Morphological and flow cytofluorometrical analyses of regenerated rat thymus after irradiation

Reconstituted rat thymuses were studied by immunohistochemistry, transmission electron microscopy (TEM) and flow cytofluorometry on days 0, 1, 2, 3, 5, and 7 after whole-body sublethal irradiation (6 Gy). One day after irradiation, numerous apoptotic cells were seen in the cortical thymus; the percentage of the sub-G1 peak representing apoptotic cells was 8.9% in the DNA content histogram of cytofluorometry. On day 3, the thymic structure had been destroyed and no distinction was drawn between the cortex and medulla. In this stage, few thymocytes but many macrophages were present, and the percentage of the sub-G1 peak reached a peak at 13.0%. Bromodeoxyuridine (BrdU) incorporated cells gradually increased after irradiation, and immunohistochemically numerous apoptotic cells were found primarily in the cortex on day 7. These thymocytes showed some levels of electron density of the nucleus as revealed by TEM. The percentage of S phase cells did not change markedly (20-30%) based on one-color DNA content histograms, but the percentage of early S and S phase cells was extremely high on day 7 (70%). These data indicate that a part of DNA synthetic cells may result in apoptosis. The combination of immunohistochemistry, TEM and flow cytofluorometry to analyze DNA content and BrdU incorporation proved a useful tool for investigating the reconstituted thymus.