Fig. 1

Effect of inhibition of VEGF/FGF on krasV12- and Myc-induced liver enlargement.

7 dpf fabp10+, kras+ or Myc+ larvae were treated with either 1 μM SU5402 or 1 μM SU6668 in the presence of 10 μg/ml Dox and 2D liver size was measured based on images. (A–C) Representative images of 7 dpf fabp10+ control larvae. (D–F) Representative images of 7 dpf kras+ larvae. (G–I) Representative images of 7 dpf Myc+ larvae. (J) Quantification of liver sizes for kras+ larvae (K) Quantification of liver sizes for Myc+ larvae. N = 20 from each groups; statistical significance: *p < 0.05, Scale bar = 20 μm.
Fig. 2

Effect of inhibition of Wnt signaling pathway on krasV12- and Myc-induced liver enlargement.

7 dpf fabp10+, kras+ or Myc+ larvae were treated with either 10 μM IWR1 or 10 μM Cardionogen 1 in the presence of 10 μg/ml Dox and 2D liver size was measured based on images. (A–C) Representative images of 7 dpf fabp10+ control larvae. (D–F) Representative images of 7 dpf kras+ larvae. (G–I) Representative images of 7 dpf Myc+ larvae. (J) Quantification of liver sizes for kras+ larvae. (K) Quantification of liver sizes for Myc+ larvae. N = 20 from each groups; statistical significance: *p < 0.05, Scale bar = 20 μm.
Fig. 3

Effect of inhibition of Hedgehog signaling pathway on krasV12- and Myc-induced liver enlargement.

7 dpf fabp10+, kras+ or Myc+ larvae were treated with either 10 μM cyclopamine or 1 μM GANT61 in the presence of 10 μg/ml Dox and 2D liver size was measured based on images. (A–C) Representative images of 7 dpf fabp10+ control larvae. (D–F) Representative images of 7 dpf kras+ larvae. (G–I) Representative images of 7 dpf Myc+ larvae. (J) Quantification of liver sizes for kras+ larvae. (K) Quantification of liver sizes for Myc+ larvae. N = 20 from each groups; statistical significance: *p < 0.05, Scale bar = 20 μm.
Fig. 4

Cell proliferation analysis of krasV12- and Myc-induced carcinogenesis.

7 dpf wild type (WT), kras+ or Myc+ larvae were treated with 10 μM SU5402, 10 μM IWR1 or 10 μM cyclopamine in the presence of 10 μg/ml Dox. Cell proliferation was analyzed by immunohistochemical staining with PCNA primary antibody. (A–D) Representative liver image of 7 dpf WT larvae. (E–H) Representative liver image of 7 dpf kras+ larvae. (I–L) Representative liver image of 7 dpf Myc+ larvae. (M) Statistical analysis of numbers of proliferating cells in the livers of kras+ larvae. (N) Statistical analysis of numbers of proliferating cells in the livers of Myc+ larvae. N = 20 from each groups; statistical significance: *p < 0.05, Scale bar = 20 μm.
Fig. 5

Cell apoptosis analysis of krasV12- and Myc-induced carcinogenesis.

7 dpf WT, kras+ or Myc+ larvae were treated with 10 μM SU5402, 10 μM IWR1 or 10 μM cyclopamine in the presence of 10 μg/ml Dox. Apoptosis was analyzed by immunohistochemical staining with digoxigenin-conjugated nucleaotide and incubated with anti-digoxigenin secondary antibody. (A–D) Representative liver images of 7 dpf WT larvae. (E–H) Representative liver images of 7 dpf kras+ larvae. (I–L) Representative liver images of 7 dpf Myc+ larvae. (M) Statistical analysis of numbers of apoptotic cells in the liver of kras + larvae. (N) Statistical analysis of numbers of apoptotic cells in the liver for Myc+ larvae. N = 20 from each groups; statistical significance: *p < 0.05, Scale bar = 20 μm.
Fig. 6

Histological examination of krasV12- and Myc-induced carcinogenesis.

7 dpf WT, kras+ and Myc+ larvae were treated with 10 μM SU5402, 10 μM IWR1 or 10 μM cyclopamine in the presence of 10 μg/ml Dox, and subjected histological analysis. (A–D) Representative liver images of 7 dpf WT larvae. Inset in (A) is a magnified area in the box with arrows pointing nucleoli. (E–H) Representative liver images of 7 dpf kras+ larvae. Inset in (E) is a magnified area in the box with arrows pointing to nucleoli of condensed nuclei. (I–L) Representative liver images of 7 dpf Myc+ liver larvae. Inset in (I) is a magnified area in the box with arrows pointing to nucleoli of condensed nuclei. (M) Quantification of liver histology observed for kras+ larvae. (N) Quantification of liver histology observed for Myc+ larvae. N = 10 from each group; scale bar = 20 μm.
Acknowledgments
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