Effects of TGF-beta1 and IGF-1 on proliferation of human nucleus pulposus cells in medium with different serum concentrations.
BACKGROUND
The low proliferative viability of human nucleus pulposus(NP) cells is considered as a cause of intervertebral discs degeneration. Growth factors, such as TGF-beta1 and IGF-1, have been implicated in cell proliferation and matrix synthesis.


OBJECTIVE
To investigate the dose-response and time-course effect of transforming growth factorbeta1(TGF-beta1) and insulin-like growth factor-1(IGF-1) on proliferation of NP cells.


METHODS
3-(4,5-dimethylthiazolyl)-2,5-diphenyl-tetrazolium bromide (MTT) is reduced by dehydrogenase in mitochondria of live cells. The proliferative viability of cells corresponds to the amount of MTT reduced, which is measured with an enzyme-linked immunosorbent assay plate reader. In this study, we assessed dose- and time-dependent effects of NP cells to TGF-beta1 and IGF-1 in medium with different serum concentrations by MTT assay.


METHODS
After release of informed consent, tissue samples of NP were obtained from anterior surgical procedures performed on five donors with idiopathic scoliosis. Isolated cells were cultured in F12 medium supplemented with 10% fetal bovine serum(FBS). Cells were seeded in 96-well plates at 1 x 103 cells/well. After synchronization, medium was replaced by F12 containing 1% or 10% FBS with either single or combination of TGF-beta1 and IGF-1. Dose-response and time-course effect were examined by MTT assay.


RESULTS
In the presence of 1% FBS, the response to IGF-1 was less striking, whereas TGF-beta1 had a remarkably stimulating effect on cell proliferation. In 10% FBS, both of the two growth factors had statistical significant mitogenic effects, especially TGF-beta1. The dose-dependent effect of TGF and IGF on cell proliferation was found within different concentrations of each growth factor(TGF-beta1 1-10 microg/L, IGF-1 10-100 microg/L). The time-course effect showed a significant elevation three days later.


CONCLUSIONS
TGF-beta1 and IGF-1 were efficient to stimulate cell proliferation of human NP cells in vitro with a dose- and time-dependent manner. These results support the therapeutic potentials of the two growth factors in the treatment of disc degeneration.
Zhang
Rongfeng
R
Department of Orthopaedic Surgery, Navy General Hospital of PLA, Beijing, 100037, People's Republic of China. zhangrongfeng200@163.com
Ruan
Dike
D
Zhang
Chao
C