Figure 3

A. Confocal immunomicrographs of microtubules (green) and DNA (red) in vehicle (0.01% DMSO) treated A549 cells displaying hallmarks of a typical cell division process in a cell-cycle, in particular, interphase (a), metaphase (b), early anaphase (c), late anaphase (d), telophase (e), and cytokinesis (f, g). B. In contrast, EM011 treatment arrested A549 cells at mitosis, impaired bipolar spindle formation, induced mitotic slippage, aneuploidy and apoptosis. EM011 treatment did not affect the radial array network of microtubules until 6 hrs of exposure (0 hr, (a); 6 hrs (b)). Aberrantly arrested cells with multipolar spindles were seen at 12 hrs (c) and 24 hrs (d) post-treatment. Cells started to exit mitosis without cell division beginning at about 30 hrs post-treatment (e). Thereafter, at 36-40 hrs post-treatment, several aberrant phenotypes were observed suggesting variability in cellular fate upon drug treatment. Multinucleate cells with small fragments of DNA indicating apoptotic bodies suggested cell death from G1-like tetraploid state (f). Some cells showed signs of DNA fragmentation while in mitosis suggesting mitotic death (g, h). However, some cells were seen to undergo asymmetric chaotic cytokinesis with triple midbody (i) and perhaps, led to the generation of aneuploid cells (j, k). Occasionally, aberrant separation into more than two abnormal cells (even multinucleated) were seen (l, m). At 48 hrs of drug exposure, several fragmented DNA pieces and clusters of small apoptotic bodies were observed (n).