Figure 6

U73343 rescues BAECs from MTX-induce lytic cell death: Evaluation at the single cell level. BAECs, cultured on circular glass coverslips, were visualized using an inverted microscope equipped for epifluorescence as described in Materials and Methods. The cells were maintained at 35–37°C and both phase and fluorescence images were acquired at 30-sec intervals. Each bar of the histogram shows 1) the percentage of dead cells (black; indicated by uptake of ethidium and presence of large membrane blebs), 2) the percentage of cells undergoing zeiosis (red), and 3) the percentage of cells protected (green; indicated by the lack of nuclear staining and membrane blebbing) for the experimental condition given below each bar. The total number of cells examined for each condition is given in parenthesis above each bar. In each experiment 0.3 nM MTX was added at time 5 min; Bar 1, 0.2% DMSO added before MTX; bar 2, 10 μM U73343 added before MTX; bars 3, 4, 5, and 6, U73343 added 4, 6, 7, and 10 min after MTX, respectively. Percentages were determined at time 40 min using time-lapse videos to determine the number of cells undergoing zeiosis.