Effects Of Entinostat On Proliferation,Migration,Invasion And Apoptosis Of Osteosarcoma Cells And Its Mechanism

Research objectiveTo observe the effect of histone deacetylase inhibitors Entinostat on the proliferation,colony formation,migration,invasion and apoptosis in osteosarcoma MG63 cells,and to further explore whether Entinostat exerts its anti-osteosarcoma effect through the Wnt/β-catenin signaling pathway.Research methodsOsteosarcoma is a malignant tumor that seriously threatens the health of children and adolescents.Osteosarcoma has high malignancy and easy early metastasis,and the prognosis is very poor.Entinostat is a promising class I histone deacetylase inhibitor,which has good curative effect on many malignant tumors.However,there are few studies on the treatment of osteosarcoma with Entinostat.In order to study the effect of entinostat on the proliferation,migration,invasion and apoptosis of osteosarcoma cells and its mechanism,the following assays were carried out in this study.The osteosarcoma cell line MG63 cells were purchased for in vitro culture,and the log phase cells were used for assays.According to the needs of the assays,The groups of assay include the control group and the experimental groups with different concentrations of Entinostat.The control group was only given 0.1% DMSO for 24 to72 hours,and the experimental groups were given different concentrations of Entinostat for 24 to 72 hours,and then Cell Proliferation assay(CCK-8)were performed to observe the effects of different Entinostat concentrations and different intervention time on the proliferation of MG63 cells.According to the results of the Cell proliferation experiment(CCK-8),the appropriate Entinostat’s concentration and intervention time were selected for follow-up assays.Follow-up assays include Cell morphology observation,Colony formation assay,Wound healing assay,Transwell invasion assay,Apoptosis analysis by flow cytometry,QRT-PCR and Western blotting.The next step was to do Cell morphology observation,Colony formation assay,Wound healing assay,Transwell invasion assay and Apoptosis analysis by flow cytometry.Finally,mRNA and protein were extracted for QRT-PCR and Western blotting to observe the mRNA and protein expression of β-catenin,E-cadherin and Caspase9.Research result1.The results of Cell proliferation assay(CCK-8)showed that Entinostat has a significant proliferation inhibitory effect on MG63 cells,and it is positively correlated with drug concentration and intervention time within a certain range;2.The observation of cell morphology showed that Entinostat could shrink and elongate the cytoplasm of MG63 cells,and also decreased the cell adhesion ability.3.The results of Colony formation assay showed that Entinostat can significantly reduce the in vitro proliferation and colony formation ability of MG63 cells;4.Wound healing assay and Transwell invasion assay showed that Entinostat can also significantly reduce the migration and invasion ability of MG63 cells;5.Result of Apoptosis analysis by flow cytometry showed that Entinostat significantly promoted the apoptosis of MG63 cells;6.Entinostat can promote the mRNA and protein expression of MG63 cell apoptosis gene(Caspase9)and tumor metastasis related genes(E-cadherin);Entinostat can also inhibit the mRNA and protein expression of β-catenin,which can inhibit the Wnt/β-catenin signaling pathway.Conclusion1.Entinostat can significantly reduce the proliferation,colony formation,migration and invasion ability of osteosarcoma MG63 cells and promote the apoptosis of osteosarcoma MG63 cells.2.Entinostat can inhibit the Wnt/β-catenin signaling pathway by inhibiting the mRNA and protein expression of β-catenin,thereby exerting an anti-osteosarcoma effect.3.As a histone deacetylase inhibitor,Entinostat is a kind of potentially effective targeted drug for the treatment of osteosarcoma,which is of great significance for the clinical treatment of osteosarcoma.