Apoptosis And Proliferation As Well As Regulation Mechanism Of Cancer Cells Transfected With Somatostatin Plasmid

With a wide variety of cancer spread in the population, development of efficient-low toxicity anti-cancer drugs has become the urgent task to the current scientific research workers. Somatostatin is widely concerned for its characteristics of growth inhibition and the widespread somatostatin receptors. Recent studies have shown that somatostatin and its analogues not only inhibit tumor cell proliferation, but also induced apoptosis. The objective of this study was to determine the effects of the somatostatin gene expression on proliferation, apoptosis, cell cycle of Hela, HepG2 and MCF-7 cells with a series of techniques. Research contents and results are as follows:1 Eukaryotic expression plasmid of somatostatin transfected tumor cells successfully and expresses the target proteinThe concentration of the plasmid pcS/2SS is 1629.7ng/μL, the pEGS/2SS is 1526.9 ng/μL, the pcDNA3.0 is 2072.4 ng/μL, and the pEGFP-N1 is 1770.4 ng/μL.After plasmids of somatostatin transfected into Hela, HepG2 and MCF-7 cells 24h, the transcription process of somatostatin gene was confirmed by RT-PCR, and the localization as well as expression of EGFP fusion protein is observed by confocal microscopy.2 Proliferations of tumor cells are inhibited after transfected with plasmids of somatostatin.Proliferation is assessed using thiazolyl blue tetrazolium bromide (MTT). The experimental groups (pcS/2SS, pcDNA3.0, pEGS/2SS, pEGFP-N1) compared with control group in Hela cells, inhibition rates were 33.59%±3.60%,21.50%±1.51%, 28.08%±1.32%,22.58%±2.12%; and 27.12%±2.06%,20.47%±0.72%,27.64%±1.44%, 24.35%±1.47% in HepG2 cells; and 38.64%±1.52%,25.32%±1.34%,37.40%±1.90%, 20.43%±1.73% in MCF-7 cells. The results show that proliferations of Hela, HepG2 and MCF-7 cells are inhibited after transfection.Cell cycle is analyzed by flow cytometry. Compared with control group, G0/G1 phase was the same proportions, S phase cells are significantly reduced and G2/M phase is significantly increased in Hela cells. Somatostatin would affect the cell cycle of HepG2 cells, but not significantly. In MCF-7 cells, somatostatin would increase the number of the cells in the S phase and reduce the number in the G2/M phase. 3 Apoptosis can be induced after transfected with plasmids of somatostatin.After transfection for 48h, apoptosis (V-FITC/PI) is analyzed by flow cytometry. The results show that transfection can induce significant apoptosis. After transfection for 48h, the apoptosis rates of Hela cells in the control group, experimental groups (pcS/2SS, pcDNA3.0, pEGS/2SS, pEGFP-Nl) were:2.07%±2.88%,17.52%±1.49%, 12.15%±5.14%,16.35%±1.82%,13.21%±1.32%; the apoptosis rates of HepG2 cells were: 0.75%±1.03%,27.62%±1.50%,22.93%±3.49%,26.35%±2.31%,22.15%±1.54%; the apoptosis rates of MCF-7 cells were:4.88%±1.62%,25.06%±1.52%,21.50%±4.29%, 28.61%±1.76%,22.58%±3.21%.After transfection for 48h, four genes related apoptosis are detected by RT-PCR for further study. The results showed that levels expression of pro-apoptotic gene Bax and wild type p53 were up-regulation and anti-apoptotic genes bcl-2 was down regulation compared with controls in Hela cells. Levels expression of anti-apoptotic genes bcl-2 and pro-apoptotic genes (Bax, p53, NFKB) all were up-regulation in HepG2 cells. And in MCF-7 cells. Levels expression of pro-apoptotic genes(Bax, p53) were up-regulation, anti-apoptotic genes bcl-2 and NFKB were down regulation. All the results proved that apoptosis-related genes are changed during apoptosis and apoptosis is a multi-channel interaction.These results indicate that somatostatin gene has significantly effect on apoptosis, proliferation and cell cycle of Hela, HepG2 and MCF-7cells.