Effects of PP7 on cell viability, motility, invasion, and endothelial tube formation in vitro. a HepG2 and HUVECs cells were treated with indicated concentrations of PP7 for 24 h and the cell viability was assessed using an MTT assay. b HepG2 and HUVECs cells were scratched with a pipette tip, and the cells were exposed to different concentrations of PP7 for 24 h. c Quantification of relative distance of the denuded zone of b. d HepG2 and HUVECs cells were incubated with various concentrations of PP7 for 24 h. Representative images are shown. e Quantification of migrating and invasive cells of d. f HUVEC cells were plated on Matrigel and treated with increasing concentrations of PP7 for 24 h. Capillary tube formation was observed and photographed at × 200. g Quantification of branch points of f. Values are expressed as means ± SD of triplicate experiments. *P < 0.05, **P < 0.01, versus control groups

Effect of PP7 on the NF-κB/MMP-9/VEGF pathway in HepG2 cells. a Western blot analysis for NF-κB p65 in the nuclear and cytoplasmic extracts of HepG2 cells. (B) MMP-9 and VEGF protein levels were determined via Western blotting. c, d were densitometric analysis of a. e, f were densitometric analysis of b. Values are expressed as means ± SD of triplicate experiments. *P < 0.05, **P < 0.01, versus control groups

Effects of PP7 on morphological changes, heart beat rate, survival rate, ISVs development and SIVs formation in zebrafish embryos. a The morphological features of zebrafish embryos observed and captured after treating embryos with PP7 for 24, 48 and 72 h. Scale bars represent 200 μM. b The average heart beating rates per minute of PP7 treated embryos were recorded at 72 h. c Survival rates of zebrafish embryos after treatment with PP7 for 24, 48 and 72 h. Lateral view of transgenic Tg(fli1:EGFP) zebrafish showing ISV (d) and SIV (g) growth after treatment for 24 h. Scale bars represent 500 μm (d) or 150 μm (g). Arrow showed the characteristic pattern of SIV. e and f was statistical analysis of number and length of ISVs of d, respectively. h, i was statistical analysis of number and length of SIVs of g, respectively. n = 10/group, *P < 0.05, **P < 0.01 as compared with control

Inhibitory effects of PP7 on the invasion, dissemination and metastasis of HeLa cells, A549 cells and HepG2 cells in zebrafish xenografts. CM-Dil-labeled (red) HeLa cells (a), A549 cells (e) and HepG2 cells (i) were microinjected into the yolk sac of 2dpf embryos, and treated with different concentrations of PP7. Tumor cell invasion, dissemination and metastasis were detected using a laser scanning confocal microscope at day 3 post-injection. Arrows indicate primary tumors, white arrowheads indicate disseminated tumor foci. Scale bars represent 250 μm. Quantification of tumor volume of HeLa cells (b), A549 cells (f) and HepG2 cells (j). Quantification of numbers of disseminated tumor foci of HeLa cells (c), A549 cells (g) and HepG2 cells (k). Averages of maximal distances of metastatic foci of HeLa cells (d), A549 cells (h) and HepG2 cells (l). Data are represented as means ± SD from 3 independent experiments. n = 10/group, *P < 0.05, **P < 0.01 as compared with control

Effects of PP7 on metastasis in H22 lung metastasis mice model. After 14-day treatment, the mice were all euthanized and the lungs were dissected out. a The numbers of the lung metastasis nodes were photographed using a camera after fixing with Bouin’s solution. b The number of metastasis nodules on the lung surface. c The body weights of tumor-bearing mice before and after the 14-day treatment. d Lung weights of the mice at the end of the study. Values represent the mean ± SD, n = 6. *P < 0.05 and **P < 0.01 were compared with model group