Figure 5.

Mcl-1 downregulation regulates cytocidal effects of UNBS1450. SH-SY5Y cells were transfected and treated as reported in Material and methods with two plasmids bearing WT Mcl-1, (1) and (2) (a and b) or KR Mcl-1 plasmid (c). Induction of apoptosis was analyzed by nuclear morphology after 16 h UNBS1450 (25 nM) treatment. In parallel, Mcl-1 level and caspase−3/−7 cleavage were assessed by western blot. (d) A concentration of 5 μM of MG132 was selected to study the impact on apoptosis (e) and (f) Mcl-1, caspase-3/7 cleavage in presence/absence of UNBS1450. MG132 was added after 10 h treatment with 25 nM UNBS1450, the minimum time required to induce Mcl-1 modulation in these cells (not shown), to minimize any side toxic effects of a longer exposure to the proteasome inhibitor. Western blots are representative of three independent experiments. The results are the mean of three independent experiments +/− S.D. Statistical analysis was performed by two-way ANOVA test (post-hoc test: Dunnett or Tukey) Significance is reported as **P<0.01, ***P<0.001, ****P<0.001 and ^#P<0.05 against the respective control values