Major DNA fragmentation is a late event in apoptosis.
Apoptosis, the terminal morphological and biochemical events of programmed cell death, is characterized by specific changes in cell surface and nuclear morphology. In addition, DNA fragmentation in an internucleosomal pattern is detectable in mass cultures of apoptotic cells. However, DNA fragmentation and nuclear morphological changes may not necessarily be associated events. In this study, we examined OVCAR-3 and KB human carcinoma cells using time-lapse video phase-contrast microscopy to characterize the surface and nuclear morphological features of apoptosis in response to treatment with either taxol or ricin. The surface morphological features of apoptosis were the same in both cell types and with both drugs. Using an in situ nick-translation histochemical assay, these single cells were also examined for DNA strand breaks during apoptosis. Surface morphological changes demonstrated discrete stages of cell rounding, surface blebbing, followed by cessation of movement and the extension of thin surface microspikes, followed much later by surface blistering and cell lysis. Nuclear features examined by DAPI cytochemistry demonstrated apoptotic nuclear condensation very early in this sequence, usually at the time of initial surface blebbing. The nick-translation assay, however, demonstrated DNA strand breaks at a much later time, only after the formation of separated apoptotic bodies or after final cell lysis. This study points out the differences between surface and nuclear morphological changes in apoptosis, and the large temporal separation between nuclear morphological changes and major DNA fragmentation detectable by this in situ technique. This result suggests caution in using in situ nick-translation as a direct correlate of internucleosomal DNA fragmentation in apoptosis.
Collins
J A
JA
Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston 29425, USA.
Schandi
C A
CA
Young
K K
KK
Vesely
J
J
Willingham
M C
MC
D017209:Apoptosis; D002277:Carcinoma / Q000503:physiopathology*; D002462:Cell Membrane / Q000473:pathology; D053938:DNA Fragmentation; D004273:DNA, Neoplasm / Q000378:metabolism*; D005260:Female; D006801:Humans; D018715:Microscopy, Video; D008954:Models, Biological; D009336:Necrosis; D017239:Paclitaxel / Q000494:pharmacology; D012276:Ricin / Q000494:pharmacology; D013997:Time Factors; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D012276:Ricin; D017239:Paclitaxel