Morphological and histological analysis of testis

To observe testis gross morphology and measure testicular weight, mice over 12 weeks of age were euthanized after measuring their body weight. Numerical data is available in Table ^S2. The whole testis was observed using BX50 and SZX7 (Olympus, Tokyo, Japan) microscopes. For histological analysis, testes were fixed with Bouin's fixative (16045-1, Polysciences, Warrington, PA, USA) at 4 °C O/N, dehydrated in increasing ethanol concentrations and 100% xylene, embedded in paraffin, and sectioned (5 µm). The paraffin sections were hydrated with Xylene and decreasing ethanol concentrations and treated with 1% periodic acid (26605-32, Nacalai Tesque, Kyoto, Japan) for 10 min, treated with Schiff's reagent (193-08445, Wako) for 20 min, counterstained with Mayer's hematoxylin solution (131-09665, Wako) for 3 min, dehydrated in increasing ethanol concentrations, and finally mounted with Permount (SP15-100-1, Ferma, Tokyo, Japan). The sections were observed using a BX53 (Olympus) microscope. Seminiferous tubule stages were identified based on the morphological characteristics of the germ cell nuclei³².