Effect of postmortem change on development of apoptosis in rats

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Terminal deoxynucleotidyl transferase (TdT) -mediated dUTP-digoxigenin nick end labeling (TUNEL) assay is useful to detect apoptotic cells in situ. We examined whether or not postmortem delay affects the development of apoptotic signals of cells in various organs by hematoxy lin-eosin (H-E) and TUNEL assay.Wistar Imamichi rats were radiated by X-ray and sacrificed six hours after radiation. The spleen, thymus, adrenal and testis were excised and kept in moist chamber at room temperature. Each tissue was fixed after different time intervals 0,6,12,24 hours, and paraffin-embedded sections were made. In no radiation group, a little of TUNEL positive cells were observed in the spleen, thymus and testis sections, but not in the adrenal. No increase in the number of apoptotic cells was observed with postmortem delay. In the radiation group, much increase in the number of TUNEL positive cells of which nuclei were clearly and deeply stained was observed in the spleen and thymus corresponding to the area where shrinking nuclei were observed in H-E section. In testis sections, there was a little increase in the number of positively stained cells, and no change was observed in H-E section. With postmortem delay, the margin of the TUNEL positive cells changed from clear to indistinct, and the positive area was spread around. Our results shows that it is difficult to distinguish apoptotic cell from postmortem change. It is possible, however, to detect TUNEL positive cell with postmortem changes as the spread TUNEL positive area.