Fig. 2

Characterization of lrrc50^Hu255h zebrafish TGCT suggests analogy to human seminoma.

(A–D) Histological characterization of wild-type testis (left panels) and lrrc50^H255h tumors (right panels). Two magnifications shown A, B (largest in insert) all scale bars; 50 μm. The characterization indicates the presence of predominantly early germ cells and loss of differentiated germ cells in the tumors. (A) Morphological tissue analysis of toluidine blue stained sections indicates the presence of all stages of spermatogenesis in normal tissue (extensively described in Figure S3A), and shows a dramatic loss of differentiated germ cells in the tumor. (B) IHC characterization with proliferation marker α-phospho-HistoneH3 (pH3) shows synchronously dividing cell-clusters in normal tissue, indicative of differentiated germ cells. Early SPG is the only germ cell that can divide as a single cell, and the tumors show mostly single proliferating cells. Increased pH3 staining suggests the tumor tissue is highly proliferative (quantified in Figure S4A). (C) IHC using α-Ziwi (strong cytoplasmic expression). In normal tissue, Ziwi expression is restricted to early SPG and gradually and diffusely lost differentiated germ cells. With the exception of somatic tissue, the tumors are almost completely composed of early SPG. (D) IHC with meiosis marker α-γ-H2Ax shows normal tissue that is composed of various stages of differentiation, whereas these are predominantly absent in tumor tissue. (E) Chromatograms of WT zebrafish, heterozygote lrrc50^Hu255h and lrrc50^Hu255h tumors. We observe a loss of the remaining wild-type allele c.263T>A/p.Lys88* in 44.4% of the tumors (LOH).