Skip to main content
NCBI home page
NIHPA Author Manuscripts logoLink to NIHPA Author Manuscripts
. Author manuscript; available in PMC: 2012 Jul 20.
Published in final edited form as: J Intern Med. 2009 Jan;265(1):67–77. doi: 10.1111/j.1365-2796.2008.02051.x

Antiviral CD8+ T cells in the genital tract control viral replication and delay progression to AIDS after vaginal SIV challenge in rhesus macaques immunized with virulence attenuated SHIV 89.6

M Genescà 1,2, MB McChesney 2, CJ Miller 1,2
PMCID: PMC3401014  NIHMSID: NIHMS153063  PMID: 19093961

Abstract

Genescà M, McChesney MB, Miller CJ (Center for Comparative Medicine and California National Primate Research Center, University of California, Davis, CA, USA). Antiviral CD8+ T cells in the genital tract control viral replication and delay progression to AIDS after vaginal SIV challenge in rhesus macaques immunized with virulence attenuated SHIV 89.6 (Review).

The recently failed clinical efficacy trial of an acquired immunodeficiency syndrome (AIDS) vaccine that elicits antiviral CD8+ T-cell responses has emphasized the challenge of producing an effective vaccine against human immunodeficiency virus (HIV). In the simian immunodeficiency virus (SIV)/rhesus monkey model of AIDS, live-attenuated lentivirus ‘vaccines’ provide the best protection from uncontrolled viral replication and clinical disease after pathogenic SIV challenge. This review summarizes a recent series of studies in which we show that after vaginal SIV challenge of rhesus macaques immunized with an attenuated lentivirus protection from uncontrolled viral replication is primarily mediated by CD8+ T cells in the vaginal mucosa. Immunization with a chimeric simian/human immunodeficiency virus (SHIV) results in a systemic infection that induces a moderate population of SIV-specific CD8+ and CD4+ T cells with cytolytic potential in the vaginal mucosa. Depletion of CD8+ T cells at the time of SIV challenge completely abrogates the protection mediated by prior infection with attenuated SHIV. Further after vaginal SIV challenge, the only significant expansion of SIV-specific T cells occurs in the vagina in these animals. No significant expansion of T-cell responses was observed in systemic lymphoid tissues. Thus, the presence of SIV-specific CD8+ T cells in the vagina on the day of vaginal SIV challenge and a modest expansion of local effector T cells is sufficient to stop uncontrolled SIV replication. It seems that T-cell based vaccine strategies that can elicit mucosal effector CD8+ T-cell populations and avoid inducing systemic T-cell proliferation upon exposure to HIV have the greatest potential for mimicking the success of live-attenuated lentiviral vaccines.

Keywords: caspase-3, CD107, cytolytic T cells, Ki67, T-cell activation, vagina

Introduction

Sexual transmission is the predominant mode of human immunodeficiency virus (HIV) transmission. Whilst education and behavioural approaches can reduce HIV transmission, in the end a vaccine that can induce immune responses capable of containing the virus in the genital tract before the establishment of a systemic infection offers the greatest hope of stopping the HIV pandemic. Uncertainty as to the nature of the immunological response that is needed to control viral replication remains the greatest hurdle to developing an effective HIV vaccine [1]. Live-attenuated acquired immunodeficiency syndrome (AIDS) ‘vaccines’ provide the most durable protection from HIV challenge in the simian immunodeficiency virus (SIV)/rhesus macaque model of AIDS [2, 3] and whilst safety concerns preclude the use of live-attenuated lentiviral vaccine in humans, understanding the nature of protective immunity in these models may provide insight into the nature of protective anti-HIV immunity.

