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New whole-body and organ systems were established to detect interphase cell death in the thymus, spleen and epithelial cells of intestinal crypts by low-dose radiation. Frozen sections of the thymus, spleen and intestine as thick as 8 microns were made after X-irradiation of whole body or removed organs, and then sections were stained with 0.02% erythrosin B solution. In unirradiated controls, a few numbers of erythrosin B positive cells (dead or dying cells) were observed in the thymus, spleen and intestinal crypt as a single cell death. When X rays were given to various strains of mice as a whole body dose, clusters of erythrosin B positive cells were produced. They appear at 2 hr after irradiation and reached maximum at 4 hr, remaining at a similar level until 8 hr after irradiation. The number of erythrosin B positive cells decreased after then by the elimination of dead cells, and they were observed like a single cell death at 24 hr after irradiation. When erythrosin B positive cells were scored 4 hr after irradiation, their total number and the number of cluster increased with increasing doses of X rays in the dose range from 0.05 to 0.5 Gy. It is noted that there were large differences in the radiation susceptibility among the inbred strains of mice for the induction of interphase cell death of thymic lymphocytes: e.g., high susceptibility in C57BL/6J and AKR/J, intermediate in N4, A/J, PT and ST, low in C3H/HeJ, HT, 101/H and DBA/2J, indicating that interphase cell death is genetically programmed. Similar results were observed with some chemical mutagens. Although a large increase of erythrosin B positive cells was observed in the thymus and spleen with methylprednisolone, there was no increase in the intestinal crypt, and vice versa with bleomycin, suggesting the organ specificity for the induction of interphase cell death by chemicals. For the in vitro method, the removed thymus was irradiated on the agar plate, and then incubated on the agar plate which was placed on the grid in the medium, so that the medium comes up to the organ through the agar plate. Frozen sections were made and stained with erythrosin B solution in the same way as the in vivo method. The number of erythrosin B positive cells in the organ culture system reached maximum at 5 hr after X-irradiation, e.g. slightly later than in the whole-body system. The efficiency was about 60% in C57BL/6J mice when compared with whole-body system.

Programmed Cell Death in Whole Body and Organ Systems by Low Dose Radiation