Fig. 6. Differential morphological responses of wild-type and MHCIIB^–/– embryonic fibroblasts during TNF+CHX-induced apoptosis. Wild-type and MHCIIB^–/– cells were incubated in the absence (control) or in the presence of TNF+CHX for 24 hours. (A) Representative bright-field micrographs of control and treated cells. TNF+CHX induced cell the shrinkage and detachment of wild-type and MHCIIB^–/– embryonic fibroblasts; however, more MHCIIB^–/– than wild type cells remained attached. Scale bar: 250 µm. (B) Adherent cells in ten different areas per experimental condition were counted. Quantification of the responses demonstrated a larger number of adherent MHCIIB^–/– than wild-type cells after the treatment. Data shown are the mean ± s.e.m. of six independent experiments. ^*P<0.0001, treated MHCIIB^–/– vs treated wild-type fibroblasts. (C) Wild-type and MHCIIB^–/– embryonic fibroblasts were incubated for increasing periods of time with TNF+CHX. Following treatments, the cells were processed for immunofluorescence microscopy with anti-active caspase 3 to identify the apoptotic cells and with Rhodamine-phalloidin to facilitate cell visualisation. Adherent, flat cells that were positive for active caspase 3 were counted and the percentage of these cells with respect to the total adherent cell population was calculated. The percentage of apoptotic cells that were flat and adherent was higher in MHCIIB^–/– than in wild-type fibroblasts from 16 hours onwards. More than 300 cells per experimental condition were counted. The values shown are the mean ± s.e.m. of three independent experiments. ^**P<0.00001, wild-type cells vs MHCIIB^–/– cells 16 and 24 hours treatment.