Our previous research demonstrated that hepatic-protectant
silibinin induced autophagy in human fibro-
sarcoma HT1080 cells through
reactive oxygen species (ROS) pathway.
Pifithrin-α (PFT-α), a specific inhibitor of p53, reduced autophagy and reversed
silibinin's growth-inhibitory effect; besides, PFT-α decreased the activation of
caspase-3, a crucial executor of apoptosis.
Silibinin upregulated expression of p53/phosphorylated-p53 (p-p53) in a time-dependent manner.
Catalase (scavenger of H(2)O(2)),
superoxide dismutase (SOD) (scavenger of O(2)(•-)), and
SB203580 (inhibitor of p38) attenuated upregulation of p53 expression, suggesting that p53 might be partially regulated by ROS-p38 pathway. On the other hand,
c-Jun N-terminal kinase (JNK) increased autophagic death in
silibinin-treated cells, and JNK/p-JNK expression was upregulated by
silibinin time-dependently. Inhibition of JNK by
SP600125 did not influence generation of ROS. Scavengers of H(2)O(2) or O(2)(•-) showed no effect on expression of JNK/p-JNK, indicating that JNK might not correlate with ROS in this process. However, activation of p53 was suppressed by
SP600125; therefore the function of p53 was possibly controlled by JNK as well. Western blotting analysis showed that PFT-α reduced activation of extracellular regulated kinase1/2 (ERK1/2) and expression of
protein kinase B (PKB, or Akt)/p-Akt.
PD98059 (inhibitor of
mitogen-activated protein kinase kinase (
MEK)/ERK) and
wortmannin (inhibitor of
phosphoinositide 3-kinase (PI3K)/Akt) enhanced
silibinin's cytotoxicity.
Wortmannin augmented
silibinin-induced autophagy, while
PD98059 did not affect autophagic ratio. These results suggest that
silibinin might induce p53-mediated autophagic cell death by activating ROS-p38 and JNK pathways, as well as inhibiting
MEK/ERK and PI3K/Akt pathways.