Fig. 5

Icaritin inhibited the proliferation and promoted the apoptosis of HepG2/SMMC7721 cells. a Icaritin promoted apoptosis and inhibited the proliferation in HCC cells. Pro-caspase-3, cleaved-caspase-3, PTEN and GAPDH (as a control) were determined by western blot in HepG2 and SMMC7721 cells with DMSO or icaritin. b P53 knockdown or AFP overexpression attenuated icaritin-induced growth inhibition. The cell growth of icaritin-treated L02 and HepG2 and SMMC7721 cells was analyzed by MTT (top panel). HepG2 and SMMC7721 cells were stably transfected with or without pLL3.7-shp53 or pLL3.7 (as a control) individually (middle panel), or transfected with pcDNA3.1-AFP or pcDNA3.1 (as a control), treated with DMSO or 20 μM icaritin (bottom panel), and then cell growth was analyzed by MTT assay. c P53 knockdown or AFP overexpression attenuated icaritin-induced proliferation inhibition. The cell lines from (b) were treated with 20 μM icaritin or DMSO and then cell proliferation was analyzed by EdU assay. d P53 knockdown or AFP overexpression attenuated icaritin-induced apoptosis. The cell lines from (b) were treated with icaritin or DMSO for 20 h and then cell apoptosis was analyzed by flow cytometry. e Icaritin inhibited tumor activity in a xenograft mouse model, and the relationship between p53 and AFP in xenograft mouse. HepG2 cells were subcutaneously injected into nude mice. Icaritin was given at a dose of 100 mg/kg via intraperitoneal injection three times a week. The control group was given an equivalent amount of vehicle solvent. Four weeks after injection, the tumors were weighed, and size was measured. Data are shown as mean ± S.D. (n = 4). Expression levels of p53, AFP and GAPDH (as a control) in mice tumors were determined by western blot. *, p < 0.05; **, p < 0.01. The full-length images for blots in Fig. 5a and e were presented in Supplementary Fig. 8