Mediation of interleukin-1beta-induced transforming growth factor beta1 expression by activator protein 4 transcription factor in primary cultures of bovine articular chondrocytes: possible cooperation with activator protein 1.
OBJECTIVE
Interleukin-1 (IL-1) and transforming growth factor beta1 (TGFbeta1) play major roles in osteoarticular diseases, exerting opposite effects on both the catabolism and anabolism of cartilage matrix. Previous findings suggest that IL-1 and TGFbeta1 could function in a feedback interaction. However, the effect exerted by IL-1 on expression of TGFbeta by articular chondrocytes is, so far, poorly understood. The present study was carried out to determine the influence of IL-1beta on the expression of TGFbeta1 by bovine articular chondrocytes (BACs) in primary culture.


METHODS
BAC primary cultures were treated with IL-1beta, and TGFbeta1 messenger RNA (mRNA) steady-state levels and protein expression were measured by real-time reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Transient transfection of TGFbeta1 gene promoter constructs was performed to delineate the DNA sequences that mediate the IL-1beta effect. Electrophoretic mobility shift assays (EMSAs) and supershift analysis were used to characterize the transcription factors binding to these sequences.


RESULTS
Cultured BACs responded to IL-1beta exposure by exhibiting an increase of TGFbeta1 expression at both the mRNA and protein levels. The effect was found to be mediated by a major 80-bp sequence located between -732 and -652 upstream of the transcription initiation site. EMSA and supershift analysis revealed that the transcription factors activator protein 4 (AP-4) and AP-1 specifically bound to the -720/-696 part of this sequence under IL-1beta treatment. Overexpression of AP-4 in the BAC cultures resulted in stimulation of the transcriptional activity of the -732/+11 TGFbeta1 promoter construct through the same IL-1beta-responsive element.


CONCLUSIONS
IL-1beta induces an increase of TGFbeta1 in articular chondrocytes through activation of AP-4 and AP-1 binding to the TGFbeta1 gene promoter. These findings may help us understand the role of IL-1beta in the disease process. Notwithstanding its deleterious effect on cartilage, IL-1 could initiate the repair response displayed by injured cartilage in the early stages of osteoarthritis through its ability to enhance TGFbeta1 expression by local chondrocytes.
Andriamanalijaona
R
R
Laboratory of Connective Tissue Biochemistry, Caen, France.
Felisaz
N
N
Kim
S-J
SJ
King-Jones
K
K
Lehmann
M
M
Pujol
J-P
JP
Boumediene
K
K
D000818:Animals; D001483:Base Sequence; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D015180:Electrophoresis, Gel, Two-Dimensional; D024202:Electrophoretic Mobility Shift Assay; D004797:Enzyme-Linked Immunosorbent Assay; D007375:Interleukin-1 / Q000494:pharmacology*; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000737:chemistry; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000096:biosynthesis* / Q000235:genetics; D053773:Transforming Growth Factor beta1
D004268:DNA-Binding Proteins; D007375:Interleukin-1; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D014157:Transcription Factors; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C055473:enhancer-binding protein AP-4