Effect of Shugan Yishen Prescription on Tamoxifen-resistant Cell Line from Exosome-mediated Crosstalk in Breast Cancer Microenvironment / 中国实验方剂学杂志

ABSTRACT

Objective:To explore the effect of Shugan Yishen prescription（SGYS） on the tamoxifen （TAM） -resistant cell line LCC9 by the intervention of exosome-mediated crosstalk in the breast cancer microenvironment. Method:Four groups of serum were set up， specifically， a blank group， a TAM （2 mg·kg^-1·d^-1） group，an SGYS（113.2 g·kg^-1·d^-1） group，and a combination group. The exosomes of LCC9 cells were extracted by ultracentrifugation and identified by transmission electron microscopy （TEM），nanoparticle tracking analysis （NTA）， and Western blot. Then the collected LCC9 exosomes （LCC9-EXO） were co-cultured with bone marrow mesenchymal stem cells（BMMSCs），and 10% of the above four groups of serum were added to the co-culture system. After 48 hours of co-culture，the exosomes of BMMSCs （BMMSCs-EXO） were extracted and incubated with LCC9 cells. Fluorescence microscope was used to observe the uptake of exosomes by cells. Cell counting kit-8 （CCK-8） assay，flow cytometry， and Transwell assay were used to detect the effects of drug-containing serum in the four groups on the proliferation，apoptosis， and migration of LCC9 cells. Western blot was used to detect the protein expression of CD24，CD44，human epidermal growth factor 2（HER2）， and estrogen receptor <italic>α</italic> （ER<italic>α</italic>） in each group. Result:Fluorescence microscope observed that LCC9-EXO could be taken up by BMMSCs，and BMMSCs-EXO could be taken up by LCC9 cells. CCK-8 assay revealed that compared with the TAM group，the SGYS group and the combination group showed reduced cell proliferation ability at each period （<italic>P</italic><0.05），especially the combination group，but no statistically significant difference between the SGYS group and the combination group was observed （<italic>P</italic><0.05）. Flow cytometry revealed that compared with the TAM group，the SGYS group and the combination group showed increased levels of apoptosis （<italic>P</italic><0.05）. Transwell assay revealed that compared with the TAM group，the SGYS group and the combination group showed decreased cell migration ability （<italic>P</italic><0.05）. Western blot revealed that compared with the TAM group，the SGYS group and the combination group showed up-regulated expression of ERα and CD24（<italic>P</italic><0.05），and down-regulated expression of HER2 and CD44 （<italic>P</italic><0.05）. The effect of the combination group on protein expression was superior to that of the SGYS group （<italic>P</italic><0.05）. Conclusion:SGYS reverses the TAM resistance of LCC9 cells by interfering with the crosstalk between BMMSCs-EXO and LCC9-EXO.