Study On Apoptosis Mechanism Of Human Colorectal Cancer SW480 Cell Induced By Isobavachalcone

Objectives: Colorectal cancer(CRC)is one of the most common malignancies in the human digestive tract.It is also the third largest malignancy and the leading cause of cancer death in men and women worldwide.Even today,its prevalence remains high,mostly among people between the ages of 40 and 50,with a male to female ratio of 2:1 to 3:1.Due to the characteristics of slow onset,high metastasis rate and poor prognosis of colorectal cancer,it is important to study the pathogenesis of colorectal cancer for early diagnosis and individualized treatment.Effective method for treating colorectal cancer at present includes surgery,radiotherapy and chemotherapy.A series of targeted therapies have been developed to treat cancer,but serious side effects and drug resistance have limited their application.The effect of the treatment of patients with colorectal cancer is still poor.Therefor it is of great importance to develop new drugs.Isobavachalcone is a kind of ketone compoundsisolated from herbs malaytea scurfpea fruit.Studies have shown that it has a wide range of pharmacological and biological activities,including antifungal,antibacterial and anticancer and parasitic resistance,etc.Isobavachalconecan induce the apoptosis of many kinds of tumor cells at the same time,however,the anticancer activity and potential molecular mechanismof isobavachalcone in colorectal canceris still unclear.In this study,human colorectal cancer SW480 cells were taken as the research material to preliminarily investigate the mechanism of isobavachalcone inducing apoptosis of human colorectal cancer SW480 cells,provide theoretical and experimental basis for the application of isobavachalcone as a drug for the treatment of colorectal cancer.Methods: After stimulating human colorectal cancer SW480 cells with different concentrations of isobavachalcone,the effect of isobavachalcone on cell proliferation was determined by CCK8 test.Cell cycle arrest,intracellular ROS levels and apoptosis were detected by flow cytometry with PI single staining,DCFH-DA probe staining and Annexin V-FITC/PI double staining.Western blotting(WB)was used to detect changes in cell cycle,apoptosis and expression of autophagy-related proteins after stimulation with different concentrations of drugs,SW480 cells were pretreated with reactive oxygen scavenger NAC and autophagy inhibitor 3-MA and then stimulated with different concentrations drugs.The effects of reactive oxygen on cell proliferation,autophagy and AKT/ GSK-3? pathway were detected by CCK8 assay and WB.Results: 1.The results of CCK8 test showed that the cell activity significantly decreased with the increase of isobavachalcone concentration,and it was concentration-dependent.2.Flow cytometry results showed that isobavachalcone could arrest SW480 cell cycle in G0/G1 phase and increase the proportion of cells in G0/G1 phase in a gradient dependent manner.WB results showed that isobavachalcone decreased the expression of Cyclin D1 protein in SW480 cells,while the expression of p21 protein was up-regulated.3.Flow cytometry results showed that isobavachalcone induced apoptosis of SW480 cells and promoted the increase of intracellular ROS production.WB results showed that Cleaved PARP expression was up-regulated in SW480 cells after isobavachalcone stimulation.4.WB results show isobavachalcone significantly raised SW480 cells LC3B-?,p-AKT,p-GSK-3? protein expression,at the same time,the protein expression of three decreased because of NAC pretreatment.In addition,In addition,3-MA pretreatment can significantly reduce the inhibitory effect of isobavachalconeon SW480 cell proliferation.Conclusion: The above experimental results indicated that isobavachalcone had a significant inhibitory effect on the proliferation of SW480 cells and arrested the cell cycle in the G0/G1 phase.ROS mediated activation of autophagy and inhibition of AKT phosphorylation,which further inhibitedphosphorylation of GSK-3? and activated its activity,finally induced cell apoptosis.Therefore,isobavachalcone is a potential anti-tumor drug,and the use of isobavachalcone in the treatment of colorectal cancer may be a promising adjuvant chemotherapy.