Figure 4

Effect of paclitaxel on the growth of murine melanoma B16-LS9-luc⁺ xenografts. (A) Immunohistochemical analysis of zebrafish embryo eyes at 1 h (t₀) and 4 days (t₄) after orthotopic injection of B16-LS9-luc⁺ cells. Ki-67 (left panel) and vimentin (right panel) are detected in brown. Tumor area is highlighted in yellow. L, lens. Scale bar: 50 µm. (B) B16-LS9-luc⁺ bioluminescence signal was evaluated 1 h (t₀), 1 day (t₁), 2 days (t₂), 3 days (t₃), and 4 days (t₄) post implantation in the lysates of the whole embryos. Data are the mean ± SEM of five independent experiments. ** p < 0.01 vs. t₀ and t₁, ANOVA. (C) Effect of paclitaxel on the proliferation of B16-LS9-luc⁺ cells in vitro. Viable cells were counted after 72 h of incubation with increasing concentrations of the drug. Data are the mean ± SEM of three independent experiments. (D) B16-LS9-luc⁺ cells were cultured for 24 h in vitro in the absence or in the presence of 0.5 µM paclitaxel or with the corresponding volume of DMSO and then grafted in the zebrafish eye. Tumor growth was evaluated at t₄ by measuring the cell luminescence signal in the lysates of the whole embryos. Data are the mean ± SEM (n = 20). * p < 0.05 vs. DMSO, Student’s t-test. (E) After injection of B16-LS9-luc⁺ cells into the zebrafish eye, embryos were incubated at t₀ with 10 µM paclitaxel or with the corresponding volume of DMSO, both dissolved in fish water. Tumor growth was evaluated at t₄ by measuring the cell luminescence signal. Data are the mean ± SEM (n = 35). (F) After B16-LS9-luc⁺ cell grafting into the zebrafish eye, 0.4 pmoles/embryo of paclitaxel or of the corresponding volume of DMSO were injected in the same eye. Tumor growth was evaluated at t₄ by measuring the cell luminescence signal. Data are the mean ± SEM (n = 45). In (D–F), each dot represents one embryo. *** p < 0.0001 vs. DMSO, Student’s t-test.