| Objective To explore the effects of matrine and docetaxel on cell proliferation and apoptosis in human breast cancer MDA-MB-231 cells.On this basis,the further molecular mechanism of matrine combined with docetaxel against human breast cancer MDA-MB-231 cells was studied by proteomics and bioinformatics technology,and verified in vitro and in vivo experiments.Methods 1.To explore the effects of different concentrations of matrine and docetaxel on the proliferation of human breast cancer MDA-MB-231 cells by CCK-8,clone formation,transwell,scratch test and flow cytometry assays.On this basis,matrine(1 mM),docetaxel(10 nM)and combination therapy(matrine 1 mM+docetaxel 10 nM)were selected for the clonal formation,invasion,migration and pro-apoptotic effects of human breast cancer MDA-MB-231 cells.Western blotting was used to detect the effects of the above-mentioned medication modes on the expression of Caspase and Bcl-2 family-related proteins in human breast cancer MDA-MB-231 cells.2.The target molecules of matrine combined with docetaxel against human breast cancer MDA-MB-231 cells were studied by proteomics and bioinformatics technology,and verified by relevant experiments.After the molecule was overexpressed by transfection plasmid,the phenotype and Caspase family related proteins of human breast cancer MDA-MB-231 cells were detected by the treatment of combination therapy.3.The BALB/c nude mice model of human breast cancer MDA-MB-231 cells was established.When the tumor volume reached about 100 mm3,the nude mice were randomly divided into the following 4 groups,with 8 mice in each group,and were treated as follows:①Control:An equal volume of normal saline was injected intraperitoneally,5 times/week,for a total of 15 times.②Mat:Intraperitoneal injection of matrine 50mg/kg,5 times/week,a total of 15 times.③Doc:Intraperitoneal injection of docetaxel 8 mg/kg,1 time/week,3 times in total.④ Comb:Matrine+Docetaxel(intraperitoneal injection of matrine 50mg/kg,5 times/week,a total of 15 times;intraperitoneal injection of docetaxel 8mg/kg,once/week,a total of 3 times).During the study,the general conditions of the tumor-bearing nude mice in each group were observed,and the body weight and tumor volume were measured.After meeting the experimental requirements,blood was collected,and the nude mice were sacrificed by de-neck.HE staining,Tunel staining and Immunohistochemical detection of related proteins were performed on tumor tissues respectively.Results 1.Matrine and docetaxel inhibited the proliferation,clone formation,invasion,migration and apoptosis of human breast cancer MDA-MB-231 cells in a concentration-and time-dependent manner,and the combination of the two drugs had a synergistic effect.WB assays confirmed that the expression of pro-apoptotic proteins such as Cleaved-caspase-3,-8,-9 and Bax were all up-regulated after matrine and docetaxel single-drug treatment on human breast cancer MDA-MB-231 cells,the expression of anti-apoptotic protein Bcl-2 decreased,and the combination of the two drugs had a synergistic effect.2.Proteomic analysis showed that HN1 was one of the down-regulated proteins in human breast cancer MDA-MB-231 cells treated with matrine,and the proliferation,clone formation,invasion,migration and apoptosis of MDA-MB-231 cells induced by matrine combined with docetaxel could be counteracted by overexpression of HN1.3.The combined administration of matrine and docetaxel had a synergistic effect on the inhibition of tumor volume and tumor weight of human breast cancer MDA-MB-231 cells transplanted nude mice.Immunohistochemistry suggested that the combined administration mode had a synergistic effect in inhibiting the expression of CD31,VEGF,Cleaved-caspase-3 and HN1 in tumor tissue,and this medication model had no significant effect on the general condition,body weight,liver function and blood routine of nude mice.Conclusion The anti-tumor effect of matrine(1mM)combined with docetaxel(10nM)on human breast cancer MDA-MB-231 cells was synergistic,which may be achieved by down-regulating the expression of HN1.The combined administration of matrine and docetaxel had a synergistic effect on the inhibition of tumor volume and tumor weight of human breast cancer MDA-MB-231 cells transplanted nude mice,and had no significant effect on the general condition,body weight,liver function and blood routine of nude mice. |
