title: Regulation of insulin-like growth factor-binding protein-5 by insulin-like growth factor I and interleukin-1alpha in ovine articular chondrocytes.
abstract: Insulin-like growth factors (IGFs) contribute to the maintenance of the cartilage matrix by stimulating proteoglycan synthesis. In contrast, interleukin-1 (IL-1), an inflammatory cytokine, suppresses the synthesis of proteoglycans. In pathological conditions the chondrocytes' responsiveness to IGF-I is decreased, and elevated levels of IGF-binding proteins (IGFBPs) have been implicated as a possible cause. The aim of this study was to investigate the effects of IGF-I and IL-1 on IGFBP production by ovine articular chondrocytes (OAC) and the roles of these IGFBPs in the regulation of proteoglycan synthesis. As revealed by Western ligand and immunoblotting, OACs secreted IGFBP-2 and a 24-kDa IGFBP in culture medium under basal conditions. Exposure of the cells to IGF-I for 48 h resulted in the appearance of IGFBP-5 in the medium. Des(1-3)IGF-I, an IGF-I analog with reduced affinity for IGFBPs, also increased the level of IGFBP-5, but to a lesser extent than IGF-I, whereas LR3IGF-I, which has virtually no affinity for IGFBPs, had no effect on IGFBP-5. Furthermore, IGFBP-5 underwent a time-dependent limited proteolysis when incubated with OAC-conditioned medium, degrading into 22- and 16-kDa fragments. The degradation of IGFBP-5 was significantly inhibited by IGF-I, but not by des(1-3)IGF-I or LR3IGF-I. Basic fibroblast growth factor, transforming growth factor-beta, and platelet-derived growth factor had no effect on OAC IGFBPs. However, IL-1alpha increased the IGFBP-5 level in a dose-dependent manner, showing maximum activity at 200 U/ml. Furthermore, IL-1alpha, but not IGF-I, induced IGFBP-5 messenger RNA expression, as assessed by Northern blot analysis. Coincubation of IGF-I with IL-1alpha resulted in a substantially increased IGFBP-5 protein level, suggesting a synergism between the mechanisms of action of these two factors. Des(1-3)IGF-I and LR3IGF-I were 10 times more potent than IGF-I in stimulating proteoglycan synthesis, indicating inhibition of IGF-I activity by endogenous IGFBPs. IL-1alpha reduced the IGF-I bioactivity, but had no effect on the activities of the IGF-I analogs, thus implying that locally produced IGFBPs, particularly IGFBP-5, which was substantially increased when IGF-I and IL-1alpha were coincubated, mediated the reduction of the IGF-I activity. Our results demonstrate that IGF-I and IL-1alpha synergistically increase the level of IGFBP-5 in OAC by inhibiting the proteolysis and stimulating the expression of IGFBP-5, respectively. Furthermore, the attenuation of IGF-I-stimulated proteoglycan synthesis by IL-1alpha in OAC appears to be mediated by chondrocyte IGFBPs. We conclude that locally produced IGFBPs, in particular IGFBP-5, may play a critical role in the regulation of cartilage matrix degradation in inflammatory and degenerative arthritides.
authors.affiliation: Department of Medicine, Modbury Public Hospital, South Australia, Australia.
authors.forename: D
authors.initials: D
authors.lastname: Sunic
authors.forename: J D
authors.initials: JD
authors.lastname: McNeil
authors.forename: T E
authors.initials: TE
authors.lastname: Rayner
authors.forename: D L
authors.initials: DL
authors.lastname: Andress
authors.forename: D A
authors.initials: DA
authors.lastname: Belford
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002358:Cartilage, Articular / Q000166:cytology* / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D017077:Culture Media, Conditioned; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D015870:Gene Expression; D015151:Immunoblotting; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000235:genetics / Q000378:metabolism*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007335:Insulin-Like Growth Factor II / Q000378:metabolism; D007375:Interleukin-1 / Q000494:pharmacology*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011509:Proteoglycans / Q000096:biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D012756:Sheep; D016212:Transforming Growth Factor beta / Q000494:pharmacology
chemical_list: D017077:Culture Media, Conditioned; D018975:Insulin-Like Growth Factor Binding Protein 5; D007375:Interleukin-1; D010982:Platelet-Derived Growth Factor; D011509:Proteoglycans; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D016222:Fibroblast Growth Factor 2; D007334:Insulin-Like Growth Factor I; D007335:Insulin-Like Growth Factor II