Agaricus blazei Extract Induces Apoptosis through ROS-Dependent JNK Activation Involving the Mitochondrial Pathway and Suppression of Constitutive NF-κB in THP-1 Cells
The role of ROS on ABE-mediated apoptosis. THP-1 cells
(2 × 105 cells ml−1) were pretreated with 10 mM NAC, and then
4 mg ml−1 of ABE was treated for 12 h (a) and 24 h (b–d). (a)
The redox status was monitored by the oxidation-sensitive fluorescent dyes
for and H2O2. The mitochondrial
membrane potential was also determined by flow cytometer using DiOC6 labeling of mitochondria.
(b) DNA contents were analyzed by flow cytometer. The sub-G0/G1
DNA fractions, indicating apoptosis, were determined as a percentage of the total number of cells.
(c) Equal amounts of cell lysate were resolved on SDS-PAGE, transferred to
nitrocellulose membranes, and probed with specific antibodies for cytochrome c,
Bax, and PARP. β-Actin was used as an internal loading control. (d) THP-1 cells were
collected following 24 h exposure to 4 mg ml−1 of ABE with or without 2 h of pretreatment
with NAC. Caspase-3 activity was determined following the manufacturer's protocol. Each point represents
the mean ± SD of three independent experiments. The significance was determined
by Student's t-test ( versus vehicle control).
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