Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2008 Nov 10;381(1):106-15.
doi: 10.1016/j.virol.2008.08.019. Epub 2008 Sep 14.

Generation of functionally active HIV-1 specific CD8+ CTL in intestinal mucosa following mucosal, systemic or mixed prime-boost immunization

Igor M Belyakov  1 , Jeffrey D Ahlers, Gary J Nabel, Bernard Moss, Jay A Berzofsky
Affiliations

Generation of functionally active HIV-1 specific CD8+ CTL in intestinal mucosa following mucosal, systemic or mixed prime-boost immunization

Igor M Belyakov et al. Virology. .

Abstract

Gastrointestinal and vaginal mucosa are major sites of entry in natural HIV infection and therefore the preferred sites to elicit high-avidity CD8+ CTL by vaccination. We directly compare systemic and mucosal immunization in mice after DNA priming and boosting with rgp160 env expressed either in MVA or Ad for their ability to induce mucosal as well as systemic HIV-specific CTL. The optimal CTL response in the gut mucosa was observed after priming with the HIV-1 gp160 env DNA vaccine and boosting with rMVA or rAd encoding the same envelope gene all administered intrarectally (IR). Maximum levels of high-avidity CD8+ T cells were seen in intestinal lamina propria following this regimen. When the prime and boost routes were distinct, the delivery site of the boost had a greater impact than the DNA priming. IM DNA prime and IR rMVA boost were more effective than IR DNA prime and IM rMVA boost for eliciting mucosal CD8+ T-cell avidity. A systemic DNA-prime-followed by systemic rMVA boost induced high levels of high-avidity CD8+ T cells systemically, but responses were undetectable in mucosal sites. A single systemic immunization with rMVA was sufficient to induce high-avidity IFN-gamma secreting CD8+ T cells in systemic organs, whereas a single mucosal immunization with rMVA was not sufficient to elicit high-avidity CD8+ T cells in mucosa. Thus, a heterologous mucosal DNA prime-viral vectored boost strategy was needed. The requirement for a heterologous DNA prime-recombinant viral boost strategy for generation of high-avidity CD8+ T cells in mucosal sites in mice may be more stringent than for the induction of high-avidity CD8+ T cells in systemic compartments.

