Effects of transforming growth factor type β on expression of cytoskeletal proteins in endosteal mouse osteoblastic cells

Abstract

Transforming growth factor β (TGFβ) has been shown to influence the growth and differentiation of many cell types in vitro. We have examined the effects of TGFβ on cell morphology and cytoskeletal organization in relation to parameters of cell proliferation and differentiation in endosteal osteoblastic cells isolated from mouse caudal vertebrae. Treatment of mouse osteoblastic cells cultured in serum free medium for 24 hours with TGFβ (1.5–30 ng/mL) slightly (− 23%) inhibited alkaline phosphatase activity. In parallel, TGFβ (0.5–30 ng/mL, 24 hours) greatly increased cell replication as evaluated by [ 3H]-thymidine incorporation into DNA (157% to 325% of controls). At a median dose (1.5 ng/mL) that affected both alkaline phosphatase and DNA synthesis (235% of controls) TGFβ induced rapid (six hours) cell respreading of quiescent mouse osteoblastic cells. This effect was associated with increased polymerization of actin, α actinin, and tubulins, as evaluated by both biochemical and immunofluorescence methods. In addition, TGFβ (1.5 ng/mL) increased the de novo biosynthesis of actin, α actinin, vimentin, and tubulins, as determined by [ 35S] methionine labeling and fractionation of cytoskeletal proteins using two-dimensional gel electrophoresis. These effects were rapid and transient, as they occurred at six hours and were reversed after 24 hours of TGFβ exposure. The results indicate that the stimulatory effect of TGFβ on DNA synthesis in endosteal mouse osteoblastic cells is associated with a transient increase in cell spreading associated with enhanced polymerization and synthesis of cytoskeletal proteins.