Study Of The Effect And Mechanism Of Activin A On Regulation Of Migration Of Human Breast Cancer Cells

Breast cancer is the most common malignant tumor in women.It ranks first among women with new cancers in the world.It is also the most common malignant tumor among women due to cancer deaths,which seriously threatens women’s health and life.Activin A belongs to a member of the transforming growth factor beta(TGF-β)superfamily and is involved in the regulation of various cell differentiation development,activation,and biological behavior.Studies have shown that activin A is expressed in a variety of tumor tissues,such as lung cancer,liver cancer,pancreatic cancer,metastatic prostate cancer and rectal cancer,and activation of activin A signaling pathway promotes invasion of tumor cells such as esophageal squamous cell carcinoma.Studies have also shown that serum activin A levels in breast cancer patients increased,the previous study of our group found that activin A can promote the migration of human breast cancer cell line MDA-MB-231 cells.The spread and metastasis of cancer cells is an important cause of death in cancer patients,and tumor metastasis is closely related to tumor cell migration ability and epithelial-mesenchymal transition(EMT)process.MDA-MB-231 cells act as estrogen receptor-negative(ER-)breast cancer cells,and there are many biological behaviors in estrogen receptor-positive(ER+)breast cancer cells.However,the effect of activin A on the migration and proliferation of different types of breast cancer cells,as well as the signaling mechanism of action,has not yet been elucidated.Therefore,this study selected MCF-7(ER+)and MDA-MB-231(ER-)breast cancer cell lines to investigate the role of activin A in the regulation of migration and proliferation of two breast cancer cells and its related signaling mechanisms.1.METHODS(1)Microfluidic chip technology was used to detect breast cancer cell migration: using a three-channel microfluidic chip,the migration of cells under the action of activin A was recorded using a microscope imaging system at 37 °C.(2)RTCA method for detecting breast cancer cell proliferation and adhesion: Real-Time Cell-based Assay(RTCA)is a real-time cell biosensor that performs uninterrupted,label-free real-time analysis of cells during the experiment.When the cells are connected or separated from the surface electrode,the impedance changes,and the change in the electronic reading occurs,and the cell index(CI)value is plotted by computer processing.The higher the number of cells,the higher the CI value.Therefore,the number of cells in the CI range is related to the adhesion quality,which in turn reflects cell proliferation and adhesion.(3)Western blotting detect the expression of related signal pathway proteins: Western blot was used to detect the levels of N-cadherin,E-cadherin and Vimentin,and Smad3,ERK and phosphorylated ERK(p-ERK),Akt and phosphorylated Akt(p-Akt).2.RESULTS(1)Autocrine activin A in breast cancer cells: The results showed that MDA-MB-231 and MCF-7 cells secrete activin A,but MCF-7 cells secrete activin A levels significantly lower than MDA-MB-231 cells.(2)EGFR expression in breast cancer cells: The results showed that the expression of EGFR m RNA in MDA-MB-231 cells was significantly higher than that in MCF-7 cells,suggesting that the biological characteristics of the two cells are different.(3)Effect of activin A on proliferation of breast cancer cells: The results showed that activin A can promote the proliferation of MDA-MB-231 cells and inhibit the proliferation of MCF-7 cells.At the same time,activin A inhibits the apoptosis of MDA-MB-231 cells,and promoted apoptosis of MCF-7 cells.(4)Effect of activin A on migration and adhesion of breast cancer cells: The results showed that activin A promoted migration and adhesion of MDA-MB-231 cells and accelerated wound healing;while inhibiting migration of MCF-7 cells and slowing the healing of scars,and there is no significant effect on its adhesion.Activin A also promotes the expression of EMT intermediate markers N-cadherin and Vimentin and the expression of MMP9 in MDA-MB-231 cells.It is suggested that activin A may affect EMT by affecting the microenvironment of breast cancer cells,promoting infiltration and affecting its migration and adhesion.(5)Activin signaling protein expression: The results showed that activin A can promote Act RIIA expression in MDA-MB-231 cells,but has no significant effect on Act RIIB and Smad3 expression.Further studies showed that activin A increased ERK and p-Akt protein levels in MDA-MB-231 cells,whereas p-ERK and p-Akt protein levels decreased in MCF-7 cells,suggesting that activin A may pass ERK /MAPK and PI3K/AKT-related signaling pathways regulate breast cancer cell migration and proliferation.3.CONCLUSIONIn summary,the biological characteristics of MDA-MB-231 and MCF-7 cells are different.Activin A has different effects on the migration and proliferation of two breast cancer cells.The regulation of breast cancer cells may be independent of Smad signaling.However,the ERK/MAPK and PI3K/AKT signaling pathways of Smad-independent pathway regulate the proliferation and migration of MDA-MB-231 and MCF-7 cells.Therefore,this study not only described activin A regulation of migration and proliferation of two breast cancer cells and its related signaling mechanisms,but also further revealed that activin A may be an important factor affecting the development of breast cancer.