human collagenase-3 promoter sequence. Supershift assays were carried out with the specific antibodies against the Jun and Fos proteins.


RESULTS
IL-17 induced collagenase-3 expression and synthesis, with an EC50 at 10 ng/ml. Transfection experiments with wild-type -1599CAT and -133CAT and their mutated AP-1 or PEA-3 derivatives revealed that the AP-1 site was essential for basal and proinflammatory cytokine induced collagenase-3 promoter activity, whereas the PEA-3 motif exerted a cooperative effect. Of note, in OA chondrocytes, IL-17 and IL-1beta induced collagenase-3 production through AP-1 occurred with differential protein complexes: IL-17 stimulation resulted in FosB activation, while IL-1beta stimulated c-Fos. Data showed a strong activation of JunB only in cells showing a higher collagenase-3 basal level and low cytokine (IL-17 and IL-1beta) inducibility, suggesting this transcription factor protein acts as a negative regulator.


CONCLUSIONS
We demonstrated that IL-17 and IL-1beta induced collagenase-3 production in OA chondrocytes mainly through AP-1 mediated transcriptional activity but with differential protein complexes, suggesting that some AP-1 proteins play a pivotal role in the different cytokine responses in terms of collagenase-3 production. Our data might suggest that JunB protein plays a rate-limiting step in cytokine induced collagenase-3 production in OA chondrocytes.
Benderdour
Mohamed
M
Osteoarthritis Research Unit, Hĵpital Notre-Dame, Centre Hospitalier de l'Université de Montréal, Québec, Canada.
Tardif
Ginette
G
Pelletier
Jean-Pierre
JP
Di Battista
John A
JA
Reboul
Pascal
P
Ranger
Pierre
P
Martel-Pelletier
Johanne
J
D000368:Aged; D001483:Base Sequence; D015152:Blotting, Northern; D019902:Chondrocytes / Q000187:drug effects*; D017364:Collagenases / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D020381:Interleukin-17 / Q000494:pharmacology*; D008297:Male; D053509:Matrix Metalloproteinase 13; D008969:Molecular Sequence Data; D010003:Osteoarthritis / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D018808:Transcription Factor AP-1 / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D015854:Up-Regulation
D007375:Interleukin-1; D020381:Interleukin-17; D018808:Transcription Factor AP-1; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13