title: Arsenic trioxide reduces chemo-resistance to 5-fluorouracil and cisplatin in HBx-HepG2 cells via complex mechanisms.
abstract: BACKGROUND
  Multidrug resistance is one of the major reasons chemotherapy-based treatments failed in hepatitis B virus (HBV) related hepatocellular carcinoma (HCC). Hypoxia is generally associated with tumor chemo-resistance. The aim of the study was to investigate the effect of Arsenic trioxide (As2O3) on the hypoxia-induced chemo-resistance to 5-FU or cisplatin and explored its underlying mechanism in the HBx-HepG2 cells.
  
  
  METHODS
  MTT assay was used to examine the cell viability. Mitochondrial membrane potential (MMP) and cell cycle was examined by flow cytometry. qRT-PCR was employed to observe the mRNA expression level; and western blot assay was used to determine the protein expression level.
  
  
  RESULTS
  Our results showed that transfection of HBx plasmid established the HBx-HepG2 cells expressing HBx, and the expression of HBx was confirmed by qRT-PCR and western blot. Exposure of HBx-HepG2 cells to hypoxia (5 % O2, 3 % O2, 1 % O2) for 48 h increased the chemo-resistance to 5-fluorouracil (5-FU) (50-1600 µM) and cisplatin (25-800 µM), reduced MMP, and caused the cell cycle arrest at G0/G1 phase in a concentration-dependent manner. Hypoxia also concentration-dependently (5 % O2, 3 % O2, 1 % O2) reduced mRNA expression level of P-glycoprotein (P-gp), multidrug resistance protein (MRP1), lung resistance protein (LRP), and decreased the protein expression level of hypoxia-inducible factor-1α (HIF-1α), P-gp MRP1, and LRP. Following pretreatment with As2O3 at a non-cytotoxic concentration re-sensitized the hypoxia (1 % O2)-induced chemo-resistance to 5-FU and cisplatin in HBx-HepG2 cells. As2O3 pretreatment also prevented MMP reduction and G0/G1 arrest induced by hypoxia. Meanwhile, As2O3 antagonized increase of HIF-1α protein induced by hypoxia, and it also suppresses the increase in expression levels of P-gp, MRP1, and LRP mRNA and proteins. In addition, As2O3 in combination with 5-FU treatment caused up-regulation of DR5, caspase 3, caspase 8, and caspase 9, and down-regulation of BCL-2, but had no effect of DR4.
  
  
  CONCLUSIONS
  Our results may suggest that As2O3 re-sensitizes hypoxia-induced chemo-resistance in HBx-HepG2 via complex pathways, and As2O3 may be a potential agent that given in combination with other anti-drugs for the treatment of HBV related HCC, which is resistant to chemotherapy.
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Guifang
authors.initials: G
authors.lastname: Yu
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Xuezhu
authors.initials: X
authors.lastname: Chen
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Shudi
authors.initials: S
authors.lastname: Chen
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Weipeng
authors.initials: W
authors.lastname: Ye
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Kailian
authors.initials: K
authors.lastname: Hou
authors.affiliation: Department of Oncology, The Fifth Affiliated Hospital of Guangzhou Medical University, No. 621, Gangwan Road, Huangpu District, Guangzhou, 510700 China.
authors.forename: Min
authors.initials: M
authors.lastname: Liang
keywords: 5-FU; Chemo-resistance; Cisplatin; HBV; HBx-HepG2; HCC; HIF-1α; LRP; MRP1; P-gp