title: Increased apoptosis in HepG2.2.15 cells with hepatitis B virus expression by synergistic induction of interferon-gamma and tumour necrosis factor-alpha.
abstract: BACKGROUND
  Interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) were thought to be important immune mediators in host defence against hepatitis B virus (HBV) infection.
  
  
  OBJECTIVE
  To examine the synergistic effect of IFN-gamma and TNF-alpha on HBV-expressing HepG2.2.15 cells and its potential mechanisms.
  
  
  METHODS
  Cell viability was quantitatively measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assay. Cell morphology was captured using light microscopy. The typical DNA ladder test was performed using agarose gel electrophoresis. HBsAg and HBeAg titre changes were quantified by the enzyme-linked immunosorbent assay method. Gene expression was analysed using cDNA macroarrays.
  
  
  RESULTS
  Interferon-gamma (1000 U/ml) alone or combined with TNF-alpha (5 ng/ml) treatment resulted in apoptosis in HepG2.2.15 cells, but no significant apoptosis in the parent non-virus expressing HepG2 cells. IFN-gamma- and TNF-alpha-mediated apoptosis was reduced by lamivudine treatment in HepG2.2.15 cells. IFN-gamma combined with TNF-alpha reduced the titre of hepatitis B surface antigen and hepatitis B e antigen in the HepG2.2.15 cell line. For apoptosis-related gene changes, IFN regulatory factor 1 (IRF-1) (12.2-fold), c-myc (V00568 4.7-fold, L00058 2.4-fold) and caspase 7 (2.3-fold) genes were upregulated in the combination treatment group.
  
  
  CONCLUSIONS
  Interferon-gamma and TNF-alpha play a role in the cell death of HBV-expressing HepG2.2.15 cells. Expression of HBV leads to IFN-gamma- and TNF-alpha-mediated apoptosis in the cells. Increased IRF-1, c-myc and caspase 7 gene expression may be responsible for the synergistic induction of apoptosis by IFN-gamma and TNF-alpha.
authors.affiliation: Department of Gastroenterology and Hepatology, Zhongshan Hospital, Fudan University, Shanghai, China. shihongcn2000@yahoo.com.cn
authors.forename: Hong
authors.initials: H
authors.lastname: Shi
authors.forename: Shi-He
authors.initials: SH
authors.lastname: Guan
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D045744:Cell Line, Tumor; D020869:Gene Expression Profiling; D006515:Hepatitis B virus / Q000235:genetics / Q000378:metabolism*; D022781:Hepatocytes / Q000502:physiology* / Q000821:virology*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; C022616:thiazolyl blue