Transcriptional and proteolytic regulation of the insulin-like growth factor-I system of equine articular chondrocytes by recombinant equine interleukin-1beta.
Interleukin-1 (IL-1) and insulin-like growth factor-I (IGF-I), which have opposing effects on matrix metabolism within articular cartilage, are thought to play prominent roles in the pathogenesis of osteoarthritis. To better understand the link between these anabolic (IGF-I) and catabolic (IL-1) stimuli, we examined exogenous IL-1 regulation of the IGF-I signaling system of articular chondrocytes (ACs). Equine ACs from non-arthritic stifle joints were expanded in monolayer culture, encapsulated for 10 days in alginate beads, and stimulated as high-density monolayers with recombinant equine IL-1beta (0, 1, 10 ng/ml) for 48 h. IL-1beta enhanced expression of IGF-IR levels, as determined by both [125I]-IGF-I binding studies and Western blotting, while reducing the concentration of endogenous IGF-I detected in conditioned media by radioimmunoassay. Western ligand blotting revealed that chondrocytes primarily secreted IGF binding proteins (IGFBPs) with molecular weights of 28-30 and 32-34 kDa, which were identified as IGFBPs 5 and 2, respectively, and that IL-1beta treatment diminished IGFBP-2, the prominent homolog in conditioned media. Northern blot analysis suggested IL-1beta regulation of IGF-I and, to some extent, IGF-IR was mediated by transcription; however, the cytokine did not affect IGFBP-2 expression. To test for evidence of proteolysis by matrix metalloproteinases (MMPs), additional cultures were co-incubated with inhibitors for MMPs 2/9, 3, and 8. IGFBP-2 suppression was partially reversed by gelatinase (MMP-2/9) inhibition. In summary, these findings further delineate the role of IL-1 as a key regulator of the IGF-I system within articular cartilage, demonstrating that regulation occurs through both direct (transcriptional) and indirect (proteolytic) mechanisms.
Porter
Ryan M
RM
Department of Chemical Engineering, Virginia Polytechnic Institute & State University, Blacksburg, Virginia 24061-0211, USA.
Akers
R Michael
RM
Howard
Rick D
RD
Forsten-Williams
Kimberly
K
D000818:Animals; D002358:Cartilage, Articular / Q000378:metabolism*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism*; D017077:Culture Media, Conditioned; D005786:Gene Expression Regulation / Q000187:drug effects*; D006736:Horses / Q000378:metabolism*; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000378:metabolism; D018972:Insulin-Like Growth Factor Binding Protein 3 / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000378:metabolism*; D007375:Interleukin-1 / Q000494:pharmacology*; D061965:Matrix Metalloproteinase Inhibitors; D008954:Models, Biological; D011499:Protein Processing, Post-Translational; D017526:Receptor, IGF Type 1 / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D017077:Culture Media, Conditioned; D018971:Insulin-Like Growth Factor Binding Protein 2; D018972:Insulin-Like Growth Factor Binding Protein 3; D007375:Interleukin-1; D061965:Matrix Metalloproteinase Inhibitors; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1