Simian-human immunodeficiency virus SHIV89.6-induced protection against intravaginal challenge with pathogenic SIVmac239 is independent of the route of immunization and is associated with a combination of cytotoxic T-lymphocyte and alpha interferon responses

Abstract

Attenuated primate lentivirus vaccines provide the most consistent protection against challenge with pathogenic simian immunodeficiency virus (SIV). Thus, they provide an excellent model to examine the influence of the route of immunization on challenge outcome and to study vaccine-induced protective anti-SIV immune responses. In the present study, rhesus macaques were immunized with live nonpathogenic simian-human immunodeficiency virus (SHIV) 89.6 either intravenously or mucosally (intranasally or intravaginally) and then challenged intravaginally with pathogenic SIVmac239. The route of immunization did not affect mucosal challenge outcome after a prolonged period of systemic infection with the nonpathogenic vaccine virus. Further, protection from the SIV challenge was associated with the induction of multiple host immune effector mechanisms. A comparison of immune responses in vaccinated-protected and vaccinated-unprotected animals revealed that vaccinated-protected animals had higher frequencies of SIV Gag-specific cytotoxic T lymphocytes and gamma interferon (IFN-gamma)-secreting cells during the acute phase postchallenge. Vaccinated-protected animals also had a more pronounced increase in peripheral blood mononuclear cell IFN-alpha mRNA levels than did the vaccinated-unprotected animals in the first few weeks after challenge. Thus, innate as well as cellular anti-SIV immune responses appeared to contribute to the SHIV89.6-induced protection against intravaginal challenge with pathogenic SIVmac239.

FIG. 1.

Mean plasma vRNA levels and mean anti-SIV antibody titers in SHIV89.6-immunized rhesus macaques. (A) Plasma vRNA levels of SHIV89.6-infected monkeys, expressed as log₁₀ vRNA copies per ml of plasma. (B) Serum anti-SIV IgG antibody titers are reported as endpoint dilutions. The symbols represent i.v. (•), i.n. (▴), and i.vag. (▾) immunized monkeys. The 6 i.v. immunized monkeys that received a SHIV HXBc2 inoculation prior to SHIV89.6 immunization were not included in the calculation of mean plasma vRNA levels of i.v. immunized monkeys.

FIG. 2.

Plasma vRNA levels in vaccinated and naïve monkeys p.c. with SIVmac239. Each panel shows the plasma vRNA levels for individual monkeys in the vaccine-naïve (A) and in the i.v. (B), i.n. (C), and i.vag. (D) SHIV89.6-immunized monkeys p.c. with SIVmac239. Plasma vRNA levels are reported as log₁₀ vRNA copies per ml of plasma. The arrow indicates the detection limit of the assay (500 copies), and the dashed line marks the virological cutoff point for vaccine-induced protection (vRNA level of <10⁴ copies/ml). The number in the upper right corner represents the number of animals in each group. Each symbol represents results for an individual animal.

FIG. 3.

Percent change in absolute CD4 T-cell numbers p.c. with SIVmac239. p.c. CD4 T-cell numbers for monkeys categorized as vaccinated-protected and vaccinated-unprotected and for SIVmac239 control monkeys are shown as the average percent change ± standard deviation in CD4 T-cell numbers in each group relative to the mean CD4 T-cell numbers in that group prechallenge per μl of blood.

FIG. 4.

SIV Gag-specific pCTL frequencies in vaccinated-protected and vaccinated-unprotected animals. SIV Gag-specific pCTLs per 10⁶ CD8⁺ T cells of vaccinated-protected and vaccinated-unprotected animals were measured at the time of challenge and compared to pCTL frequencies during the acute phase p.c. Individual monkeys in each group are depicted by distinct symbols. Statistically significant differences in pCTL frequencies within each group and between vaccinated-protected and vaccinated-unprotected animals are indicated by P values.

FIG. 5.

Frequency of SIV Gag-specific IFN-γ-secreting cells in PBMC of naïve and vaccinated monkeys p.c. with SIVmac239. The number of SIV Gag-specific IFN-γ-secreting cells during the acute and chronic phases p.c. is reported as the number of IFN-γ SFC per 1 million PBMC. At each time point shown, the IFN-γ SFC numbers of naïve (▾), vaccinated-protected (▪), and vaccinated-unprotected (▴) animals were compared. The solid line shows the average number of SFC/10⁶ PBMC for each group. Statistically significant differences in the number of IFN-γ SFC between the groups are shown with the relevant P values. Results from individual animals within a group are represented by separate instances of each group symbol.

FIG. 6.

p.c. IFN-α mRNA levels in PBMC of vaccinated and naïve animals. The increase in PBMC IFN-α mRNA levels p.c. relative to prechallenge IFN-α PBMC mRNA levels of the same animal are shown for naïve (▾), vaccinated-protected (▪), and vaccinated-unprotected (▴) monkeys. Results from individual animals within a group are represented by separate instances of each group symbol. The dashed line indicates a twofold increase, the cutoff value. Solid lines represent the mean IFN-α mRNA levels within each group.

FIG. 7.

p.c. Mx mRNA levels in PBMC of vaccinated and naïve animals. The symbols are as described in the legend to Fig. 6.

FIG. 8.

p.c. MIP-1β mRNA levels in PBMC of vaccinated and naïve animals. The symbols are as described in the legend to Fig. 6.

FIG. 9.

TNF-α mRNA levels p.c. in PBMC of vaccinated and naïve animals. The increase in PBMC TNF-α mRNA levels p.c. compared to TNF-α PBMC mRNA levels of uninfected control animals are shown for naïve (▾), vaccinated-protected (▪), and vaccinated-unprotected (▴) monkeys. Results from individual animals within a group are represented by separate instances of each group symbol. Solid lines represent the mean TNF-α mRNA levels within each of the groups.