title: The tyrphostin AG1478 augments oridonin-induced A431 cell apoptosis by blockage of JNK MAPK and enhancement of oxidative stress.
abstract: Oridonin, a diterpenoid compound, extracted and purified from Rabdosia rubescen has been reported to have cytotoxic effect on tumour cells through apoptosis, and tyrosine kinase pathways are involved in these processes. A specific epidermal growth factor receptor (EGFR) inhibitor AG1478 was used to examine the relationship between EGFR signal pathways and oridonin-induced apoptosis and autophagy in EGFR abundant human epidermoid carcinoma A431 cells. Inhibition of EGFRaugmented oridonin-induced A431 cell apoptosis, while the changes of expression of downstream proteins, Bcl-2, Bcl-xL, Bax, cytochrome c, pro-caspase-3, Fas, FADD and pro-caspase-8 suggested that both the intrinsic and extrinsic apoptotic pathways are involved in these processes. Pretreatment with AG1478 aggravated oridonin-induced loss of mitochondrial membrane potential (MMP) and increased ROS generation in A431 cells, while a ROS scavenger, N-acetylcysteine (NAC) completely reversed oridonin- and AG1478-induced ROS generation and apoptosis. Therefore, AG1478 augmented oridonin-induced apoptosis by enhancing oxidative stress. Pretreatment with AG1478 decreased the expression of downstream MAPK proteins ERK, JNK and P38 and their phosphorylated forms to varying degrees compared with oridonin alone treatment. Then after administration of ERK, JNK and P38 inhibitors, only JNK inhibitor SP600125 effectively augmented oridonin-induced apoptosis and ROS generation. Therefore, in EGFR downstream pathways, JNK played a major role in preventing oridonin-induced apoptosis. Autophagy antagonised apoptosis and exerted a protective effect in A431 cells, and both AG1478 and SP600125 decreased oridonin-induced autophagy. Inhibition of EGFR augmented oridonin-induced apoptosis and this was caused by enhanced oxidative stress, and JNK played a major protective role by increasing autophagy, leading to antagonising apoptosis and ROS generation.
authors.affiliation: China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang, P. R. China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Yu
authors.forename: Si-Miao
authors.initials: SM
authors.lastname: Fan
authors.forename: Yuan-Chao
authors.initials: YC
authors.lastname: Ye
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: D017209:Apoptosis / Q000187:drug effects*; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D045786:Diterpenes, Kaurane / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D048670:MAP Kinase Kinase 4 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D018384:Oxidative Stress / Q000187:drug effects*; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology*; D020032:Tyrphostins / Q000494:pharmacology*
chemical_list: D045786:Diterpenes, Kaurane; D047428:Protein Kinase Inhibitors; D011799:Quinazolines; D020032:Tyrphostins; C011959:oridonin; C101044:RTKI cpd; D066246:ErbB Receptors; D048670:MAP Kinase Kinase 4