Fig. 4

Representative image of an MDA-MB-468 zebrafish xenograft with a tumor in the PVS and several micrometastasis spread throughout the zebrafish larvae body, namely in brain, eye, gills and CHT (a). Human cancer cell lines (Hs578T, MDA-MB-468, HCT116, SW620 or HT29) were injected into the PVS of 2 dpf Tg(fli1:eGFP) zebrafish larvae. Zebrafish xenografts were treated in vivo with bevacizumab and compared with untreated controls. At 4 dpi, zebrafish xenografts were imaged by a fluorescent stereoscope to detect tumor cells throughout the zebrafish body (b). The percentage of xenografts that display micrometastasis in the CHT region was quantified (b, Hs578T **P = 0.0042, MDA-MB-468 **P = 0.0017, SW620 *P = 0.0112) and the outcomes are expressed as AVG ± SEM. Results are from 5 (Hs578T and SW620), 3 (MDA-MB-468) and 2 (HCT116 and HT29) independent experiments. The presence of micrometastasis in other organs besides the CHT was also quantified, namely in the brain, eye and gills, in untreated and bevacizumab-treated xenografts and the incidence of micrometastasis in two or more metastatic sites was determined (c, MDA-MB-468 from ~43.7 to ~23.4%, **P = 0.0081; HCT116 from ~19 to ~2.6%, *P = 0.0308; and HT29 from ~44 to ~20%, **P = 0.0061). Statistical analysis was performed using a Fisher’s exact test. Statistical results: (ns) > 0.05, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.