β-elemene Inhibits The Proliferation,Migration,Invasion And Epithelial-to-mesenchymal Transition Of Cervical Cancer Cells Through Mediating β-catenin /TCF7/Sox2 Signaling Pathway

IntroductionCervical cancer is the world’s 4th most common cancer among women,accounting for 4th of all cancer deaths among women.The most common causes of cervical cancer are invasion and distant metastasis.The invasion and metastasis of cervical cancer is a complicated and multifactorial process.At present,the mechanism of its development is not completely clear.In the process of tumor progression,many kinds of tumor cells show the change of plasticity through morphologic and phenotypic changes,among which epithelial stromal transformation is a highly conserved process,which occurs in embryogenesis.Chronic inflammation and fibrosis,as well as the progression of cancer,are now widely regarded as an important marker of the progression of malignant tumors,in breast,liver cancer,lung cancer and colon cancer play an important role in the progression.EMT is characterized by the loss of epithelial properties and the increase of gene expression in mesenchymal cells.The down-regulation or even deletion of E-cadherin is an important component of adhesion junctions by binding to its extracellular domain.In the extracellular matrix,E-cadherin binds to β-catenin,α-catenin and p120-catenin,which makes the cell contact between β-catenin and α-catenin p120-catenin stable.The latter plays a key role in extracellular signal transduction and adhesion junctions to actin cytoskeleton.Radiotherapy and chemotherapy,as the main adjuvant treatment for recurrent and metastatic cervical cancer,did not significantly improve the prognosis.It also increases the systemic side effects of chemotherapy and the occurrence of chemotherapeutic resistance.Therefore,a new adjuvant drug is urgently needed to treat uncontrolled and metastatic cervical cancer.At present,a large number of studies have confirmed that β-elemene has anticancer sibling in many kinds of malignant tumors,such as breast cancer.Ovarian cancer,liver cancer,pancreatic cancer,However,the mechanism of β-elemene in cervical cancer is not clear,and the mechanism of β-elemene in cervical cancer is not clear.In this study,we first studied the proliferation and cycle of β-elemene on cervical cancer cells.Secondly,in order to study the effect of β-elemene on emt,we established a cervical cancer emt model by TGF-β1 stimulation of cervical cancer cells.To investigate the effect of β-elemene on emt of cervical cancer cells induced by TGF-β1.The wnt/β-catenin signaling pathway consists of proteins encoded by oncogenes and anti-oncogenes,and plays an important role in embryonic development,intracellular transport and apoptosis.In addition,the abnormal activation of this signal transduction pathway is associated with the occurrence of many cancers including invasion and metastasis.β-catenin itself cannot directly bind to DNA during the activation of the wnt-/ β-catenin signaling pathway,so it must interact with dna.The binding protein TCF/ LEF activates downstream target genes and promotes cell proliferation and invasion and metastasis.Studies have shown that the migration and invasiveness of sox 2 overexpression cells in laryngeal carcinoma is enhanced,which is closely related to the activation of β-catenin.In this study,we examined the influence ofβ-elemene on the proteins of Wnt/β-catenin signaling pathway.To provide theoretical basis for the future study of β-elemene in the treatment of cervical cancer.MethodsThe first part: The effect of β-elemene on the proliferation of cervical cancer cells was detected by MTT.Flow cytometry was used to detect the effect of β-elemene on cell cycle and apoptosis;Cell scratch test and transwell assay were used to detect the effect of β-elemene on the migration and invasion of cervical cancer cells.Western blot was used to detect the changes of the protein of Wnt/β-catenin after β-elemene treatment.The second part:Observe the morphological changes of the cells stimulated by TGF-β1 by microscope,detect the changes of emt related proteins by western blot,detect the effect of TGF-β1 on the proliferation ability of cervical cancer cells by mtt.The effect ofTGF-β1 on migration and invasion of cervical cancer cells was studied by scratch test and transwell test.The third part: The effects of β-elemene and TGF-β1 on the migration and invasion of cervical cancer cells were detected by transwell assay,and the changes of WNT/β-catenin and sox2 related proteins were detected by western blot.The expression and correlation of β-catenin,TCF7 and sox2 in cervical carcinoma tissues with different pathological grades were detected by immunohistochemistry and The effect of β-elemene on β-catenin in the nucleus induced by TGF-β1 and the effect of β-elemene on β-catenin and sox2 binding were detected by western blot and immunoprecipitation.ResultsThe first part:β-Elemene inhibited the proliferation of cervical cancer cells.β-elemene induced cell cycle arrest of cervical cells in G1 phase in a time-and dose-dependent manner.β-elemene could promote the apoptosis of cervical cancer siha cells and hela cells and increase the concentration of β-elemene.The number of apoptosis increased in a dose-dependent manner.β-elemene inhibited the migration and invasion of siha and hela cells in a dose-dependent manner and increased with the concentration of β-elemene.The ability of migration and invasion gradually decreased in a dose-dependent manner(P<0.05).β-elemene could decrease the protein expression of β –catenine,TCF7,Bcl-2,cyclin D1,c-myc,MMP-2 in a dose-dependent manner and increase the protein expression of p53.The second part: After treatment with TGF-β1 10 ng / ml,the cell morphology increased and prolonged,and the morphological changes were obvious after 4 days of fusiform.Compared with the control group,the cell length increased significantly,the expression of E-cadherin protein decreased significantly and the expression of vimentin protein increased.The proliferation of siha cells and hela cells could be promoted by the increase of TGF-β 1.In a time-and dose-dependent manner,TGF-β 1 could promote the migration and invasion of siha cells and hela cells in a dose-dependent manner.The third part: β-Elemene inhibits the migration and invasion induced by TGFβ-1 in SiHa and HeLa cells.β-Elemene inhibited the migration and invasion of cervical cancer cells induced by TCFβ-1 by inhibiting sox2 and β-catenin signal transduction.β-catenin was expressed in cell membrane,cytoplasm and nucleus.It was higher in poorly differentiated cervical cancer than in moderately and highly differentiated cervical carcinoma.TCF7 protein was mainly expressed in the nucleus of cervical cancer and was higher in poorly differentiated cervical carcinoma than in moderately and highly differentiated cervical carcinoma.The difference was significant in the nuclei of cervical carcinoma.The expression of β-elemene in poorly differentiated cervical carcinoma was significantly higher than that in moderately differentiated and highly differentiated cervical carcinoma,and the difference was statistically significant(P<0.05).The expression of β-catenin TCF 7 was positively correlated with the expression of β-catenin TCF7 in different degree of differentiation of cervical carcinoma,and β-elemene could reverse the expression of β-elemene.β-catenin induced by TGF-β1 was transferred into the nucleus,and β-elemene could inhibit β-catenin/TCF7/Sox2.Conclusions1.β-elemene inhibited the proliferation of cervical cancer cells by inhibiting wnt / β-catenin pathway,induced cell cycle arrest to G1,induced apoptosis,inhibited migration and invasion.2.TGF-β1 induced epithelial interstitial transformation of cervical cancer cells,and significantly promoted the proliferation of cervical cancer cells.3.β-elemene inhibits the emt of cervical cancer through β-catenin/TCF7/Sox2 pathway,which provides a new mechanism for preventing invasion and metastasis of cervical cancer and the treatment of cervical cancer.