Fig. 4

Typical images of γH2AX-stained seminiferous tubules at stage I to VIII of the seminiferous cycle in rats. Images of the seminiferous tubules of a MMC-treated rat (a) and a vehicle-treated rat (b). Male rats (RccHan™:WIST) were intravenously treated with saline (vehicle) or MMC at 2 mg/kg/day for 2 consecutive days and the testes were obtained 24 h after the last dose. The testes were fixed with 4% paraformaldehyde, and histopathological specimens were prepared. The specimens were stained immunohistochemically with anti-γH2AX antibody (Abcam) and with hematoxylin and eosin (HE). Magnified images are shown in boxes below. γH2AX foci in spermatids or pachytene spermatocytes are shown (red arrows). Spermatocytes (pre-leptotene or leptotene), spermatogonia, and XY-bodies (sex vesicles, indicated with blue arrows) in pachytene spermatocytes are stained due to spontaneous physiological phosphorylation of H2AX, as previously reported [35, 36]