Fig. 1

BHB significantly suppresses the proliferation, migration, and invasion of PCa cells. A, LNCaP, PC3, DU145, and RWPE-1 cells treated with various concentrations of BHB for 48 h were subjected to CCK8 assay. B, Colony formation assays were used to assess the in vitro tumor growth ability of cells treated with 0 or 15 mM BHB. The representative images and statistical charts are presented. C, Cell cycle assays were performed via flow cytometry in cells treated with or without 15 mM BHB. The ratio of cells distributed in three different phases was calculated. D, LNCaP, PC3, and DU145 cells were exposed to the corresponding concentrations of BHB for 48 h. The apoptotic rates were analyzed via flow cytometry using Annexin V-FITC/PI double staining methods. The representative images and statistical charts are shown. E, Migrative and Matrigel invasive ability of cells treated with or without 15mM BHB was tested via transwell assays. Representative images of migrating and invading cells on polycarbonate transwell membranes and statistical plots are shown. F, Western blot assays were used to detect the expression of representative epithelial-to-mesenchymal transition (EMT)-associated proteins in cells treated with or without BHB. All data were obtained by performing at least three replicate experiments and expressed as mean ± standard deviation (SD). Paired and two-tailed Student’s t-tests were performed, and significant differences were designated as *p < 0.05, **p < 0.01, and ***p < 0.001