title: [The comparative study on the regulation of apoptosis by hepatitis B virus X protein between B and C genotype].
abstract: OBJECTIVE
  To investigate the apoptosis regulation on hepatoma cells by HBx between genotype B and C.
  
  
  METHODS
  Genotype B and C HBx gene fragments were amplified and inserted into green fluorescent protein (GFP) eukaryotic expression vector pEGFP-C1 to construct recombinant pGFP-XB and pGFP-XC. The pEGFP-C1, pGFP-XB and pGFP-XC were introduced into Bel-7402 cells by Fugene HD to obtain Bel-7402 cells expressing GFP. The transcription and expression of HBx gene were demonstrated by RT-PCR and Western Blot analysis. Bel-7402, Bel-7402/GFP, Bel-7402/GFP-XB, Bel-7402/GFP-XC cells were treated with adriamycin (2.5 microg/ml), and the apoptosis of the cells was determined by trypan blue exclusion, and flow cytometry analysis.
  
  
  RESULTS
  RT-PCR and Western Blot analysis showed that HBx genes of genotypes B and C were transcribed and expressed in Bel-7402/GFP-XB, Bel-7402/GFP-XC cells. Trypan blue exclusion showed adriamycin induced time-dependent cell death in Bel-7402, Bel-7402/GFP cells while no significant cell death was observed in Bel-7402/GFP-XB, Bel-7402/GFP-XC cells. Flow cytometry analysis indicated that no significant differences of apoptosis rates of Bel-7402/GFP-XB (3.87%) and of Bel7402/GFP-XC (4.01%) were observed (P > 0.05), moreover, no significant differences of Bel-7402/ GFP-XB (3.87%), Be17402/GFP-XC (4.01%) and of the untreated cells. Apoptosis rates in Bel-7402/GFP-XB (3.87%), Bel-7402/GFP-XC (4.01%) cells were significantly lower than those in Bel-7402 (27.05%) and Bel-7402/GFP (29.14%) cells at 48 hours after the adriamycin treatment (P < 0.01).
  
  
  CONCLUSIONS
  Bel-7402 cell lines expressing GFP, GFP-XB and GFP-XC fusion proteins were successfully established. HBV X protein blocks adriamycin-induced apoptosis of Bel-7402 cells. There is no difference between HBx of genotype B and C in inhibiting apoptosis induced by adriamycin.
authors.forename: Yue
authors.initials: Y
authors.lastname: Ming
authors.forename: Qi-Feng
authors.initials: QF
authors.lastname: Xie
authors.forename: Lin
authors.initials: L
authors.lastname: Yang
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002460:Cell Line; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000494:pharmacology; D006509:Hepatitis B / Q000188:drug therapy / Q000503:physiopathology* / Q000821:virology; D006515:Hepatitis B virus / Q000145:classification / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D000998:Antiviral Agents; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D004317:Doxorubicin