C-FOS PROTOONCOGENE IS INVOLVED IN THE MITOGENIC EFFECT OF TRANSFORMING GROWTH-FACTOR-BETA IN OSTEOBLASTIC CELLS

We investigated the contribution of c-fos protooncogene in the mitogenic effect of transforming growth factor-beta (TGF beta) in serum-deprived, confluent rat calvaria osteoblastic cells. The TGF beta-induced growth in these cells was associated with an immediate and transient c-fos mRNA accumulation, similar to the inductive effect of fetal calf serum. To assess the role of c-fos in the response to TGF beta, we used a c-fos antisense (AS) oligonucleotide displaying duplex formation with rat c-fos mRNA. Studies of AS and sense (S) uptake by osteoblastic cells demonstrated that incorporation of labeled oligomers was maximal at 2 h, and the incorporated AS oligonucleotide remained intact for 24 h. Immunofluorescence analysis of c-Fos-labeled cells demonstrated that AS, but not S, oligonucleotide reduced c-Fos protein expression, suggesting specific efficient inhibition of c-fos translation by the AS oligomer. Proliferation assays showed that cell growth induced by fetal calf serum was inhibited by the AS, but not by the S oligonucleotide, in both normal rat osteoblasts and ROS 17/2.8 osteosarcoma cells, demonstrating efficient and specific blockage of cell growth by the AS oligomer. The mitogenic effect of TGF-P was abolished in cells cultured in the presence of AS, whereas S had no effect, showing that c-fos is required for TGF beta-induced osteoblast cell growth. The results show that the induction of c-fos is implicated in the mitogenic effect of TGF beta in osteoblastic cells and provide a cellular mechanism involved in the response of these cells to TGF beta.