Fragmented mitochondria are engulfed by the cytoplasmic vacuoles and
extruded from c-Flip-/-
MEFs during cell death. A and B, c-Flip-/-
MEFs stably expressing GFP-COX IV were unstimulated (A) or stimulated
with TNFα for 60 min (B). Scale bars, 25 μm.
C and D, c-Flip-/- MEFs were stimulated with
TNFα for 60 (C) or 90 (D) min and analyzed by
transmission electron microscopy. The enlarged images of the boxed
areas are shown in the right panels. The arrowheads and
arrows indicate extruded and fragmented mitochondria, respectively.
Scale bars, 500 nm. E, the percentages of cells showing
typical apoptosis, necrosis, and atypical apoptosis characterized by numerous
cytoplasmic vacuoles and mitochondrial extrusion were calculated by counting
randomly selected areas (total 100-200 cells/sample). Three independently
prepared samples were counted and are presented as the means ± S.D.
F, c-Flip-/- MEFs were stimulated with TNFα for 90
min and then analyzed by immunoelectron microscopy using anti-COX IV antibody,
followed by 10-nm colloidal gold-conjugated secondary antibody. Scale
bar, 200 nm.