FIG 4.
VLV-MHBs-immunized mice are protected from HBV hydrodynamic challenge. Mice were immunized with 107 IFU of VLV-HBcAg or VLV-MHBs or with 107 PFU of VSV-MHBs or VSV-HBcAg. HBV replication was then induced by hydrodynamic injection of pHBV1.3 plasmid (10 μg) at 6 weeks postimmunization. (A) Serum HBeAg levels were determined by ELISA at days 1, 4, and 7 after challenge (n ≥ 4). Percentages of day 1 levels are shown. (B) HBcAg expression in the liver was determined by immunohistochemistry at day 7 postchallenge. Quantifications represent average numbers of HBcAg-positive cells per 100× field (n = 3 with 5 fields counted per sample). Representative images are shown. (C) At day 7 postchallenge, Northern (NB) and Southern blot (SB) analyses were used to determine liver-associated HBV RNA and DNA levels, respectively (n ≥ 3). Representative samples are shown. RC, relaxed circular DNA; SS, single-stranded DNA. (D and E) The CD8 T cell recall response was measured at day 7 postchallenge by IFN-γ ELISPOT assay in either splenocytes (n ≥ 4) (D) or intrahepatic leukocytes (pooled samples from 2 or more mice) (E). Un, unimmunized.