Enhancement of Toluidine Blue Staining by Transforming Growth Factor-β, Insulin-Like Growth Factor and Growth Hormone in the Temporomandibular Joint of Aged Mice

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Abstract
Osteoarthritic lesions appear in the articular cartilage of the temporomandibular joint of mice aged 7 months and older. Reduced rate of proteoglycan (PG) synthesis leading to destruction of the articular cartilage was observed in this joint. The purpose of the present study was to test the ability of transforming growth factor-β1 (TGF-β1), insulin-like growth factor-1 (IGF-1) and growth hormone (GH) to induce PG synthesis in joint cartilage of aged animals and to compare it with the effect of interleukin-1α (IL-1α). Mandibular condyle explants from 18-month-old mice were cultured up to 72 h in serum-free medium, supplemented with IL-1α (TGF-β1 (0.1–5.0 ng/ml), TGF-β1 (1.0 ng/ml) + IGF-1 (2 ng/ml) or GH (10 ng/ml). The incorporation of 35S-SO4 into sulfated PG was tested. Cartilage samples were processed for histomorphometry using sections stained with 0.1% toluidine blue (TB), pH 1.8. Results indicated that in cultures supplemented (48 h) with either TGF-β, TGF-β + IGF-1 or with GH, an increased height and area of TB-positive staining as well as increased incorporation of 35S-SO4 into sulfated PG were observed. In contrast, the cytokine IL-1α exerted an inhibitory effect on TB staining and on 35S-SO4 incorporation. The present study demonstrated that in vitro supplementation of IL-1α to mandibular condyle cartilage reduced the height and area of TB staining and incorporation of 35S-SO4, whereas TGF-β1, TGF-β1 + IGF-1 or GH increased the height and area of TB staining and increased incorporation of 35S-SO4. The two parameters used to identify increased PG synthesis were shown to reveal similar results and were useful for studying the dynamic events taking place in cartilage destruction and repair in osteoarthritis.