Fig. 4.

DARac1 accelerates DAAkt1-induced glioma. (A) Two-month-old zebrafish showing bumps at the head. Inlets are merged bright field and fluorescence images. (B) Histologic findings of gliomas in Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1/UAS^GFP-DARac1 zebrafish. (a-g) Hematoxylin and eosin staining. Tumors frequently reveal heterogeneous grade of glioma showing a hypercellular nest within the tumor (red arrowheads) and increased vascularity (black arrowheads). (g and h) IHC and ISH for transgenes in a high-grade glioma. Inlets of (g) and (h) are IHC for Akt1 and ISH for DARac1, respectively. (i and j) ISH for DARac1. Cells at the invasion front show stronger expression of DARac1. Arrows indicate the direction of invasion. (k and l) Proliferation analyses reveal increased positivity to PCNA and PHH3, especially at the hypercellular areas, which accompanies a more robust expression of DARac1 on ISH (inlet). (C) Histologic grade of gliomas. Coexpression of DARac1 increased not only the tumor incidence but also the histologic grades of gliomas (D) Tumor incidence and mortality. 112 zebrafish from each Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1 and Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1/UAS^GFP-DARac1 line were followed. Tumor incidence was estimated by body bending, which preceded the appearance of an obvious bump in the head. The incidence rates of glioma in Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1 and Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1/UAS^GFP-DARac1 zebrafish are 14.3% and 44.4% at 3 months, 36.6% and 62.0% at 6 months, and 49.1% and 73.2% at 9 months, respectively. The mortality rates are also increased by the coexpression of DARac1: 0.9% vs. 22.2% at 3 months, 15.2% vs. 50.9% at 6 months, and 21.4% vs. 60.2% at 9 months. The tumor-free survival rates are much lower in the Ptf1a^Gal4/UAS^GFP-UAS^DAAkt1/UAS^GFP-DARac1 line. Bars, 50 μm.