Lysophosphatidate Reverses G2/M Arrest by Taxol Figure 1. Lysophosphatidate does not block the entry of Taxol treated MCF-7 cells into G2/M. Panels A–E, cells were treated with BrdU for 1 h to label cells in S-phase. Cells were then incubated for 12 h with 10% delipidated serum in the presence or absence of 50 nM Taxol or 5 mM LPA (Protocol A). The progression of cells through S, G2/M and G1 phases was quantified (Panels B–E) from the FACS analysis (Panel A). DNA histograms were composed with CellQuest software. The histogram for time 0 for all treatments shows the same results, repeated for clarity of the figure. Results are means 6 SD for three independent experiments. doi:10.1371/journal.pone.0020608.g001 test if cells recovered from Taxol treatment without LPA addition. Cells were stained with anti-tubulin antibody, anti-phospho- histone H3 antibody and DAPI for DNA (Fig. 3A). This identified mononucleated cells, multinucleated cells, those that were in apoptosis or dying (obvious deformations in cytoskeletal and nuclear structure) (Fig. 3B) and those in mitosis with abnormal spindle morphology (Fig. 3C). After incubating with Taxol for 26 h, 46% of MCF-7 cells were in mitosis (Fig. 4A). Incubating for a further 12 h in Taxol increased (p,0.05) the percentage of cells in mitosis to 52610%, whereas removal of Taxol decreased this value to 4065%, p,0.05. Incubating cells with LPA in the presence or absence of Taxol for 12 h decreased (p,0.05) the percentage of cells that were in mitosis to about 20% (Fig. 4A). LPA also decreased (p,0.05) the proportion of mitotic cells with abnormal spindles from 8669% in cells where Taxol was maintained to 4967% (Fig. 4B). Removal of Taxol decreased the percentage of the cells with abnormal spindles and the presence of LPA further amplified PLoS ONE | www.plosone.org 3 May 2011 | Volume 6 | Issue 5 | e20608