Fig. 4

Morphological changes in cells upon exposure to LIP. a Live-cell fluorescence images of MCF-7 cells treated with FITC-tagged LIP. MCF-7 cells were incubated in medium containing 0.5 μg CellMask™ orange plasma membrane stain for 4 min and then incubated with FITC-tagged LIP (1.0 μg/ml). The cells were observed using an Olympus FluoView FV1000 confocal microscope and photographed at the indicated time points. (Magnification: 63×). b Microtubule and mitochondrial fragmentation and ER vacuolation. MCF-7 cells were incubated with or without LIP (0.5 μg/mL) at 37 °C for 12 h. The cells were stained with a monoclonal antibody against tubulin and incubated in medium containing 30 nM MitoTracker Red or ER tracker. Merged images of cells double-stained with DAPI are shown. c Leakage of various proteins from the cytosol and organelles. MCF-7 cells were incubated with (+) or without (−) 0.5 μg/mL LIP at 37 °C for 12 h. The culture medium and cells were independently collected, and four marker proteins in each fraction were separated by SDS-PAGE and detected by western blotting using appropriate antibodies. M and C indicate the medium and cell fractions, respectively. d Elevation of intracellular calcium concentrations in MCF-7 cells following LIP treatment. Each bar represents the mean value from three determinations with the standard deviation (SD). Data (mean ± SD) with asterisks significantly differ (**P < 0.01) between treatments