Establishment of mice model with human viral hepatitis B

AIM:To establish a mice model of hepatitis B by usingHBV-transgenic mice,and to transfer HBV-specific cytotoxicT lymphocytes (CTL) induced from syngeneic BALB/c miceimmunized by a eukaryotic expression vector containing HBVcomplete genome DNA.METHODS:HBV DNA was obtained from digested pBR322-2HBV and ligated with the vector pcDNA3.RecombinantpcDNA3-HBV was identified by restriction endonucleaseassay and transfected into human hepatoma cell line HepG2with lipofectin.ELISA was used to detect the expression ofHBsAg in culture supernatant,and RT-PCR to determinethe existence of HBV PreS1 mRNA.BALB/c mice wereimmunized with pcDNA3-HBV or pcDNA3 by intramuscularinjection.ELISA was used to detect the expression of HBsAbin serum.MTT assay was used to measure non-specific orspecific proliferation ability and specific killing activity ofspleen lymphocytes.Lymphocytes from immunized micewere transferred into HBV-transgenic mice (2.5×10~7 permouse).Forty-eight hours later,the level of serum proteinand transaminase was detected with biochemical method,liver and kidney were sectioned and stained by HE to observethe pathological changes.RESULTS:By enzyme digestion with Eco RI,Xho Ⅰ andHind Ⅲ,the recombinant pcDNA3-HBV was verified tocontain a single copy of HBV genome,which was insertedin the positive direction.HepG2 cells transfected with therecombinant could stably express PreS1 mRNA and HBsAg.After immunized by pcDNA3-HBV for 4 weeks,HBsAb wasdetected in the serum of BALB/c mice.The potential of spleenlymphocytes for both non-specific and specific proliferationand the specific killing activity against target cells wereenhanced.The transgenic mice in model group had nosignificant changes in the level of serum protein but had anobvious increase of ALT and AST.The liver had obviouspathological changes,while the kidney had no evidentdamage.CONCLUSION:A eukaryotic expression vector pcDNA3-HBV containing HBV complete genome is constructedsuccessfully.HepG2 cells transfected with the recombinantcan express PreS1 mRNA and HBsAg stably.Specific cellular immune response can be induced in mice immunized bypcDNA3-HBV.A mice model of acute hepatitis with HBV hasbeen established.