title: Silibinin activated p53 and induced autophagic death in human fibrosarcoma HT1080 cells via reactive oxygen species-p38 and c-Jun N-terminal kinase pathways.
abstract: 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.
authors.affiliation: China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016, China.
authors.forename: Wen-Jun
authors.initials: WJ
authors.lastname: Duan
authors.forename: Qi-Sheng
authors.initials: QS
authors.lastname: Li
authors.forename: Ming-Yu
authors.initials: MY
authors.lastname: Xia
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Tashiro
authors.forename: Satoshi
authors.initials: S
authors.lastname: Onodera
authors.forename: Takashi
authors.initials: T
authors.lastname: Ikejima
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology; D001343:Autophagy / Q000187:drug effects*; D045744:Cell Line, Tumor; D005354:Fibrosarcoma / Q000188:drug therapy*; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D017382:Reactive Oxygen Species; D000077385:Silybin; D012838:Silymarin / Q000494:pharmacology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
chemical_list: D000970:Antineoplastic Agents; D017382:Reactive Oxygen Species; D012838:Silymarin; D016159:Tumor Suppressor Protein p53; D000077385:Silybin; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases