Ligand-induced downregulation of receptors for TGF-beta in human osteoblast-like cells from adult donors

High concentrations of transforming growth factor b (TGF-beta) are found in the bone matrix, reflecting a pivotal role of this growth factor in the co upling of bone resorption and formation. TGF-beta strongly stimulates the s ynthesis of extracellular matrix proteins, but in vitro studies show an inh ibitory effect on the final mineralization process, which in vivo occurs de spite high concentrations of TGF-beta. Little is known about how bone-formi ng cells respond to different concentrations of TGF-beta and ii they can tr ansiently adapt receptor numbers in order to modulate cellular activity. Ag ainst this background, we studied the cell-surface expression of TGF-beta r eceptors (T beta R) I, II and III (betaglycan) on human osteoblast-like cel ls from adult donors, and examined the T beta R presentation on these cells after a preceding exposure to TGF-beta 1. Affinity crosslinking studies wi th disuccinimidylsuberate showed the presence of all three receptor types. Preincubation with TGF-beta 1 markedly reduced I-125-TGF-beta 1 binding in a time-dependent and dose-dependent manner and revealed a 95% reduction aft er an 18-h preincubation with 200 pM TGF-beta 1. In parallel, Scatchard ana lysis showed that the binding affinity did not change as a consequence of T GF-beta 1 preincubation. Immunoblotting analyses revealed an almost complet e disappearance of immunoreactive T beta R-II and T beta R-III proteins aft er a 24-h preincubation with TGF-beta 1. Using semi-quantitative reverse tr anscription PCR, no effect of TGF-beta 1 on the expression of T beta R-II m RNA was observed. These studies demonstrate a ligand-induced downregulation of T beta Rs-II and -III on human osteoblast-like cells, without any evide nce for recovery within the first 24 h, both in the presence and after the removal of the ligand. The underlying mechanism appears to be based on post -transcriptional events. The results suggest that high concentrations of ac tive TGF-beta 1 decrease the responsiveness of osteoblasts towards this gro wth factor.