High expression of hepatitis B virus based vector with reporter gene in hepatitis B virus infection system.
OBJECTIVE
To construct a hepatitis B virus (HBV)-based vector with a reporter gene and to establish an HBV infection system to evaluate the availability of the vector.


METHODS
The HBV-based vectors with green fluorescence protein (GFP) were packaged into the liver of immunodeficient mice through transfer and helper plasmid using hydrodynamic technology. Wild type HBV (wt HBV) was provided by plasmid MC2009. Primary human hepatocytes (PHH) were isolated and infected with recombinant HBV (rHBV) or wt HBV. GFP expression was monitored by confocal and flow cytometry. HBV DNA and HBV surface antigen (HBSAg) were analyzed by PCR and ELISA.


RESULTS
3 x 10(7) wt HBV copies/mL and 5 x 10(6) rHBV copies/mL were collected from mice serum. In the wt HBV infected group, HBV progeny was 2 x 10(7) copies/mL and HBSAg was 770 ng/mL. In the rHBV infected group, GFP fluorescence was detected on d 3 post-infection and over 85% of the parenchymal cells expressed green fluorescence on d 12 post-infection. Compared with wt HBV in the PHH infection system, no rHBV DNA or HBSAg were detected in PHH culture media.


CONCLUSIONS
An effective HBV based vector was developed, which proved to be a useful HBV infection system. This vector and infection system can be applied to develop a therapeutic vector and study the HBV life cycle and viral pathogenesis.
Li
Shi-Hong
SH
Center of Experimental Animals, Sun Yat-sen University, 58 Zhongshan 2 Road, Guangzhou 510080, Guangdong Province, China.
Huang
Wen-Ge
WG
Huang
Bing
B
Chen
Xi-Gu
XG
D000818:Animals; D002478:Cells, Cultured; D004279:DNA, Viral / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D006509:Hepatitis B / Q000235:genetics* / Q000503:physiopathology; D006515:Hepatitis B virus / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology; D022781:Hepatocytes / Q000473:pathology / Q000821:virology; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D010957:Plasmids / Q000235:genetics; D019076:Transgenes / Q000235:genetics; D014771:Virion / Q000235:genetics
D004279:DNA, Viral