Study On The Effect Of Danzhi Xiaoyao Powder On Regulating Intestinal Flora And Its Mechanism Of Anti-breast Cance

Objective:In this study,the sequencing analysis of intestinal flora was used to investigate the inhibition of breast cancer development and its possible molecular mechanism,and to provide a reference for the basic research on the prevention and treatment of breast cancer by Danzhi Xiaoyao Powder.Materials and methods.1.To observe the effect of Danzhi Xiaoyao Powder on the intestinal flora of human breast cancer MDA-MB-231 cells in tumor-bearing nude miceForty-four SPF 6-week-old BALB/c-nu female nude mice were fed for 7 days.The tumour model was established by inoculating a stable transfer of MDA-MB-231-GFP cell suspension（density 5×10⁵/m L）under the fat pad of the nude mammary cavity at logarithmic growth stage.The mice were divided into normal control group,model group,Danzhi Xiaoyao Powder group(11.69g-kg^-1)and Xi huang wan group(0.78g-kg^-1)by random grouping method,11 rats in each group.After 1 day of inoculation,breast cancer cells were gavaged once a day at a fixed time for 22 days.Equal amounts of saline were given to the normal control group and the model group for gavage.After 2 hours of the final administration,the samples were removed by execution and the intestinal contents and faeces were collected and sequenced for microbial diversity amplicons.The differential flora were screened by Alpha diversity analysis,Beta diversity analysis,and relative abundance bar cumulative plot analysis of intestinal microorganisms,and the flora diversity,abundance and structure were analysed.2.Observation of the effect of Danzhi Xiaoyao Powder-containing serum on human breast cancer MCF-7 cellsHigh and low doses of Danzhi Xiaoyao Powder(17.98,8.99 g-kg^-1)were prepared and the blank serum was prepared.Danzhi Xiaoyao Powder-containing serum was used to interfere with human breast cancer MCF-7 cells.The expression of inflammatory factors IL-10 and IL-23 in MCF-7 cells was detected by ELISA and the expression of STAT3,P-STAT3,JAK2,P-JAK2 and IL-6 in MCF-7 cells was detected by Western blot.Results.1.Effect of Danzhi Xiaoyao Powder on the intestinal flora of tumour-bearing nude mice with human breast cancer MDA-MB-231 cellsDanzhi Xiaoyao Powder restored the species richness and uniformity of the samples to some extent.It restored the altered condition of intestinal flora,which had an ameliorating effect on the imbalance of intestinal microecology,and restored the composition and structure of intestinal flora to some extent.Compared with the model group,At the phylum level,the Danzhi Xiaoyao Powder group could elevate the relative abundance of Bacteroidetes and decrease the relative abundance of Cyanobacteria.At the phylum level,the relative abundance of Bacteroidetes was increased and that of Oxyphotobacteria was decreased.At the order level the relative abundance of Bacteroidales can be raised and that of Chloroplast lowered.The relative abundance of Muribaculaceae can be increased at the family level.The relative abundance of Prevotellaceae＿UCG-001 can be increased and that of Lachnospiraceae＿UCG-006 can be decreased at the genus level.It is suggested that Danzhi Xiaoyao Powder may exert its anti-breast cancer effect through regulating intestinal flora.2.Effects of Danzhi Xiaoyao Powder-containing serum on human breast cancer MCF-7 cellsIn the scratch healing assay,the healing rate of the high and low doses of Danzhi Xiaoyao Powder-containing serum group was significantly lower than that of the blank serum group.In comparison with the blank serum group,the expression of inflammatory protein IL-10 and IL-23 was significantly down-regulated in the Danzhi Xiaoyao Powder(17.98,8.99g-kg^-1)serum group was detected by ELISA.In comparison with the blank serum group,the expression of inflammatory protein IL-6 was significantly down-regulated in the Danzhi Xiaoyao Powder(17.98,8.99g-kg^-1)serum group,and the expression of P-JAK2/JAK2,P-STAT3/STAT3 protein expression were significantly reduced was detected by Western blot.Conclusion.1.Danzhi Xiaoyao Powder may exert its anti-tumor effects through the regulation of intestinal flora in human breast cancer cell-bearing nude mice to influence the inflammatory response pathway.2.Danzhi Xiaoyao Powder-containing serum significantly altered the inflammatory changes in human breast cancer MCF-7 cells,probably through the regulation of the JAK2/STAT3signaling pathway.