Localization of caspase 8 with displaced occludin and DISC proteins after LYHY treatment of Eph4 monolayers on glass slides. A. Cells were treated with LYHY at 350 μM for 30 minutes and stained live during peptide treatment for activated caspase 8 (green, A1, 2) and activated caspase 3 (Red, A2). Cells were then fixed and stained for occludin (red pseudocolor, A1) and nuclei (Blue). Activated caspase 3 co-localizes with disrupted occludin and with activated caspase 8 in the right hand cell only. B. EPH4 cells on glass slides were treated for 2 hr with LYHY then stained live for activated caspase 8 (green B1, B2). After fixation monolayers were stained for occludin (pseudocolored red, B1), the DISC adapter protein FADD (red, B2) and nuclei (blue). The smaller panel under B5 shows an image that has been exposed for a longer time, and suggests that FADD is present at lower intensity in regions free of non-junctional occludin. C. EPH4 cells were treated with LYHY and stained for activated caspase 8 (green C1, C2; C4) and occludin as well as the death receptor Fas (red, C1 and white C5), and nuclei (blue). The smaller panel under C5 shows a more highly contrasted view of the region within the white box of the Fas panel. Like FADD, low levels of Fas appear to be present in cellular regions free of non-junctional occludin.