Protective effect of RIP and c-FLIP in preventing liver cancer cell apoptosis induced by TRAIL.
TRAIL (TNF-related apoptosis-inducing ligand) is a member of the tumor necrosis factor superfamily that can induce tumor selective death by up-regulating death receptor 4 (DR4) and DR5 expression. The study aimed to explore the role of RIP and c-FLIP genes in TRAIL induced liver cancer cell HepG2 and Hep3B apoptosis and related mechanism. RIP and c-FLIP silenced HepG2 and Hep3B cell model were established through siRNA. Western blot was applied to test c-FLIP, RIP, DR4, DR5, FADD, Caspase-3/8/9, ERK1/2, and DFF45 protein expression. Caspase-8 kit was used to detect Caspase-8 expression. Flow cytometry was performed to measure cell apoptosis rate. Acid phosphatase method was applied to determine cell cycle. TRAIL had no significant effect on Caspase-3/8/9, DR4, DR5, ERK1/2, and DFF45 protein expression, but up-regulated c-FLIP and RIP protein expression and reduced FADD expression level. After treated by the chemotherapy drug mitomycin and adriamycin, c-FLIP and RIP expression decreased significantly, while FADD increased. After knockout c-FLIP and RIP gene, HepG2 and Hep3B cell apoptosis rate induced by TRAIL increased obviously. Meanwhile, cell subG1 percentage increased markedly and exhibited G1 phase growth retardation. In addition, after two kinds of gene knockout, Caspase-8 was activated and produce Caspase-3 P20 and P24, leading DFF45 appeared DNA fragment P17 and P25. c-FLIP and RIP can inhibit Caspase-8 activation and prompting HepG2 and Hep3B resistant to cell apoptosis induced by TRAIL.
Department of Surgery, The 3rd Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Jichun
J
Sun
Department of Surgery, The 3rd Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Hongwu
H
Luo
Department of Surgery, The 3rd Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Wanpin
W
Nie
Department of Surgery, The Second Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Xundi
X
Xu
Department of Surgery, The Second Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Xiongying
X
Miao
Department of Surgery, The 3rd Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Feizhou
F
Huang
Department of Surgery, The 3rd Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Haiyan
H
Wu
Department of Surgery, The Second Xiangya Hospital, Central South University Changsha, Hunan Province, China.
Xiaoxin
X
Jin
D000971:Antineoplastic Combined Chemotherapy Protocols / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000235:genetics / Q000378:metabolism*; D053181:Caspase 8 / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm; D004789:Enzyme Activation; D059585:G1 Phase Cell Cycle Checkpoints / Q000187:drug effects; D056945:Hep G2 Cells; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D034622:RNA Interference; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000494:pharmacology*; D013997:Time Factors; D014162:Transfection
D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C505530:CASP8 protein, human; D053181:Caspase 8
RIP; TRAIL; apoptosis; c-FLIP; liver cancer