| Objective: To investigate the expression,function and related mechanisms of CHAF1 A in cervical cancer.Method: The m RNA expression level of CHAF1 A and ERα in cervical cancer tissues and normal cervical tissues adjacent to cancer was detected by RT-PCR,and the protein expression of CHAF1 A and ERα in cervical cancer tissues was detected by immunohistochemical staining;the relationship between the expression of the two genes and clinical characteristics was analyzed to understand their clinical significance;further statistical analysis was carried out to understand the correlation of their expression based on their expression level.Based on the expression of CHAF1 A in cervical cancer cells and human normal immortalized epithelial cells,Siha and He La cells were selected for cell functional assays;lentivirus was used to interfere with the expression of CHAF1 A in cervical cancer cells,and cells transfected with negative control virus were used for control studies.RT-PCR and western blot were used to verify the effective transfection and then the cell proliferation was assessed by the cell colony formation assay,cell proliferation assay,CCK8 assay,RTCA(proliferation)assay.The cell cycle and apoptosis were monitored by flow cytometry assay.Scratch assay and RTCA(migration)assay were performed to detect the effect of CHAF1 A on cell invasiveness.Based on the biological expression of CHAF1 A in cervical cancer cells,changes in the expression levels of key molecules related to proliferation and apoptosis in proliferation-related signaling pathways were further detected by RT-PCR.To further elucidate the upstream and downstream functions of CHAF1 A,changes in ERα expression after knockdown of CHAF1 A were detected.Result: Both CHAF1 A and ERα were differentially expressed in CSCC.The trend of CHAF1 A expression in CSCC was that the rate of positive expression increased with cervical cancer progression,and the increased expression of CHAF1 A was correlated with cervical cancer stage and differentiation with statical significance(P < 0.05).ERα expression was negatively correlated with cervical cancer progression,Erα was correlated with cervical cancer stages,differentiation and lymph node metastasis(P < 0.05),but not with patients’ age and depth of infiltration.r=-0.300 in Spearman’s test revealing that the expression of the two was negatively correlated with p < 0.05,which was statistically significant.The expression of CHAF1 A in cervical cancer cell lines and human immortalized epithelial Ha Ca T cells by RT-PCR showed that the expression of CHAF1 A was higher in cervical cancer cell lines than in Ha Ca T cell lines.The Si Ha and He La cell lines with elevated expression underwent lentivirus-mediated knocked down of CHAF1 A,and the cells transfected with negative control virus were used for cell functional assays.The results showed that the reduced expression of CHAF1 A significantly inhibited the proliferation of cervical cancer cells,promoted apoptosis and reduced the number of cells in S-phase.The migration ability of cervical cancer cells was reduced.The expression of key molecules of proliferation-related signaling pathways was further examined by Western blot,which showed that the levels of phosphorylated PI3K(tyr458)and Akt(ser473)decreased significantly after CHAF1 A knockdown,while the levels of PI3 K and Akt in total proteins remained unchanged,and the levels of total Fox O1 remained unchanged,while the levels of phosphorylated Fox O1 increased.The expression of the apoptosis-related Bcl-2 gene was also significantly decreased after the downregulation of CHAF1 A expression,while the expression of the apoptosis-promoting BAX was increased.Meanwhile,the expression of ERα was increased after the knockdown of CHAF1 A,and there was a negative correlation between the expression of the two genes.Conclusion: CHAF1 A expression was elevated in cervical cancer tissues and cells compared with normal ones.Elevated CHAF1 A expression mediates the malignant biological behavior of cervical cancer. |
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