Fig. 1: Molecular characteristics of spermatogonia remaining in adult testis following BU-induced damage.
From: Distinctive molecular features of regenerative stem cells in the damaged male germline
a Spermatogonial hierarchy and marker expression associated with distinct populations of Aundiff. b Kinetics of adult male germline recovery in response to treatment with low dose BU. Initiation of regenerative response occurs between D8 and 10. D10 post-BU was selected for subsequent analysis. c Representative wholemount IF of tubules D10 post-BU (n = 2 mice per group). d Flow-sorting strategy for isolation of Aundiff (E-Cadherin+ c-KIT– α6-integrin+) for RNA-seq. Percentages of cells within gates are indicated (n = 4 mice per group). e Venn-diagram comparing differentially expressed genes (DEGs) in regenerative versus control Aundiff (false discovery rate < 0.05 and absolute fold change > 1.5) and genes marking progenitor (Oct4-GFP+) or stem-associated (Oct4-GFP–) Aundiff fractions. Heatmap shows selected DEGs in regenerative vs. control Aundiff associated with stem and progenitor Aundiff. Genes of interest are highlighted. f Flow cytometry of adult Oct4-GFP testis D10 post-BU. Oct4-GFP expression within Aundiff population is shown (E-Cadherin+ c-KIT– α6-integrin+). Percentages of Aundiff Oct4-GFP + are included (n = 3 mice per group). g Representative wholemount IF of tubules D10 post-BU. Graph shows ratio of RARγ + (progenitor-enriched) to GFRα1 + (stem-enriched) spermatogonia (n = 3 mice per condition, 31–50 mm tubule length scored per animal). h Volcano plot of DEGs from e (analysis by Limma-voom and empirical Bayes method). Genes regulated by GDNF and FGF are highlighted, and relevant genes are indicated. i Representative flow cytometry analysis of adult WT testis D10 post-BU. EpCAM expression in Aundiff from control versus BU-treated mice and Adiff (E-CAD + c-KIT+) of controls is shown in histogram. Percentages of cells EpCAM+ are indicated. EpCAM is upregulated in E-Cadherin+ c-KIT + differentiating cells compared to Aundiff. Graph shows relative levels of EpCAM (median fluorescent intensity) on Aundiff of control and BU-treated mice (n = 3 per group). Scale bars: 50 μm. Dashed lines in wholemount IF indicate seminiferous tubule profile. Data are presented as mean values ± SEM in d, f, g, i. Significance determined by two-tailed unpaired student t test in d, f, g, i. Source data are provided as a Source Data file.
