Proliferation Effect Of NS In Embryonic Esophageal Carcinoma Cells

Background China’s esophageal cancer incidence and mortality rates are much higher than the world average level, the development of esophageal cancer and invasion transfer mechanism of the study found that abnormal cell proliferation is closely related to the occurrence of esophageal cancer development. Nucleostemin(NS) is an important regulation of cell proliferation factor can maintain the proliferation of stem and carcinoma cell, and keeps cells in the undifferentiated state. On the basis of previous studies,we will use the immortalized Shantou Human esophageal epithelial cell line SHEE and its malignant cell line SHEEC as the research object, determination the expression of NS in SHEE and SHEEC lines, and know the proliferation and cell cycle related index. Using RNA interference technology to reduce the expression of NS, observe the effect on cell malignant transformation. This project is to reveal the effect on cell proliferation of NS in esophageal carcinoma cell SHEEC has important significance.Objectives To reveal the proliferation of NS in SHEEC cells.Methods 1. This research adopts the immortalized esophageal epithelial cell line SHEE and its malignant cell line SHEEC. By RT-PCR and Western Blot methods respectively to detect the expression of NS changes in the two cell lines,and the cell proliferation relevant indicators of cell cycle protein cyclin B1 and tumor suppressor gene PTEN protein expression levels of change. 2. By si RNA interference technique to transient transfection the immortalized esophageal epithelial malignant cell SHEEC, SHEEC cells were divided into three groups: reagent control group(Mock), nagtive control group(NC) and RNAi group. By RT-PCR and Western Blot methods respectively to detect the expression of NS, cyclin B1 and PTEN changes in the SHEEC cells.3. Detection activity of cell proliferation of SHEEC cells before and after the inhibiting the expression of NS, and flow cytometry to detect cell cycle and apoptosis.4. The statistical analysis was performed using SPSS17.0 statistical software, the two groups were compared using t-test,compared with many groups was the single factor analysis of variance(one-way ANOVA), different groups were compared with LSD method, significant level is 0.05.Results1. Expression of NS, Cyclin B1 and PTEN m RNA(2.047±0.507, 2.214±0.213, 0.668±0.188) in SHEEC was higher than SHEE, P<0.05. PTEN m RNA and protein was higher than that in SHEEC, P<0.05.2. The expression of NS, Cyclin B1 and PTEN protein in SHEE and SHEEC were(1.142±0.113, 3.161±0.108),(1.585±0.056, 2.173±0.058),(1.894±0.341, 1.162± 0.524), P<0.05. 3. After reduce the expression of NS in SHEEC, the expression of Cyclin B1 and PTEN m RNA and protein in Mock, NC and NS-1543 group were(1.057±0.054, 0.998±0.104, 0.621±0.133),(1.069±0.059, 0.999±0.085, 2.319±0.148) and(1.913±0.177, 1.735±0.275, 0.597±0.103),(0.791±0.382, 0.880±0.077, 2.409±0.058), P<0.05. 4. After reduce the expression of NS in SHEEC, the rate of apoptotic cells in Mock, NC and NS-1543 group were(7.233±0.427, 9.318±0.703, 33.667±2.517), P<0.05. The cell cycle of G0/G1 phase cells percentage increased, S phase and G2/M phase cells decreased percentage. 5. The cell proliferation activity after down regulation of NS in the SHEEC cells were different of each groups, compared with the Mock and NC group, the cell proliferation activity was decreased at 24 h, 36 h and 48 h, P<0.05.Conclusion 1. Silencing of NS expression can block the cell cycle at the G0/G1 phase, the inhibition of SHEEC cell proliferation, promote apoptosis, and accompanied by increased expression of Cyclin B1,reduced expression of PTEN. 2. NS can promote proliferation of SHEEC cell.