The Study On The Effect And Mechanism Of Radiosensitivity Change Of MDA-MB-435 Cells And T-47D Cells Of Breast Cancer By Artemisinin

Objective:Radiation therapy is one of the important treatments for malignant tumors, but in the course of radiotherapy,ionizing radiation can also make normal tissue and cell damage,as well as the destruction of tumor cells at the same time.In order to improve efficacy to tumor and reduce injury to normal tissues and cells caused by radiotherapy, studies on tumor radio sensitization have been carried out.The radiosensitivity of tumor cells has the closest relationship to their p53 gene status.This study compares the different changes of radiosensitivity in p53 mutant human breast cancer MDA-MB-435 cells and wild-type p53 human breast cancer cells T-47D cells-two different cells of different p53 gene status treated with artemisinin.And try to find out the mechanism of the variations caused by artemisinin in p53 mutant cells.Methods:Use human breast cancer cells MDA-MB-435 and T-47D as the experimental cells.(1)Study the impact of artemisinin to cell survival of the two types of experimental cells by using MTT,in order to determine drug concentration and time; (2)Study the apoptosis of the two kinds of cells induced by artemisinin combined with ⁶⁰Co-γ-ray by using Annexin V/PI double staining method;(3)Study the effect to MDA-MB-435 cells caused by artemisinin combined with ⁶⁰Co-γby using cytokinesis block method(CB method) micronucleus method and conventional chromosome examination;(4)Study the changes of cell cycle distribution of two types of cells detected by flow cytometry;(5)Study the changes in expression of cyclin B1 using western blot analysis.The results:(1)MTT analysis showed that:survival of the two types of experimental cells treated with artemisinin reduced along with the increase of drug concentration;but overally toxicity of artemisinin to the two cells is low,and determine the concentration of artemisinin in MDA-MB-435 is 250μmol/L,in T-47D is 200μmol/L,acting time is 24h; (2)Annexin V/PI double staining showed:artemisinin had no significant impact to apoptosis of cells without exposure to radiation,artemisinin united with radiation can effectively increase apoptosis of p53 mutant MDA-MB-435 cells,the apoptosis rate in simple radiation cell group is 11.573±1.832%and in artemisinin united with irradiation group is 32.833±10.105%,there is significant difference(p＜0.05) between two groups; however,p53 wild-type T-47D cells are in the absence of such effects,in simple radiation cell group apoptosis rate is 46.393±3.871%and in artemisinin united with irradiation group apoptosis rate is 46.617±11.859%,and there wasn't significant differences(p＞0.05) between the two groups;(3)micronucleus and chromosome analysis showed that: chromosome damage of MDA-MB-435 cells treated with artemisinin united with irradiation of the same dose is more serious than treated only with radiation;(4)Flow cytometry analysis showed that:by 0,5,10Gy the MDA-MB-435 cells in artemisinin united with irradiation group,content of G2 phase cells have significant decrease compared to simple radiation cell group(p＜0.05),and decrease increase with the dose;and in the two experimental groups of T-47D cells,the contents of G2 phase cells had no significant differences(p＞0.05);(5)Western blot analysis showed artemisinin united with radiation could reduce expression of G2 phase protein cyclinB1 in p53 mutant cells MDA-MB-435, but the effect to p53 wild-type T-47D cells was not obvious.Conclusion:Artemisinin can increase the radiation-induced apoptosis rate,MNF, MNCF and chromosome aberration rate in p53 mutant breast cancer cells MDA-MB-435, which confirmed that after the treatment with artemisinin the radiosensitivity of the p53 mutant-type MDA-MB-435 cells will be increased.However,artemisinin can not change the radiation-induced apoptosis of p53 wild-type T-47D cells.MDA-MB-435 cells,treated with artemisinin and irradiation,the content of G2 phase cells is significantly lower than the cells treated only with irradiation,and in T-47D cells,the effect of artemisinin is not so significant.Artemisinin can reduce the MDA-MB-435 cells' expression level of cyclin B1 protein after radiation,and cannot change the T-47D cells' expression level of cyclin B1,it can be speculated that artemisinin can inhibit G2 phase cell block and the expression of cycling B1,and then increase the radio sensitization of p53 mutant MDA-MB-435 cell.