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Cytokine regulation of cartilage‐derived retinoic acid‐sensitive protein (CD‐RAP) in primary articular chondrocytes: suppression by IL‐1, bfGF, TGFβ and stimulation by IGF‐1
Author(s) -
Kondo Seiji,
Cha Steven H.,
Xie WeiFen,
Sandell Linda J.
Publication year - 2001
Publication title -
journal of orthopaedic research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.041
H-Index - 155
eISSN - 1554-527X
pISSN - 0736-0266
DOI - 10.1016/s0736-0266(00)00068-1
Subject(s) - retinoic acid , microbiology and biotechnology , chondrocyte , chondrogenesis , type ii collagen , chemistry , messenger rna , gene expression , cartilage , transforming growth factor , reporter gene , biology , gene , biochemistry , anatomy
Cartilage‐derived retinoic acid‐sensitive protein (CD‐RAP) is a secreted protein identified in our laboratory by RT‐PCR and differential display [U.H. Dietz, L.J. Sandell. Cloning of a retinoic acid‐sensitive mDNA expressed in cartilage and during chondrogenesis. J. Biol. Chem. 271 (1996) 3311–3316]. It is synthesized by chondrocytes throughout development and down‐regulated by retinoic acid in coordination with type II collagen gene expression. To further explore the regulation CD‐RAP in primary articular chondrocytes, we examined effects of selected cytokines on CD‐RAP gene expression compared to their effects on type II collagen expression. Northern blot analysis showed that expression of CD‐RAP mRNA was suppressed by bFGF, IL‐1β and retinoic acid in coordination with type II collagen mRNA. TGF‐β decreased CD‐RAP expression while increasing type II collagen mRNA whereas both mRNAs were up‐regulated by IGF‐1. In chondrocytes dedifferentiated with retinoic acid, IGF‐1 induced re‐expression of both CD‐RAP and type II collagen mRNAs. The mechanism of stimulation of CD‐RAP by IGF‐1 was further investigated. An mRNA stability assay revealed that IGF‐1 had no effect on CD‐RAP or type II collagen mRNA half life, suggesting that the enhancement by IGF‐1 is due to increased gene transcription. To study the transcriptional mechanism, we used the 5′‐flanking region of the CD‐RAP gene fused to a promoter‐less reporter plasmid encoding luciferase. Deletion analysis of the CD‐RAP promoter indicated that an IGF‐1‐responsive element is present between nucleotides –475 and –458. These data indicate that CD‐RAP expression can be regulated by cytokines known to influence chondrocyte metabolism and that IGF‐1 up‐regulates CD‐RAP gene expression through a transcriptional mechanism. © 2001 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved.