Research Of Interference Apollon Gene On Proliferation And Apoptosis Of QBC939Cells

Objective:In this experiment, in order to research the efficiency of transfection of Apollon ASODN(AntiOligodeoxynucleotide) on cell proliferation, apoptosis and Apollon protein expression of human cholangiocarcinoma cell QBC939, to provide the basis for subsequent in-depth study.Methods:Application of liposomes (Lipofectamin RNAimax),Apollon ASODN have been transiently transfected into human cholangiocarcinoma subculture QBC939cells. Experiment was divided into:Lip-ASODN group (final concentration200,400,600,800and1600nM), Lip-MSODN(missense oligodeoxynucleotides), empty liposome, cell control group. MTT assay of Inhibition at different times with different concentrations of Apollon ASODN on cell growth and proliferation; choose the best reaction time and concentration; Western Blot detection interference efficiency that Apollon protein expression changes before and after transfection.The apoptosis and cell cycle of QBC939cell was detected by flow cytometry(FCM) after the transfection.Results:Apollon ASODN can be successfully transfected into QBC939cells by liposomes (Lipofectamine RNAimax), which can be detected in higher suppression Efficiency and within a certain range of concentrations in a dose-dependent. MTT results showed that the concentration gradient of five kinds of antisense oligonucleotides(ASODN) on QBC939cells were found to inhibit cell proliferation in200～800nmol/L in a time-and dose-dependent. The QBC939cell inhibition rate was significantly higher within ASODN groups at different concentrations and time than MSODN group and liposome group and the control group cells,when Apollon ASODN concentration by800nmol/L at48h after transfectiont can get maximum inhibition rate that was63.88±0.24, the difference was statistically significant (P <0.05). Western Blot showing:the transfection of Apollon ASODN could significantly inhibit the expression of Apollon protein, the maximum reduction from81.6%, compared with the control group, the difference was statistically significant (P <0.05). PI staining flow cytometry analysis the cell cycle showed that:the role of different concentrations of apollon ASODN on QBC939cells at48h later, with increasing concentration, the percentage of cells showed a significant increase in S phase arrest also, up35.67%. Flow cytometry PI/Annexin V double staining detection of early apoptosis rate shows:after48h, ASODN early apoptosis rate was significantly higher than control group (P<0.05), MSODN group compared with the control group no significant effect on apoptosis (P>0.05), between200-800nmol/1Apollon ASODN dose-dependent manner, with the concentration increased, its early apoptosis rate was significantly higherConclusion:under the help of Liposomes (Lipofectamin RNAimax), Apollon ASODN can be successfully transfected into QBC939cell line,the transfection of Apollon ASODN can significantly decrease the expression of Apollon protein,and obviously inhibit the proliferation of QBC939cell and induce apoptosis of QBC939cells. The experiments show that by blocking the relevant functional areas Apollon gene can effectively suppress the value of cholangiocarcinoma cells and induce cell apoptosis.