J Korean Orthop Res Soc.  2000 Oct;3(2):140-147.

Fibroblast growth factor-induced Thymidylate Synthase activity and expression in the serum-starved UMR 106-01 osteoblast cells

• Kim SW
• Hwang OY
• Lee JD
• Song KY
• Kim DH
• Hong HN
• Chang JS
• Um KH
• Lee SJ

Abstract

PURPOSE
In the present study, the effects of bFGF on the early responses of proliferation of UMR 106-01 osteoblast cells during cell cycle reentry from the latent(G0/G1) to the proliferative periods(S/M) were investigated.
MATERIALS AND METHODS
The synchronized cell culture method using the serum starvation was utilized. After the addition of bFGF, the time courses of protein synthesis, DNA synthesis, thymidylate synthase(TS) activity, TS mRNA level and expression of c-fos were determined.
RESULTS
87% UMR 106-01 cells were synchronized to G0/G1 by serum starvation for seven days in the medium containing 0.1% serum. The protein level began to increase 3 hours after bFGF treatment and reached the maximum at 18 hours. TS activity began to increase 3 hours after the bFGF treatment and reached its peak at 6 hours while its mRNA level, determined by quantitative PCR, reached the maximum at 12 hours. The expression of c-fos protein, determined by western blot analysis and immunocytochemistry, increased 3 hours after bFGF treatment. On the contrary, these prominent changes and responses to bFGF were not observed in the case of using non-synchronized cells cultured in the medium containing 10% serum.
CONCLUSION
Based on these data it can be concluded that bFGF-induced DNA synthesis in the early proliferative phase is due to increases in both TS activity and mRNA amount and that the increase in c-fos expression and TS activity occur before the increase in TS mRNA level.

Keyword

bFGF; serum starvation; UMR 106-01 osteoblast cell; thymidylate synthase; c-fos

MeSH Terms

Blotting, Western
Cell Culture Techniques
Cell Cycle
DNA
Fibroblasts*
Immunohistochemistry
Osteoblasts*
Polymerase Chain Reaction
RNA, Messenger
Starvation
Thymidylate Synthase*
DNA
RNA, Messenger
Thymidylate Synthase