Many abnormal events were identified and these abnormalities became frequent 24 hours after the addition of HU, and at subsequent time points. In particular, the results suggested that cells bearing micronuclei frequently underwent abnormal mitoses, producing daughter cells with many disorders. Disorders were delivered to descendent cells and typically amplified. In contrast, cells with nuclear buds generally did not result in such catastrophic consequences after mitoses. Some cells showed no apparent abnormality until 72 hours after the addition of HU. The other finding from a series of these experiments was that the behavior of most micronuclei could be Trilostane followed for a long time. Some micronuclei could be followed for the duration of one cell cycle, from one mitosis to the next. Therefore, micronuclei in HeLa H2B-GFP cells were relatively stable. As a BIX-01294 control, we repeated similar five independent time-lapse observations in the absence of HU. By comparing these two types of experiments, we found that the average length of interphase as well as the length of mitosis were increased by the presence of 150 mM HU, which suggests transient cell cycle arrest by HU-induced DNA damage. Consistent with this notion, the frequencies of interphase cells with nuclear abnormalities and apoptosis, as well as the frequencies of abnormal mitosis increased dramatically in an HU-concentration dependent manner. The observed decrease in micronuclei frequency at 250 mM HU may be explained by growth arrest caused by higher HU concentration. If cells were cultured in the absence of HU for 3 days after they were treated with 150 mM HU for 3 days, some of the nuclear abnormalities still remained but mitotic abnormalities disappeared. These data suggest that most of the nuclear as well as the mitotic abnormalities seen in the HU-treated cultures were induced by replication stress caused by low HU concentrations. Most micronuclei appeared directly after completion of mitosis, although some appeared long after mitosis was completed. Early micronuclei were derived from either the chromatin bridge between separating anaphase chromosomes or the chromatid detached from the bulk of the chromosomes during the transition from metaphase to anaphase.