Effect Of Silencing COPB2 Gene On Proliferation,Migration And Invasion Of Human Triple-negative Breast Cancer Cells

ObjectiveTriple-negative breast cancer has strong invasiveness,high lymph node metastasis rate,high recurrence rate in five years,poor prognosis and lack of therapeutic targets.In recent years,silencing the expression of tumor-specific genes and developing new drugs that against tumor-specific genes has become a new direction of tumor gene therapy.COPB2 plays an important role in intracellular transport.Previous study had found that COPB2 gene was highly expressed in cancer tissues,and COPB2 gene silencing could cause cell cycle arrest,inhibition of cell proliferation and induction of apoptosis.In this study,the changes of proliferation,migration and invasion after COPB2 silencing were investigated in triple-negative breast cancer cell lines,and the mechanism of action after COPB2 silencing will be explored.Method1.Western blotting was used to quantify the expression of COPB2 and the expression of E-cadherin,vimentin and N-cadherin in MCF-7 non-triple-negative breast cancer cells and HS-578 T triple-negative breast cancer cells.2.MTT assay,cloning formation assay and transwell assay were used to detect the differences of proliferation,migration and invasion between HS-578 T and MCF-7.3.Lentiviral interference vector of COPB2 gene sh-COPB2 was used to transfect HS-578 T cells as silencing group.After transfection for 96 hours,the silencing efficiency was determined by Western blotting.4.MTT assay,cloning formation assay and transwell assay were used to detect cell proliferation,migration,invasion after COPB2 silencing in HS-578 T.5.Western blotting was used to detect the expression of E-cadherin ?N-cadherin?vimentin and p-AKT in HS-578 T after silencing.6.After addtion of AKT agonist SC79,the changes of p-AKT and COPB2 in HS-578 T were detected by western blotting and the changes of cell migration andinvasion were detected by transwell assay.Result1.The level of COPB2 expression in HS-578 T cells was higher than that in MCF-7 cells(P <0.05).2.HS-578 T cells have stronger abilities of proliferation,migration and invasion than MCF-7 cells.The level of E-cadherin expression in MCF-7 cells was higher than that in HS-578 T cells,while the levels of N-cadherin and vimentin expression in HS-578 T cells were higher than those in MCF-7 cells(P <0.05).3.After HS-578 T cells were transfected with sh-COPB2,the silencing efficiency was more than 80%.After COPB2 silencing,the proliferation,migration and invasion of HS-578 T cells were reduced compared with the control group(P<0.05).4.After silencing COPB2,there was no significant difference in E-cadherin expression compared with control group(P>0.05).The expression of N-cadherin,vimentin,and p-AKT in HS-578 T cells was significantly down regulated compared with control group(P <0.05).5.After addtion of AKT agonist SC79,migration and invasion were upregulated compared with silencing group(P <0.01).6.After addtion of AKT agonist SC79,there was no significant difference in COPB2 expression compared with silencing group.ConclusionThe COPB2 expression level of HS-578 T cells was significantly higher than MCF-7 cells,and the abilities of proliferation,invasion and migration of HS-578 T cells were stronger than MCF-7 cells.COPB2 silencing could inhibit the proliferation,migration and invasion of HS-578 T cells.These findings suggested that COPB2 may be involved in the development of triple negative breast cancer,and its mechanism is related to the upregulation of p-AKT.