Inhibitory Effect Of Sijunzi Decoction Extract On The Growth Of Human Triple Negative Breast Cancer MDA-MB-468 Cells

Background and ObjectiveBreast cancer is the most common tumor in women and the second leading cause of cancer death in women.Breast cancer is a disease that affects many women worldwide.In Asian countries,the incidence of breast cancer has been increasing over the past few decades.Surveys in most cities in China show that breast cancer incidence and mortality rank first.Triple negative breast cancer(TNBC)is a special type of breast cancer that is difficult to treat.It has estrogen receptor(ER),progesterone receptor(PR)and human epidermal growth factor receptor(HER2)immunohistochemical markers.Negative characteristics.Compared with other breast cancer types,TNBC is more malignant and more aggressive.Single treatment and lack of specific targeted drugs are currently the biggest obstacles to clinical treatment of this disease.The key to the treatment of TNBC by traditional Chinese medicine is Yiqi Jianpi,a classic prescription of Yiqi Jianpi,which is composed of ginseng,Baizhu,Poria,and licorice.It has been initially proved that Sijunzi Decoction can significantly improve breast cancer chemotherapy and postoperative complications,significantly improve the quality of life of breast cancer patients,and can regulate the immune system function of breast cancer patients.However,the mechanism of Sijunzi Decoction on tumor is not clear.Therefore,we used the method of extracting Sijunzitang concentrated dry powder and adding cells to investigate the effects of Sijunzitang on the proliferation and apoptosis of triple-negative breast cancer cells through cell proliferation experiments,clone formation experiments,Hoechst33342 staining technology and flow cytometry.Methods1.The dry powder of Sijunzi Decoction was extracted and quality control was performed by detecting the content of glycyrrhizic acid.Filter and sterilize before use.2.CCK-8 was used to test the effects of Sijunzi Decoction extracts with different concentrations on the inhibition rate and proliferation activity of breast cancer cells MDA-MB-468.3.The effects of Sijunzi decoction extracts on colony forming ability of breast cancer cell MDA-MB-468 cells were tested by clone formation experiments.4.Effects of Sijunzi Decoction Extracts on MDA-MB-468 Migration in Breast Cancer Cells.5.Hoechst33342 staining technique was used to observe the effect of Sijunzi Decoction extract on the apoptosis of breast cancer cell MDA-MB-468 by fluorescence microscope.6.Flow cytometry was used to detect the effects of Sijunzi Decoction extract on the apoptosis and cycle of human breast cancer cells MDA-MB-468.7.Western Blot to detect STAT3 protein expression in MDA-MB-468 cells.8.SPSS 25.0 software was used for statistical analysis of the data.The experimental data were expressed as,and the differences between the groups were analyzed by one-way ANOVA.P<0.05 indicated that the differences were significant.Results1.Sijunzitang can inhibit the proliferation of MDA-MB-468 cells with IC50 of 9.745 mg/mL.According to the IC50,the low,medium,and high Sijunzitang concentrations in the subsequent experiments were 2.5,5.0,and 10.0 mg/mL,respectively.The CCK-8 method was used to detect the effects of different concentrations of Sijunzitang on the proliferation of MDA-MB-468 cells at different times.It was found that the inhibitory effect of Sijunzitang on the proliferation of MDA-MB-468 cells was time and mass concentration dependent.2.Compared with the control group,the size and number of cell clones were significantly reduced after Sijunzitang was applied to MDA-MB-468 cells,and the cells in the high-dose Sijunzitang group could hardly form visible clones.3.Compared with the control group,the scratch healing ability of the cells in the Sijunzitang 5,10 mg/mL group was significantly reduced 48 h after the scratch(P<0.05,0.001).It shows that Sijunzitang can inhibit the migration of MDA-MB-468 cells,and it is mass concentration dependent.4.Hoechst 33342 staining results showed that the apoptotic morphology of Sijunzitang groups was obvious.5.Flow cytometry apoptosis test results showed that after dealing with Sijunzitang,the early apoptosis rate of MDA-MB-468 cells gradually increased,which was significantly different from the control group.And the total cell apoptosis rate gradually increased,showing a mass concentration dependence.6.The flow cytometry cycle test results showed that compared with the control group,the G2 phase cells in the Sijunzitang 5,10 mg/mL group were significantly reduced,and there was a mass concentration dependence(P<0.01).7.Western Blot showed that histone expression at Sijunzitang 5,10 mg/mL was significantly reduced,compared with the control group.Conclusion1.Sijunzitang decoction can inhibit the proliferation and colony formation of breast cancer cell MDA-MB-468.2.Sijunzitang decoction can promote the apoptosis of breast cancer cells MDA-MB-468,and reduce G2 cells.3.The inhibitory effect of Sijunzitang decoction on breast cancer cell MDA-MB-468 may be related to the STAT3 pathway.