Transforming Growth Factor-β and Insulin-like Growth Factor-1 Restore Proteoglycan Metabolism of Bovine Articular Cartilage After Depletion by Retinoic Acid

Abstract

Previous studies showed that retinoic acid is a powerful resorbing agent for articular cartilage at physiological doses (10⁻⁸ to 10⁻¹⁰ M); the possible role of individual cytokines in the reversal of this effect is now explored in bovine articular cartilage organ cultures. Seven days of treatment with the retinoid under serum-free conditions, at 1 × 10⁻⁸ M, led to a suppression of proteoglycan synthesis of 90 ± 5% (n = 6; n = cultures from different animals; mean ± SD) and to a net loss of 64 ± 14% (n = 6). Removal of the retinoid from the feeding medium did not significantly increase proteoglycan synthesis nor diminish the further loss of proteoglycans. Thus, transforming growth factor-β (TGF-β) and insulin-like growth factor-1 (IGF-1), cytokines which independently maintain proteoglycan homeostasis (Morales and Roberts, 1988, J. Biol. Chem. 263, 828; and Luyten et al., 1988, Arch. Biochem. Biophys. 267, 416), were tested. TGF-β (10 ng/ml) or IGF-1 (10 ng/ml) added for 7 days to serum-free medium following retinoic acid treatment led to recoveries of proteoglycan synthesis of 74 ± 24% (n = 12) and 69 ± 18% (n = 12), respectively, as compared to controls switched from serum-free conditions to corresponding cytokine treatments. TGF-β + IGF-1 restored activity to 95 ± 17% (n = 12) of controls. TGF-βs 1-3 exhibited identical responses in control and experimental cultures. IGF-2 replaced IGF-1, but a fourfold higher concentration was required; insulin also had IGF-1-like effects, but even at 500 ng/ml it was 25% less effective than IGF-1. In contrast to the cultures switched from retinoic acid treatment to serum-free conditions, the cultures switched to IGF-1, TGF-β, or IGF-1 + TGF-β were stabilized from further proteoglycan loss by the treatment; after 1 week, tissue levels were 97 ± 19, 96 ± 22, and 114 ± 15% (n = 6), respectively, compared to the content before switching. Measurements of catabolism were in agreement with these observations. It is proposed that retinoic acid, TGF-β, and IGF-1 are parts of an endogenous system involved in the reversible modulation of proteoglycan homeostasis in articular cartilage. While the cytokines are able to reestablish the balance of the metabolic pathways to prevent further loss of proteoglycans, the finding that they did not replenish lost material during the 1-week recovery suggests that other factors, such as mechanical forces and/or other cytokines may be contribute to a more effective repair.