Prostaglandin E2 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 C

Abstract

Insulin-like growth factor-1, IGF-1, is believed to be an important anabolic modulator of cartilage metabolism and its bioactivity and bioavailability is regulated, in part, by IGF-1 binding protein 3 (IGFBP-3). Prostaglandin E2 (PGE2) 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 culture, Western and Western ligand blotting to measure secreted IGFBP-3 protein, and Northern analysis to monitor IGFBP-3 mRNA levels, we demonstrated that PGE2 provoked a 3.9 +/- 1.1 (n = 3) fold increase in IGFBP-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+2 ionophore, ionomycin, mimicked the effects of PGE2 as did the phorbol ester PMA, which activates Ca++/-phospholipid-dependent protein kinase C (PKC). Cyclic AMP mimetics, such as forskolin, IBMX, Ro-20-1724, and Sp-cAMP, inhibited the expression and synthesis of the binding protein. PGE2 did not increase the levels of cAMP or protein kinase A (PKA) activity in chondrocytes. The PGE2 secretagogue, IL-1 beta, down-regulated control levels of IGFBP-3 which could be completely abrogated by pre-incubation with the tyrosine kinase inhibitor, erbstatin, and partially reversed (50 +/- 8%) by KT-5720, a PKA inhibitor. These observations suggested that PGE2 does not mediate the effect of its secretagogue and that IL-1 beta signalling in chondrocytes may involve multiple kinases of diverse substrate specificities. Dexamethasone down-regulated control, constitutive levels of IGFBP-3 mRNA and protein eliminating the previously demonstrated possibility of cross-talk between glucocorticoid receptor (GR) and PGE2 receptor signalling pathways. Taken together, our results suggest that PGE2 modulates IGFBP-3 expression, protein synthesis, and secretion, and that such regulation may modify human chondrocyte responsiveness to IGF-1 and influence cartilage metabolism.

Differential regulation of IGF-binding proteins in rabbit costal chondrocytes by IGF-I and dexamethasone

“…In contrast, IGFBP-2 was the most abundant species in sheep chondrocytes (Borromeo et al 1996, Sunic et al 1998. IGFBP-3 production was prominent in human articular chondrocytes (Olney et al 1995, DiBattista et al 1996 and weaker in IGF-I-stimulated bovine chondrocytes of various origin (Olney et al 1993). Further study will be needed to establish whether the observed differences are caused by species differences, the type of cartilage from which the chondrocytes were derived, or variation in experimental procedures.…”

“…The regulation of expression of IGFBPs and their proteases has been extensively characterized in many types of cultured cells, including osteoblasts (Hassager et al 1992, Conover & Kiefer 1993, Mohan 1993, McCarthy et al 1994 and articular chondrocytes (Froger-Gaillard et al 1989, Olney et al 1993, DiBattista et al 1996, Matsumoto et al 1996a,b, Sunic et al 1998. Much less information is available on IGFBP production by growth plate cartilage.…”

Voltage-Dependent Calcium Channels in Chondrocytes: Roles in Health and Disease

“…This observation was supported by in vitro data; treatment of mouse embryonic limb bud-derived micromass cultures with nifedipine or verapamil during the early stages of differentiation inhibited chondrogenesis, reflecting on an important role of L-type VDCCs during cartilage formation [24]. Furthermore, in high-density primary cultures of human articular chondrocytes, application of verapamil or nifedipine abrogated the stimulatory effect of prostaglandin E2 (PGE2) on IGFBP-3, a protein that regulates the bioactivity and bioavailability of insulin-like growth factor-1 (IGF-1), an important mediator of chondrocyte metabolism [25]. In a followup study, IGF-1, IGF-2 and insulin all increased cytosolic Ca 2+ levels in cultured rabbit articular chondrocytes but through different mechanisms; Ca 2+ influx mediated by L-type VDCCs was involved downstream of IGF-1 and insulin, since verapamil diminished these responses [26].…”

Long-chain polyunsaturated fatty acids: Selected mechanisms of action on bone

“…PGE 2 also promotes the expression of various IGF-binding proteins [77,80,82] suggesting that PGE 2 could keep IGF available for stimulation of osteoblasts at a later phase of bone remodeling [77]. McCarthy et al [48] suggested that the ability of PGE 2 to enhance osteoblastic IGF-I synthesis could explain its anabolic potential, and furthermore suggests a role for PGE 2 in coupled bone remodeling.…”