Effect Of CENP-O On Proliferation And Apoptosis Of Ovarian Cancer Cells And Discussion Of Significance

Background:Ovarian cancer is a common female malignant tumor,and its mortality rate is the highest among gynecological tumors.At present,there is insufficient understanding of the etiology,pathogenesis,recurrence and metastasis of ovarian cancer.The early diagnosis and treatment of ovarian cancer still face great challenges.The sensitivity and specificity of existing ovarian cancer screening methods are still unsatisfactory.With the continuous development of biotechnology,detection instruments and methods,various potential biomarkers and therapeutic targets have initially shown broad application prospects and have attracted widespread attention.Centromere protein O（CENP-O）is a newly discovered structural centromere protein related to cell death.It is necessary for bipolar spindle assembly,chromosome separation,and checkpoint signaling during mitosis^[1].According to reports,CENP-O has been detected to have abnormally high expression in a variety of tumors(such as bladder cancer^[2,3],gastric cancer^[4],etc.)and participates in the regulation of tumor cell proliferation and other processes,but in ovarian cancer Whether there is a high expression of CENP-O and the possible role of CENP-O in the occurrence and development of ovarian cancer have not been reported in related studies.In this study,we designed and constructed a lentiviral vector containing CENP-O RNA interference sequence to infect ovarian cancer cell line SKOV3,knock down the expression of CENP-O gene,detect cell proliferation,apoptosis and other biological function changes and CENP-O in the ovaries The role of cancer cell proliferation and apoptosis,to explore the possibility and value of CENP-O as a target for diagnosis and treatment of epithelial ovarian cancer.Objective:Detect the expression of CENP-O gene in different ovarian cancer cell lines,and construct a stable transfected ovarian cancer cell line with CENP-O gene knockdown to study the effect of CENP-O on the proliferation and apoptosis of ovarian cancer cells.Method:1.Real-time quantitative PCR（qPCR）detects the mRNA expression abundance of CENP-O gene in different ovarian cancer cells（SKOV3,OVCAR-3,ES-2）,and selects the cell line with the highest expression abundance（CENP-O strong positive Cell line）as experimental cells.2.Design and construct a lentiviral vector containing CENP-O RNA interference sequence to infect CENP-O strongly positive cell lines;construct and identify a stable transfected cell line with CENP-O gene knockdown.The knockdown rate of CENP-O gene was detected by qPCR.3.Caspase3/7 test verifies the effect of knockdown of CENP-O gene on cell apoptosis;clone formation experiment and MTT test verifies the effect of knockdown of CENP-O on cell proliferation;Western Blot method detects the protein after CENP-O gene knockdown in cells Changes in the amount of expression.Results:1.The results of RT-PCR showed that the expression abundance of CENP-O gene was high in three different ovarian cancer cell lines,and the expression abundance of SKOV3 was the highest.2.After shRNA lentivirus infection of SKOV3 cells,the expression of CENP-O gene was inhibited（p<0.01）,and the knockdown efficiency reached 79.8%.3.The results of caspase3/7 detection showed that:knocking down the CENP-O gene increased SKOV3 cell apoptosis（p<0.01）;the results of the clone formation experiment showed that knocking down the CENP-O gene reduced the number of SKOV3 cell clones（p<0.01）;The MTT test results showed that the proliferation of SKOV3 cells was reduced after the CENP-O gene was knocked down;the Western Blot test results showed that in SKOV3 cells,the endogenous knockdown of the CENPO gene by the target had a significant knockdown effect at the protein level.Conclusion:1.CENP-O gene is significantly expressed in three ovarian cancer cell lines（SKOV3,OVCAR-3,ES-2）.2.After the CENP-O gene is knocked down,the proliferation and cloning ability of ovarian cancer cells is reduced.3.After the CENP-O gene is knocked down,the apoptosis of ovarian cancer cells increases.4.After CENP-O gene knockdown,the protein expression of CENP-O gene in ovarian cancer cells decreases.5.CENP-O has the potential to target molecular therapy.Down-regulating the expression of CENP-O gene can break the infinite proliferation ability of cancer cells and promote their apoptosis,which provides a basis and new ideas for subsequent molecular mechanism research and targeted therapy.