Article Figures & Data
Figures
- Figure 1.
Construction and expression of LMP2/1-hsp chimeric gene. A, scheme of the construction of a rAAV with LMP2/1-hsp chimeric gene that expresses a sequence encoding for a polyepitope protein that contains 13 HLA class I–restricted LMP1 and LMP2 epitopes. B, representative Western blot. Samples of 293 cells were prepared 24 h after infection with rAAV-LMP2/1 CTL-hsp or rAAV-GFP and results showed that LMP2/1-hsp chimeric gene (110 kDa) expression in infected cells (lane 1) but not in the control cells (lane 2).
- Figure 2.
In vivo tumor elimination assay. Mice were immunized with 5 × 1010 viral particles of rAAV-LMP2/1-hsp or rAAV-GFP or PBS (mock), and after 3 wk, were injected s.c. with 1 × 106 SP2/0-LMP2 or SP2/0 tumor cells. A, the tumor volume was monitored once every 5 d. B, the tumor load was weighted by 35 d. Columns, mean; bars, SE; *, P < 0.001 versus SP2/0-LMP2+rAAV-LMP2/1-hsp group. Every experiment was repeated thrice.
- Figure 3.
Survival time assay. BALB/c mice were challenged with 106 SP2/0-LMP2 tumor cells. Eight days after the challenge, when the tumor size was 0.2 cm in diameter, these mice were immunized with either rAAV-LMP2/1-hsp or rAAV-GFP (5 × 1010 viral particles/mouse) and then survival time was recorded. The data were the averages of 10 vaccinated mice from each group and presented as a percentage of mice surviving after immunization with rAAV-LMP2/1-hsp or rAAV-GFP.
- Figure 4.
LMP2 peptide131–139 responses induced by immunization with rAAV-LMP2/1CTL-hsp. BALB/c mice were immunized i.m. with rAAV-LMP2/1CTL-hsp. Four weeks after the vaccination, the T cells were collected and analyzed for in vitro CTL assay. A, SP2/0-LMP2 cells; B, SP2/0 cells; C, SP2/0 cells pulsed with LMP2 peptide131–139; and D, SP2/0 cells pulsed with nonspecific peptide. The data were the averages of six vaccinated mice. E/T, effector/target ratio; *, P < 0.05 versus controls and **, P < 0.01 versus controls.
- Figure 5.
Proliferation assays of rAAV-LMP2/1-hsp–vaccinated lymphocytes in response to LMP2 peptide 131 to 139. A, BALB/c mice were immunized i.m. with rAAV-LMP2/1-hsp, rAAV-GFP, or mock. Four weeks after the vaccination, the spleens were collected and analyzed for T-cell proliferation as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The data were the averages of six vaccinated mice of each group. Bars, SEs. Stimulation index was defined as the ratio of mean absorbance values for cells with antigen (LMP2 peptide 131–139) to mean absorbance values for cells without any added antigen; **, P < 0.001 versus controls. B, representative flow cytographs of T-cell subsets after LMP2 peptide131–139 stimulation for 72 h. C, proliferation kinetics model of T subsets after peptide stimulation.
- Figure 6.
Humoral immune responses in mice. Serum samples (n = 6) were collected at various time points. Antibodies against EBV LMP2 were detected with ELISA using a single dilution (1:500). Columns, mean (absorbance values); bars, SD; **, P < 0.01 versus the indicated groups.
Volume 8, Issue 9