PubMed Disclaimer

Figures

Fig. 1
Fig. 1
The optimal CD8+ CTL response in the gut mucosa was observed after priming with DNA vaccine and boosting with HIV-1 rMVA or rAd both through the intrarectal route. Groups of BALB/c mice (five per group) were primed IM or IR with HIV DNA 3 times at three-week intervals between each immunization and boosted with HIV-1 rMVA (A, B) or HIV-1 rAd (C, D). Three weeks after the last immunization, SP (A, C) and PP (B, D) cells were stimulated in vitro for 7 days before assay. Cytolytic activity of CTL was measured by a 4-hr assay with 51Cr-labeled targets. 51Cr-labeled P815 targets were tested in the presence or absence of P18-I10 peptide (1 µM). E:T 100:1 ratio means 100 effectors to 1 target. Negative control values were subtracted from experimental lysis. SEM of triplicate cultures were all <5% of the mean.
Fig. 2
Fig. 2
Mucosal prime-boost immunization is essential for high-avidity CD8+ T cells in inductive mucosal sites (Peyer's patches), whereas a single systemic immunization with recombinant vector was sufficient to induce high-avidity CD8+ T cells in systemic sites. (A) CD8+ T-cell activity measured as IFN-γ-producing cells by ELISPOT as a function of peptide concentration for HIV-1 P18-I10 peptide in the PP after IR DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). T cells were activated with different concentrations of P18-I10 peptides (1, 0.01, 0.0001 µM and no peptide, as indicated). (B) IFN-γ-producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the SP after IR immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). (C) IFN-γ producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the PP after IM immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). (D) IFN-γ producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the SP after IM immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). Error bars represent SD of four mice/group. Ratio high/(high + low) CD8+ T-cell avidity (E–H). (E, G) PP; (F, H) SP. Ratio of the number of IFN-γ+ cells against 0.0001 µM peptide (high-avidity T cells) to the number of IFN-γ-producing cells against 1 µM peptide (total low- and high-avidity T cells). Prevalent high-avidity CD8+ T cells generated after prime-boost immunization, whereas recombinant vector alone was not effective in induction of functionally active CD8+ T cells in mucosal sites (I, J). To quantify the improvement in CD8+ T-cell avidity in Peyer's patches and spleen after prime-boost mucosal vs systemic immunizations, we calculated the fold difference in the avidity ratio after prime-boost immunization vs immunization with rMVA alone in Peyer's patches (I) or spleen (J) after IR vs IM immunizations. rMVA alone was delivered IR for mucosally immunized animals and IM for systemically immunized mice.
Fig. 2
Fig. 2
Mucosal prime-boost immunization is essential for high-avidity CD8+ T cells in inductive mucosal sites (Peyer's patches), whereas a single systemic immunization with recombinant vector was sufficient to induce high-avidity CD8+ T cells in systemic sites. (A) CD8+ T-cell activity measured as IFN-γ-producing cells by ELISPOT as a function of peptide concentration for HIV-1 P18-I10 peptide in the PP after IR DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). T cells were activated with different concentrations of P18-I10 peptides (1, 0.01, 0.0001 µM and no peptide, as indicated). (B) IFN-γ-producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the SP after IR immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). (C) IFN-γ producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the PP after IM immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). (D) IFN-γ producing cells as a function of peptide concentration for HIV-1 P18-I10 peptide in the SP after IM immunization with DNA prime and rMVA boost (closed diamond) or rMVA alone (closed square). Error bars represent SD of four mice/group. Ratio high/(high + low) CD8+ T-cell avidity (E–H). (E, G) PP; (F, H) SP. Ratio of the number of IFN-γ+ cells against 0.0001 µM peptide (high-avidity T cells) to the number of IFN-γ-producing cells against 1 µM peptide (total low- and high-avidity T cells). Prevalent high-avidity CD8+ T cells generated after prime-boost immunization, whereas recombinant vector alone was not effective in induction of functionally active CD8+ T cells in mucosal sites (I, J). To quantify the improvement in CD8+ T-cell avidity in Peyer's patches and spleen after prime-boost mucosal vs systemic immunizations, we calculated the fold difference in the avidity ratio after prime-boost immunization vs immunization with rMVA alone in Peyer's patches (I) or spleen (J) after IR vs IM immunizations. rMVA alone was delivered IR for mucosally immunized animals and IM for systemically immunized mice.
Fig. 3
Fig. 3
Mucosal prime-boost immunization is most effective for high-avidity CD8+ T cells in a mucosal effector site (lamina propria), whereas IR immunization with recombinant vector alone is not sufficient. (A) CD8+ T-cell activity measured as IFN-γ-producing cells by ELISPOT as a function of peptide concentration for HIV-1 P18-I10 peptide in the LP after IR immunization with DNA prime and rMVA boost or rMVA alone. T cells were activated with 1 µM (closed bar) or 0.0001 µM (open bar) concentration of P18-I10 peptides. (B) Ratio of the number of IFN-γ+ cells against 0.0001 µM peptide (high-avidity T cells) to the number of IFN-γ-producing cells against 1 µM peptide (total low- and high-avidity T cells) after IR or IM immunizations. The avidity ratio in LP after prime-boost immunization — closed bar and rMVA immunization alone — open bar. (C) Fold increase in the avidity ratio after prime-boost (IR/IR and IM/IM) and criss-cross (IR/IM and IM/IR) immunizations vs immunization with rMVA alone in lamina propria. rMVA alone delivered IR or IM and depending on the route of boost in the group of comparison. For IR/IR and IM/IR groups, rMVA was injected IR and for IM/IM and IR/IM, it was given IM.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Acierno PM, Schmitz JE, Gorgone DA, Sun Y, Santra S, Seaman MS, Newberg MH, Mascola JR, Nabel GJ, Panicali D, Letvin NL. Preservation of functional virus-specific memory CD8+ T lymphocytes in vaccinated, simian human immunodeficiency virus-infected rhesus monkeys. J. Immunol. 2006;176(9):5338–5345. - PubMed
    1. Ahlers JD, Belyakov IM, Thomas EK, Berzofsky JA. High affinity T-helper epitope induces complementary helper and APC polarization, increased CTL and protection against viral infection. J. Clin. Invest. 2001;108:1677–1685. - PMC - PubMed
    1. Ahlers JD, Belyakov IM, Berzofsky JA. Cytokine, chemokine and costimulatory molecule modulation to enhance efficacy of HIV vaccines. Current Molecular Medicine. 2003;3:285–301. - PubMed
    1. Alexander-Miller MA, Leggatt GR, Berzofsky JA. Selective expansion of high or low avidity cytotoxic T lymphocytes and efficacy for adoptive immunotherapy. Proc. Natl. Acad. Sci. U. S. A. 1996;93:4102–4107. - PMC - PubMed
    1. Allen TM, Vogel TU, Fuller DH, Mothe BR, Steffen S, Boyson JE, Shipley T, Fuller J, Hanke T, Sette A, Altman JD, Moss B, McMichael AJ, Watkins DI. Induction of AIDS virus-specific CTL activity in fresh, unstimulated peripheral blood lymphocytes from rhesus macaques vaccinated with a DNA prime/modified vaccinia virus Ankara boost regimen. J. Immunol. 2000;164(9):4968–4978. - PubMed

Publication types