Figure 1

Induction of tumor metastasis in twist1a+/kras+ transgenic zebrafish with Dox treatment. All zebrafish were treated with 20μg/mL Dox and 1μg/mL 4-OHT at 4 mpf, and samples were taken at 2 and 4 wpi. (A) Representative images of wild-type, twist1a+, kras+, and twist1a+/kras+ transgenic zebrafish. The left column shows the external appearance, the middle column shows the internal organs of the abdomen with the liver outlined (white dotted line), and the right column shows H&E staining of liver tissues at 4 wpi. Compared with the wild-type group, (B) the body lengths of kras+ and/or twist1a+/kras+ transgenic zebrafish differed significantly at 2 and 4 wpi, whereas (C) the body weights of transgenic zebrafish did not differ at 2 and 4 wpi. (D) Kaplan-Meier survival curves showing the percentage of survival at 4 wpi. (E) Fluorescence analysis presenting evidence of metastatic HCC at 2 wpi in twist1a+/kras+ zebrafish (white dotted line: primary and metastatic liver tumors). (F) Histological analysis revealed that wild-type, twist1a+, kras+, and twist1a+/kras+ transgenic zebrafish developed HCC or metastatic HCC at 4 wpi. (G) Representative images of normal, HP, HCA, and HCC using histological analysis. Scale bars: 50 or 200 μm. Student’s t-test or one-way ANOVA were used to assess differences between variables: * p < 0.05, *** p < 0.001.