Opposing effects of tumor-expressed and liver-derived FVII on prometastatic genes and metastasis. (A) qPCR analysis of Slug and Sox9 expression in MDA^pcDNA and MDA^FVII after vehicle or rFVIIa (n = 4). (B) Western blot analysis of Sox9 expression in MDA^pcDNA and MDA^FVII after vehicle or rFVIIa. The analysis of prometastatic gene expression was analyzed using qPCR in MDA-MB-453 (n = 5) (C) and MDA-MB-468 (n = 5) (D). (E) Effects of rFVIIa on Wnt pathway activity as determined with a β-catenin–responsive luciferase construct in the presence or absence of the Wnt pathway activator LiCl (n = 3). (F) Effects of rFVIIa on Wnt pathway activity in the presence of Wnt3A (n = 3). Effects of mFVIIa on Sox9 in MDA^pcDNA and MDA^FVII was analyzed using qPCR (G) and western blot (H). (I) Schematic overview in time of tumor cell grafting and subsequent downregulation of liver mFVII by siRNA approaches. (J) In vivo orthotopic tumor growth of MDA^pcDNA and MDA^FVII after control (siNEG) or FVII (siF7) siRNA treatment (n = 6). Black asterisks show significant differences between tumor growth after MDA^FVII graftment in the presence of siNEG vs that after graftment of MDA^pcDNA cells in the presence of siNEG. Red asterisks show differences in tumor growth after graftment of MDA^pcDNA cells in the presence of siNEG vs that after graftment of MDA^pcDNA cells in the presence of siF7. (K) MDA^pcDNA and MDA^FVII liver metastasis after control (siNEG) or FVII (siF7) siRNA treatment as determined using qPCR and histochemistry (L). Statistically significant differences were tested using t tests.