Nicotinamide Induces Apoptosis Of F9 Mouse Teratocarcinoma Stem Cells By Down Regulation Of SATB1 Expression

The special AT-rich sequence binding protein 1(SATB1) is a tissue-specific and time-specific expression of nuclear matrix-binding protein. In recent years, there were some study reported that SATB1 was over-expression in many tumor cells and the expression was associated with the migration and proliferation of tumors. Nicotinamide, a component of vitamin B3, could change the cell’s survival way and induce some tumor cells apoptosis. We analyzed cell proliferatin, cell cycle, cell apoptosis by a series of experiments after cells were treated with nicotinamide or siRNA. The results are as follows:1. The result of RT-QPCR indicated that SATB1 was over-expression in F9 cells. When we tried to use different concentrations of NA(0 mmol/L, 1.5 mmol/L, 2 mmol/L, 2.5 mmol/L) to treat F9 cells, and analyzed the expression of SATB1 by RT-QPCR and western blotting. We found that the expression of SATB1 was decreased significantly in nicotinamide-treated groups than that in the control group, and significant negative correlation was observed with the increasing of nicotinamide concentration. In order to confirm the decreased of SATB1 expression was associated with nicotinamide, we then cultured the cells which ever treated with 0 mmol/L, 1.5 mmol/L, 2.0 mmol/L, 2.5 mmol/L nicotinamide with fresh medium without nicotinamide for 36 h. And the expression of SATB1 was also analyzed by RT-QPCR and western blotting, we found the SATB1 expression increased significantly. It indicated that it was the function of nicotinamide down-regulate SATB1 expression.2. We analyzed the cell cycle and apoptosis by flow cytometry. We found that most cells were arrested to G1 phase and the apoptosis rate was significantly increased in nicotinamide-treated groups. A CCK-8 kit was used to decide whether nicotinamide suppressed the proliferation of the F9 cells. The results indicated that the proliferation rates were significantly lower in nicotinamide-treated groups than those in the control group. But proliferation rates both at 12 h and 24 h were not significantly different among the three nicotinamide-treated groups. However the cell proliferation was significantly different among the three NA treatment groups both at 36 h, 48 h and with the increasing of nicotinamide, the proliferation rates were decreased significantly. It indicated that nicotinamide could inhibited F9 cells proliferation and induced apoptosis of F9 cells.3. We successfully constructed and screened RNAi sequence si-984 targeting SATB1 gene. We found that expression levels of SATB1 were decreased significantly after interfered with si-984 compared with control. Moreover, we found that most of F9 cells were blocked at G1 phase post transfection with si-984. At the same time, the cell proliferation was significantly restrained, and the apoptosis rate was significantly increased after down-regulation of SATB1 by transfecting with si-984 when compared with control cells.In summary, the present study described nicotinamide is an apoptosis inducer in MF9 cells, it could promote growth arrest and apoptosis of MF9 cells by down regulation of SATB1 expression. The results will not only provide theoretical basis for the application of nicotinamide in tumor research, but also make an important contribution to further understanding of tumor’s treatment.