Objective

To determine the specificity of and possible synergism among polypeptide growth factors (PGF) on the net synthesis of proteoglycan and a novel high molecular weight anionic glycoprotein (HMW AG), of roughly 540 kDa, by articular chondrocyte cultures.

Methods

Confluent articular chondrocyte cultures were labeled with either [35S] SO4 or [3H] glucosamine and [35S] SO4 and stimulated by adding the individual PGF or combinations of the PGF. Alcian blue dye precipitation was used for direct rapid quantification of newly synthesized proteoglycans from the media of the 35S labeled cultures. To assess the effects of the PGF on the synthesis of the HMW AG, a toluidine blue dye batch precipitation method was used for isolation of anionic glycoconjugates from the media of the 3H and 35S labeled articular chondrocyte cultures, followed by chondroitinase ABC digestion and gradient sodium dodecyl sulfate polyacrylamide gel electrophoresis.

Results

Optimal concentrations of transforming growth factor-beta1 (TGF-beta), insulin-like growth factor-1 (IGF), and platelet derived growth factor (PDGF) stimulated net proteoglycan synthesis per cell by 178, 106, and 101%, respectively. In combinations, TGF-beta+ PDGF, TGF-beta+ IGF, and PDGF+ IGF gave stimulations of proteoglycan synthesis of 418, 384, and 217%, respectively. HMW AG net synthesis was induced by all 3 PGF, but the degree of induction was much greater with TGF-beta.

Conclusion

Net proteoglycan synthesis by articular chondrocyte cultures is synergistically stimulated by the addition of optimal concentrations of combinations of TGF-beta+ PDGF and TGF-beta+ IGF, but not PDGF+ IGE Net HMW AG synthesis is preferentially stimulated by TGF-beta.