Apoptosis is the form of luteal microvascular endothelial cell death initiated by TNFα. (Panel A) Cells were treated TNFα (50 ng/ml) for 72 h and cell viability determined after Hoechst staining. The numbers of non-apoptotic cells per field in each of 10 separate fields of view per dish were counted. These data represent the mean ± SEM of at least three independent experiments. The ''*'' denotes significance (P < 0.05) compared to untreated control. (Panel B) Oligonucleosomal DNA fragmentation of CLENDO cells cultured for 48 h in serum and growth factor free conditions (control, lane 2); or for 12 h (lane 3), 24 h (lane 4), or 48 h (lane 5) with TNFα (50 ng/ml), and for 48 h with ceramide (50 μM, lane 6). Lane 1 shows a low molecular weight DNA ladder. (Panel C) Cleavage of Phi Phi Lux, demonstrating caspase-3 activity in CLENDO cells treated with TNFα (50 ng/ml) for 2 or 8 h.