| Objective: Breast cancer has always been one of the most important causes of female mortality in recent years.Scientists from all over the world have been working on the treatment mechanism of breast cancer.Studies have shown that obesity is one of the main factors that cause breast cancer.Curcumin has been discovered as a kind of lipidlowering drug for many years,especially its potential unique anticancer properties have been widely concerned.Curcumin has been shown to inhibit the growth of a variety of cancer cells,including breast cancer.However,the molecular mechanism of curcumin in the treatment of breast cancer has not been fully elucidated,and lack of in vivo studies.The purpose of this study was to investigate the effect of curcumin on MCF-7breast cancer cells and its potential molecular mechanism in vitro and in vivo.Methods: In our study,human MCF-7 breast cancer cells were selected.In vivo experiment: The effects of curcumin on tumor growth were observed in animal models of breast cancer.Breast cancer animal model was established to observe the effect of curcumin on tumor growth,and the positive reactions of caspase-1,IL-1β and IL-18 were detected by immunohistochemical method.LC3,p62,CTSB,ASC,pro-Caspase-1,GSDMD,NLRP3,Caspase-1,GSDMD-N,IL-1β,IL-18 proteins expression from tumor were determined by western blot.In vitro experiment: MTT assay was used to determine the effect of curcumin on survival rate of MCF-7 cells.The expression of these proteins were measured by western blot.The release of mature IL-1β and IL-18 was measured by ELISA assay.LDH release was measured by LDH assay.The content of CTSB in cytoplasm were determined by immunofluorescence assay.Transwell assay,tube formation assay and plane cloning assay were used to measure cell function.In our study,NLRP3 inflammasome inhibitor MCC950,cathepsin B inhibitor CA-074 Me and autophagy inhibitor 3-MA were also used to act on cells to investigate the role of NLRP3 inflammasome,cathepsin B and autophagy in curcumin-induced pyrotosis of MCF-7 breast cancer cells.Results: Curcumin significantly reduced the survival rate of MCF-7 breast cancer cells.MTT method was used to screen out the treatment condition of 8μm curcumin treated for 48 hours.Curcumin could induce pyroptosis of MCF-7 breast cancer cells in vitro and vivo: The immunohistochemistry of pyroptosis related proteins Caspase-1,IL-1βand IL-18 in tumor tissue sections of curcumin treated were significantly positive compared with the control;Compared with the control group,the protein expression levels of LC3,CTSB,ASC,pro-Caspase-1,GSDMD,NLRP3,Caspase-1,GSDMD-N,IL-1β and IL-18 in curcumin treatment group were significantly increased,with the protein expression level of p62 was significantly decreased;The release of IL-1β,IL-18 and LDH in curcumin treatment group were significantly increased compared with the control;Curcumin treatment significantly reduced cell migration,tube formation and plate cloning ability of MCF-7 breast cancer cells compared to the control.After 4h pretreatment with MCC950 CA-074 and 3-MA,the protein expression levels of LC3,CTSB,NLRP3,Caspase-1,GSDMD-N,IL-1β,IL-18,the release of IL-1β and IL-18 and LDH were significantly decreased compared with the curcumin treated group;The expression level of p62 protein,cell migration,tube formation and plate cloning ability were significantly increased compared with curcumin treated group.We found that curcumin-induced pyroptosis of MCF-7 cells was based on the activation of NLRP3 inflammasome which was mediated by cytoplasmic CTSB.Curcumin upregulated the autophagy level of MCF-7 cells and induced the release of CTSB.Conclusion: Curcumin activates the NLRP3 inflammasome by inducing autophagy in MCF-7 breast cancer cells,thus causing pyroptotic cell death via Autophagy/CTSB/NLRP3/Caspase-1/GSDMD signaling pathway.Curcumin also inhibits the proliferation and migration of MCF-7 breast cancer cells. |
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