Effects Of SETD8 Gene On Cellular Biology And Chemotherapy Sensitivity Of Breast Cancer Cells

Objective: To construct a recombinant lentivirus vector SETD8 to investigate the effects of SETD8 on apoptosis,cell cycle and oxidative stress of breast cancer cells,and to observe the effects of SETD8 on the sensitivity of breast cancer cells to chemotherapy drugs.Methods: MCF-7 cell lines with stable overexpression of SETD8 and MDA-MB-231 cell lines with stable underexpression of SETD8 were established by lentivirus-mediated method.The infection efficiency was observed under fluorescence microscope.The silencing and overexpression efficiency of stable cell lines were verified at the transcription and protein levels by quantitative PCR and Western Blot(WB).The changes of cell apoptosis,cell cycle and intracellular reactive oxygen species were detected by flow cytometry.The correlation between SETD8 and cyclins was analyzed by refering database,and the changes of Cyclin B1 and P21 protein contents were detected by WB.The commonly used chemotherapy drugs ADM,DOCE and 5-FU were used to treat constantly SETD8-knock-down cells.Then the sensitivity of the stable cells to different chemotherapy drugs was verified by MTT method.Finally,flow cytometry was used to observe the changes of stable cell cycle and apoptosis after 48 treatment with docetaxel.Results: MCF-7 with overexpression of SETD8 and MDA-MB-231 with knockdown of SETD8 were successfully constructed,and the efficiency of SETD8 overexpression and knockdown of MDA-MB-231 was verified by PCR and WB(P<0.01).The apoptosis of breast cancer cells with low expression of SETD8 increased and the cycle stagnated at G2/M and S stages,and the intracellular ROS level was higher than that of the control group.The cell viability of SETD8 knockdown cell lines treated with docetaxel at different concentrations was significantly lower than that of the control group(P<0.01).Conclusion: lentivirus-mediated downregulation of SETD8 expression leads to increased apoptosis of breast cancer cells,cycle arrest in G2/M phase,increased intracellular ROS,synergistic effect with docetaxel,and increased drug sensitivity.