title: Gli2 silencing enhances TRAIL-induced apoptosis and reduces tumor growth in human hepatoma cells in vivo.
abstract: Our previous studies have showed that Gli2 played a predominant role in proliferation and apoptosis resistance to TRAIL in hepatoma cells. The purpose of this study was to explore whether Gli2 silencing enhances efficiency of TRAIL for hepatoma in vivo. SMMC-7721-shRNA cells were implanted subcutaneously into nude mices and TRAIL was injected into the peritoneal space. TUNEL assay was used to detect apoptosis of tumor cells. The expression of Gli2, c-FLIPL, c-FLIPS, and Bcl-2 protein was determined by immunohistochemistry, respectively. Apoptosis and the level of caspases proteins in SMMC-7721 and HepG2 cells were detected by Flow cytometry and Western blot. Transcriptional activity of c-FLIP induced by Gli2 was measured by luciferase reporter gene assay. The results showed that lower volumes and weights of tumor were found in mice xenografted with SMMC-7721-shRNA cells as compared with control cells in the presence of TRAIL (P < 0.05). TUNEL assay showed significantly higher apoptosis index (AI) in the SMMC-7721-shRNA group than in the control groups (P < 0.05). There were remarkable positive correlations between Gli2 and c-FLIPL, c-FLIPS, Bcl-2 protein expression. Over-expression of c-FLIP or Bcl-2 in HepG2 cells attenuated TRAIL-induced apoptosis via suppression of caspase-8 or caspase-9 activity, respectively. Luciferase reporter gene assay found a regulatory sequence by which Gli2 activated transcription between -1007 to -244 in the c-FLIP promoter region. This study demonstrates that Gli2 showed regulatory activity on transcription of c-FLIP gene, and Gli2 silencing enhances TRAIL-induced apoptosis via down-regulation of c-FLIP and Bcl-2 in human hepatoma cells in vivo.
authors.affiliation: a Department of Hepatobiliary Surgery ; The Second Affiliated Hospital of Guangzhou Medical University ; Guangzhou , China.
authors.forename: Da-wei
authors.initials: DW
authors.lastname: Zhang
authors.forename: Hai-yan
authors.initials: HY
authors.lastname: Li
authors.forename: Wan-yee
authors.initials: WY
authors.lastname: Lau
authors.forename: Liang-qi
authors.initials: LQ
authors.lastname: Cao
authors.forename: Yue
authors.initials: Y
authors.lastname: Li
authors.forename: Xiao-feng
authors.initials: XF
authors.lastname: Jiang
authors.forename: Xue-wei
authors.initials: XW
authors.lastname: Yang
authors.forename: Ping
authors.initials: P
authors.lastname: Xue
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein / Q000235:genetics / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000473:pathology*; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D004195:Disease Models, Animal; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000235:genetics*; D008113:Liver Neoplasms / Q000235:genetics* / Q000473:pathology*; D008297:Male; D051379:Mice; D011401:Promoter Regions, Genetic; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000494:pharmacology*; D015533:Transcriptional Activation; D047368:Tumor Burden / Q000187:drug effects / Q000235:genetics; D023041:Xenograft Model Antitumor Assays; D000075746:Zinc Finger Protein Gli2
chemical_list: D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C000618551:Gli2 protein, mouse; D051741:Kruppel-Like Transcription Factors; D019253:Proto-Oncogene Proteins c-bcl-2; D053221:TNF-Related Apoptosis-Inducing Ligand; D000075746:Zinc Finger Protein Gli2; D020169:Caspases
keywords: FADD, fas-associated protein with death domain; Gli2; Gli2, glioma-associated oncogene 2; Hh, Hedgehog; PARP, poly ADP-ribose polymerase; TRAIL; TRAIL, tumor necrosis factor (TNF)-related apoptosis-inducing ligand; TUNEL, Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling; apoptosis; c-FLIP, cellular FLICE-inhibitory protein; hepatoma; shRNA, short hairpin RNA