In vivo induction of a high-avidity, high-frequency cytotoxic T-lymphocyte response is associated with antiviral protective immunity - PubMed (original) (raw)

In vivo induction of a high-avidity, high-frequency cytotoxic T-lymphocyte response is associated with antiviral protective immunity

C Sedlik et al. J Virol. 2000 Jul.

Abstract

Many approaches are currently being developed to deliver exogenous antigen into the major histocompatibility complex class I-restricted antigen pathway, leading to in vivo priming of CD8(+) cytotoxic T cells. One attractive possibility consists of targeting the antigen to phagocytic or macropinocytic antigen-presenting cells. In this study, we demonstrate that strong CD8(+) class I-restricted cytotoxic responses are induced upon intraperitoneal immunization of mice with different peptides, characterized as CD8(+) T-cell epitopes, bound to 1-microm synthetic latex microspheres and injected in the absence of adjuvant. The cytotoxic response induced against a lymphocytic choriomeningitis virus (LCMV) peptide linked to these microspheres was compared to the cytotoxic T-lymphocyte (CTL) response obtained upon immunization with the nonreplicative porcine parvovirus-like particles (PPV:VLP) carrying the same peptide (PPV:VLP-LCMV) previously described (C. Sedlik, M. F. Saron, J. Sarraseca, I. Casal, and C. Leclerc, Proc. Natl. Acad. Sci. USA 94:7503-7508, 1997). We show that the induction of specific CTL activity by peptides bound to microspheres requires CD4(+) T-cell help in contrast to the CTL response obtained with the peptide delivered by viral pseudoparticles. Furthermore, PPV:VLP are 100-fold more efficient than microspheres in generating a strong CTL response characterized by a high frequency of specific T cells of high avidity. Moreover, PPV:VLP-LCMV are able to protect mice against a lethal LCMV challenge whereas microspheres carrying the LCMV epitope fail to confer such protection. This study demonstrates the crucial involvement of the frequency and avidity of CTLs in conferring antiviral protective immunity and highlights the importance of considering these parameters when developing new vaccine strategies.

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Figures

FIG. 1

FIG. 1

In vivo induction of CTL responses by synthetic peptides bound to 1-μm microspheres. BALB/c mice were immunized i.p. on days 0 and 21 with 109 beads bound to various synthetic peptides: LCMV p118–132 (A), Listeria p91–99 (B), or HIV p315–329 (C). Eight or 11 days after the last injection, spleen cells from immune mice were stimulated in vitro with priming peptide p118–132 (A), p91–99 (B), or p315–329 (C) in the presence of irradiated syngeneic spleen cells. The cytotoxic activity of these effector cells on 51Cr-labeled P815 target cells pulsed with the respective peptide or incubated with medium alone was measured. Data represent the means of percent specific lysis from duplicate samples. E/T ratio, effector-to-target cell ratio.

FIG. 2

FIG. 2

Comparison of the CTL responses induced by the LCMV synthetic peptide delivered by two different types of particulate vectors. BALB/c mice were immunized i.p. on days 0 and 21 (A) with various doses of LCMV beads or with 109 control HEL beads (B) and with 10 μg of PPV:VLP-LCMV or control PPV:VLP. Seven days after the last immunization, spleen cells were stimulated in vitro with the p118–132 peptide in the presence of syngeneic spleen cells and cytotoxic activity was measured on 51Cr-labeled P815 target cells pulsed with the p118–132 peptide (solid symbols) or incubated with medium alone (open symbols). Data represent the means of percent specific lysis from duplicate samples. E/T ratio, effector-to-target cell ratio.

FIG. 3

FIG. 3

CTL responses induced by LCMV beads are mediated by MHC class I-restricted cytotoxic cells. BALB/c mice were immunized i.p. on days 0 and 21 with 109 LCMV beads. Seven days after the last injection, spleen cells from immune mice were stimulated in vitro as described for Fig. 2 and cytotoxic activity was measured on 51Cr-labeled p118–132-pulsed target cells from various H-2 haplotypes at a 60:1 effector/target ratio. Lysis obtained with the target cells incubated with medium alone was less than 10% and is not shown. Data represent the means of percent specific lysis from duplicate samples.

FIG. 4

FIG. 4

In vivo induction of CD8+ effector CTLs by LCMV beads requires a CD4+ helper T-cell activity. (A) BALB/c mice were immunized i.p. on days 0 and 21 with 109 LCMV beads or HEL beads. Ten days after the last injection, spleen cells from immune mice were stimulated in vitro as described for Fig. 2. These effector cells were then treated with PBS or anti-CD4 or anti-CD8 MAb as described in Materials and Methods. (B) Mice were treated on days −1, 0, 1, 7, 14, and 20 with anti-CD4 or anti-CD8 MAb or with PBS and were injected i.p. on days 0 and 21 with 109 LCMV beads. Spleens were removed 7 days after the last immunization and cells were stimulated in vitro as described for Fig. 2. Cytotoxic activity on 51Cr-labeled p118–132-pulsed P815 target cells (solid symbols) and on cells incubated with medium alone (open symbols) was measured. Data represent the means of percent specific lysis from duplicate samples. E/T ratio, effector-to-target cell ratio.

FIG. 5

FIG. 5

Immunization with PPV:VLP-LCMV induces a high frequency of LCMV-specific T cells compared with immunization with LCMV beads. BALB/c mice were immunized i.p. on days 0 and 21 with 109 HIV or LCMV beads or 10 μg of PPV:VLP or PPV:VLP-LCMV. Seven days after the last injection, the frequency of LCMV-specific T cells in the spleen was measured by single-cell IFN-γ ELISPOT assay in the presence of p118–132 as described in Materials and Methods. Data are expressed as the number of SFC per spleen and represent the means obtained with six mice in two independent experiments. No SFC were obtained in the absence of p118–132.

FIG. 6

FIG. 6

Comparison of the peptide affinities of the CTL response induced by the LCMV peptide delivered by two different particulate vectors. BALB/c mice were immunized i.p. on days 0 and 21 with 109 LCMV beads or with 10 μg of PPV:VLP-LCMV. Ten days after the last injection, spleen cells from immune mice were stimulated in vitro as described for Fig. 2. Cytotoxic activity of these effector cells on 51Cr-labeled P815 target cells pulsed with various concentrations of the p118–132 peptide or incubated with medium alone was measured. Data represent the means of percent specific lysis from duplicate samples obtained at a 100:1 effector/target ratio.

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