Figure 4
From: Hyperosmotic stress enhances cytotoxicity of SMAC mimetics
Hyperosmotic stress enhances SMAC mimetic-induced, TNF-mediated apoptosis in ALL cells. (a and b) REH cells were grown in medium supplemented with 50 mM NaCl and challenged with the indicated concentrations of (a) LCL161 and (b) birinapant (Biri) in the presence and absence of etanercept (400 μg/ml). Shown are data points and mean±S.E.M. from three independent experiments. (c) REH cells were left untreated or challenged with 2.5 μM LCL161 or 2.5 μM birinapant with or without addition of 50 mM NaCl. After incubation for 16 h, cells were analyzed by flow cytometry for positivity of 7-AAD and annexin-V. Data shown are representative of two experiments performed. (dand e) REH cells were challenged with 0.3 μM BV6, LCL161 and birinapant with or without addition of 50 mM NaCl for 18 h. Caspase-8 and caspase-3/-7 activity was assessed using the fluorogenic substrates (IETD)2-R110 and (DEVD)2-R110, respectively. Data points from three independent experiments are shown. A line connects caspase activation under isotonic conditions with the corresponding value under hypertonic conditions (50 mM NaCl) for each experiment. (f) REH cells were treated with the indicated concentrations of BV6, LCL161 and birinapant with or without addition of 50 mM NaCl and 50 μM zVAD-fmk. Shown are data points and mean±S.E.M. from three independent experiments. Biri, birinapant
