Figure 2.

ODC is critical for malignant transformation in prostate cells. A: Representative immunoblot illustrating the expression of the main enzymes of the polyamine pathway and spermine abundance in the normal RWPE1 cell line, compared to the prostate cancer cell lines CWR22 and LnCaP. B: Quantification of proliferation by MTS assay at 24, 48, 72, 96, and 120 hours after seeding. Three independent experiments for each time point are represented (t-test). C: Quantification of three independent experiments of Matrigel invasion assay (t-test). D: Quantification of proliferation by MTS assay at 24, 48, and 72 hours after seeding and treating with d,l-α-difluoromethylornithine (DFMO). Three independent experiments for each time point are represented (t-test). E: Quantification of proliferation by MTS assay at 24, 48, 96, and 120 hours after seeding and treating with 100 μmol/L of putrescine. Three independent experiments for each time point are represented (t-test). F: Quantification of Matrigel invasion assay comparing RWPE1_P with regular medium or with putrescine treatment. Three independent experiments for each time point are represented (t-test). G: Tumor initiation experiments showed that RWPE1_ODC cells had a high de novo tumor initiation capacity (tumors formed in 45 of 48 injection sites for the RWPE1_ODC cells). Three independent experiments were performed (t-test). H: Gross and microscopic appearance of a representative tumor that formed after s.c. injection of 1 × 10⁶ RWPE1_ODC cells in immunodeficient mice. Arrows point to mitotic figures observed in the tumor. n = 6 for each time point and cell line for experiment (B–F); n = 16 for each cell line (G). ^∗∗P < 0.01, ^∗∗∗∗P < 0.0001. Scale bars = 200 μm (H). GAPDH, glyceraldehyde-3-phosphate dehydrogenase; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium; ODC, ornithine decarboxylase; PAO, polyamine oxidase; SPSY, spermine synthase.