title: DNA-PKcs deficiency sensitizes the human hepatoma HepG2 cells to cisplatin and 5-fluorouracil through suppression of the PI3K/Akt/NF-κB pathway.
abstract: The aim of the present study was to investigate the effects of DNA-PKcs deficiency on the chemosensitivity of human hepatoma HepG2 cells to cisplatin (CDDP) and 5-fluorouracil (5-Fu), and to explore the underlying molecular mechanism. After transfection with DNA-PKcs siRNA or control siRNA, HepG2 cells were exposed to combination treatment of CDDP and 5-Fu. The cell viability, DNA damage, cell apoptosis, intracellular reactive oxygen species and glutathione (GSH) level, expression of apoptosis related proteins, activity of phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway, and nuclear factor-κB (NF-κB) pathways were assessed. The combination of CDDP and 5-Fu had a synergistic cytotoxic effect in HepG2 cells in terms of the cell viability, DNA damage, apoptosis, and oxidative stress level. DNA-PKcs siRNA could sensitize the HepG2 cells to the combined treatment. DNA-PKcs suppression further reduced the Akt phosphorylation level and Bcl-2 expression in HepG2 cells exposed to CDDP and 5-Fu, but enhanced the expression of pro-apoptotic proteins p53 and caspase-3. Moreover, CDDP could inhibit the transcriptional activity of NF-κB through degradation of IkB-α, while 5-Fu alone seemed in some extent increases the NF-κB activity. The combined treatment with CDDP and 5-Fu resulted in significantly decrease of the transcriptional activity of NF-κB, which was further aggravated by DNA-PKcs siRNA treatment. In conclusion, DNA-PKcs suppression had complementary effects in combination with CDDP and 5-Fu treatment in HepG2 cells, which was associated with suppression of NF-κB signaling pathway cascade, activation of caspase-3 and p53, as well as down-regulation of Bcl-2 and GSH.
authors.affiliation: Department of General Surgery, Zhongshan Hospital, Fudan University, Shanghai, 200032, People's Republic of China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Fang
authors.forename: Zongtao
authors.initials: Z
authors.lastname: Chai
authors.forename: Dansong
authors.initials: D
authors.lastname: Wang
authors.forename: Tiantao
authors.initials: T
authors.lastname: Kuang
authors.forename: Wenchuan
authors.initials: W
authors.lastname: Wu
authors.forename: Wenhui
authors.initials: W
authors.lastname: Lou
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D002470:Cell Survival / Q000187:drug effects; D002945:Cisplatin / Q000494:pharmacology*; D004249:DNA Damage; D051747:DNA-Activated Protein Kinase / Q000235:genetics / Q000378:metabolism*; D019008:Drug Resistance, Neoplasm; D005472:Fluorouracil / Q000494:pharmacology*; D055785:Gene Knockdown Techniques; D005978:Glutathione / Q000378:metabolism; D056945:Hep G2 Cells; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D000970:Antineoplastic Agents; D016328:NF-kappa B; D009687:Nuclear Proteins; D017382:Reactive Oxygen Species; D019869:Phosphatidylinositol 3-Kinases; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D051057:Proto-Oncogene Proteins c-akt; D005978:Glutathione; D002945:Cisplatin; D005472:Fluorouracil