PROSTAGLANDIN-E(2) UP-REGULATES INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-3 EXPRESSION AND SYNTHESIS IN HUMAN ARTICULAR CHONDROCYTES BY A C-AMP-INDEPENDENT PATHWAY - ROLE OF CALCIUM AND PROTEIN-KINASE-A AND PROTEIN-KINASEC

Insulin-like growth factor-1, IGF-1, is believed to be an important an abolic modulator of cartilage metabolism and its bioactivity and bioav ailability is regulated, in part, by IGF-1 binding protein 3 (IGFBP-3) . Prostaglandin E(2) (PGE(2)) stimulates IGF-1 production by articular chondrocytes and we determined whether the eicosanoid could regulate IGFBP-3 and, as such, act as a modifier of IGF-1 action at a different level. Using human articular chondrocytes in high density primary cul ture, Western and Western ligand blotting to measure secreted IGFBP-3 protein, and Northern analysis to monitor IGFBP-3 mRNA levels, we demo nstrated that PGE(2) provoked a 3.9 +/- 1.1 (n = 3) fold increase in I GFBP-3 mRNA and protein. This effect was reversed by the Ca++ channel blockers, verapamil and nifedipine, and the Ca++/calmodulin inhibitor, W-7. The Ca++ ionophore, ionomycin, mimicked the effects of PGE(2) as did the phorbol ester PMA, which activates Ca++-phospholipid-dependen t protein kinase C (PKC). Cyclic AMP mimetics, such as forskolin, IBMX , Re-20-1724, and Sp-cAMP, inhibited the expression and synthesis of t he binding protein. PGE(2) did not increase the levels of cAMP or prot ein kinase A (PKA) activity in chondrocytes. The PGE(2) secretagogue, IL-1 beta, down-regulated control levels of IGFBP-3 which could be com pletely abrogated by pre-incubation with the tyrosine kinase inhibitor , erbstatin, and partially reversed (50 +/- 8%) by KT-5720, a PKA inhi bitor. These observations suggested that PGE(2) does not mediate the e ffect of its secretagogue and that IL-1 beta signalling in chondrocyte s may involve multiple kinases of diverse substrate specificities. Dex amethasone down-regulated control, constitutive levels of IGFBP-3 mRNA and protein eliminating the previously demonstrated possibility of cr oss-talk between glucocorticoid receptor (GR) and PGE(2) receptor sign alling pathways. Taken together, our results suggest that PGE(2) modul ates IGFBP-3 expression, protein synthesis, and secretion, and that su ch regulation may modify human chondrocyte responsiveness to IGF-1 and influence cartilage metabolism. (C) 1996 Wiley-Liss, Inc.