Figure 1.
Schematic diagrams of recombinant baculoviruses expressing sH1N1 HA.
A pFBHERV-sH1N1 vector containing HERV env and sH1N1 HA genes under the control of a polyhedron promoter and hEF1α promoter, respectively, was constructed. The recombinant baculovirus, AcHERV-sH1N1-HA, was generated using the Bac-to-Bac baculovirus expression system.
Figure 2.
Characterization of the recombinant baculovirus, AcHERV-sH1N1-HA, and its expression in mammalian cells.
(A) PCR detection of HA and HERV. (1) pFastBacHERV-sH1N1-HA; (2) viral DNA for AcHERV-sH1N1-HA; (3) negative control.
(B) Western blot analysis of HA protein expression in 293TT cells. (1) Lysates of AcHERV-sH1N1-HA–infected 293TT cells; (2) lysates of mock-infected cells.
(C) Immunofluorescence detection of HA protein in 293TT cells.
Figure 3.
Cellular immune responses in mice immunized with AcHERV-sH1N1-HA or killed virus vaccine.
Mouse splenocytes were harvested 2 weeks after the final immunization. The number of IFN-γ–producing sH1N1 HA-specific T cells was determined using ELISPOT assays. *P<0.05 versus killed virus vaccine group; not significant (NS) versus the number of IFN-γ spots in the BALB/C group.
Figure 4.
Humoral immune response of mice immunized with AcHERV-sH1N1-HA or killed virus vaccine.
BALB/C and C57BL/6 mice were intramuscularly immunized with 5 × 107 PFU of recombinant AcHERV-sH1N1-HA, killed virus vaccine, or PBS. Antigen-specific IgG antibody titers against HA in BALB/C and C57BL/6 were determined by ELISA *P<0.05, versus BALB/C third immunization titer.
Figure 5.
HAI assays with A/California/04/2009(H1N1).
Four HAUs of A/California/04/2009(H1N1) virus was seeded into a V-shape, 96-well plate and combined with 2-fold serially diluted serum. After antigen-antibody reaction, 1% chicken blood was added for determination of HAI titer. *P<0.05, versus BALB/C third HAI titer.
Figure 6.
Protective efficacy of AcHERV-sH1N1-HA vaccine against lethal influenza challenge.
(A) Percentage change in body weight after challenge in mice. Changes in body weight (n=10) are expressed as the mean in each group. (B) Survival rate after challenge in mice. Symbols: AcHERV-sH1N1-HA–vaccinated C57BL/6, filled red diamonds; AcHERV-sH1N1-HA–vaccinated BALB/C, open red diamonds; killed virus-vaccinated C57BL/6, filled green triangles; killed virus-vaccinated BALB/C, open green triangles; PBS-injected C57BL/6, filled grey circles; PBS-injected BALB/C, open grey circles. *P<0.05 compared with PBS-injected mice; repeated measures ANOVA from day 1 to day 14 post infection.
Figure 7.
Histological lesions in the lung sections of immunized C57BL/6 mice after challenge with A/California/04/2009(H1N1).
Seven days post challenge, mice in each group were euthanized and their lungs were HE-stained for histological evaluation. (A) Normal C57BL/6 mice. (B) Non-vaccinated C57BL/6 mice challenged with virus. (C) C57BL/6 mice vaccinated with commercial killed virus vaccine. (D) C57BL/6 mice vaccinated with AcHERV-sH1N1-HA.