Infection with a virulence-attenuated simian/human immunodeficiency virus (SHIV) 89.6 consistently protects 60% female rhesus macaques from uncontrolled viral replication after vaginal challenge with SIV [4, 5]. This model is unique amongst live-attenuated lentivirus vaccine/macaque models because the immunizing virus was engineered by inserting the parts of the envelope (env) gene from two HIV-1 molecular clones into the backbone of the SIV-mac239, the challenge virus used in this study [6]. Thus, although most of the genomes of the vaccine and challenge viruses are identical, because the envelope (env) gene in the immunizing SHIV virus is derived from HIV, and only distantly related to SIV env, it is unlikely that neutralising antibodies have a role in this model of protection [4, 5]. Because the rate of protection from uncontrolled SIV replication after vaginal challenge is very high in SHV-immunized monkeys compared with other experimental AIDS vaccine models, defining protective immune responses in this animal model of effective vaccine-induced immunity may provide an important advance in the effort to develop an HIV vaccine.

Whilst it is generally accepted that virus-specific CD8+ T lymphocytes play a central role in controlling HIV and SIV replication [7–12], a recent efficacy trial of a HIV T-cell vaccine failed to show any evidence of protection from infection, or uncontrolled viral replication, in vaccine recipients. These results caused the trial to be stopped [1, 13]. The ramifications of this trial extend beyond the specific candidate vaccine. These results have raised some doubts as to the ability of vaccine-elicited T-cell responses to control HIV replication [14–17]. Recent studies suggest that the quality of the T-cell responses and not just the strength of the responses may be critical for the control of HIV replication [18, 19]. Similarly, in live-attenuated SHIV-immunized macaques, polyfunctional T-cell responses are associated with a better control of challenge virus replication [20].

The route of infection with nonpathogenic SHIV 89.6 does not alter the efficacy of SHIV 89.6 infection as vaginal, intranasal and intravenous inoculation of SHIV 89.6 all produce protection from vaginal challenge in about 60% of the immunized animals [4]. The similar level of protection following mucosal and systemic inoculation of SHIV likely reflects the fact that the genital tract immune responses are primed and maintained by viral replication in the genital tract during the 6–8 months of systemic SHIV infection prior to vaginal SIV challenge [21]. Thus, in this model the immunizing virus establishes a persistent disseminated infection and antiviral immune responses are distributed to all tissues. For simplicity, the animals in the recently published experiments [21, 22] that are summarized below were all immunized by intravenous inoculation of the attenuated SHIV89.6 prior to vaginal SIV challenge.

Vascular endothelium of the human cervicovaginal mucosa expresses intercellular adhesion molecule-1, vascular adhesion protein-1, P-selectin, vascular cell adhesion molecule-1 or E-selectin, but not MAdCAM-1 [23]. Thus, T cells expressing the adhesion molecules lymphocyte function-associated antigen-1, α4 integrins, and cutaneous lymphocyte antigen can bind to the genital tract vascular endothelium and enter the interstitial spaces of the genital mucosa [23]. In fact the vagina, cervix, skin and systemic lymph nodes share a common set of vascular addressins and recruit a similar set of lymphocytes from the circulation. In contrast, the gut and other mucosal sites have a unique set of vascular adressins that recruit a subset of lymphocytes expressing the α4β7 integrin [24]. Thus the T cells in the genital tract are distinct from mucosal T-cell populations that express the α4β7 integrin and use MAdCAM-1 to recirculate through gut tissues [24].

The female genital has a full compliment of immune functions and the distribution of immune cells in the genital tract of women and female rhesus macaques is very similar [25]. CD4+ and CD8+ T cells are found in the lamina propria and epithelium of the vagina and cervix [26]. CD8+ T cells are especially abundant in the deep layers of the vaginal epithelium and lamina propria of rhesus macaques. In both acute and chronic SIV infections, specific CD8+ cytotoxic T lymphocyte (CTL) are found in the vaginal mucosa [27, 28] of rhesus macaques and anti-HIV CTL are also present in the cervical mucosa of HIV-1-infected women [29]. Because CD8+ T-cell responses develop at 14 days postchallenge, about 7 days after the onset of uncontrolled viral replication in genital tract tissues [28], an effective vaccine-induced, mucosal memory T-cell response at the portal of entry may be able to contain viral replication in the genital tract and prevent the establishment of a systemic infection [30]. Despite the consensus that mucosal immune responses are desirable, there have been no reports of antiviral T cells in the genital tract of rhesus monkeys immunized with an attenuated lentivirus.

The ability of HIV/SIV to replicate robustly in the first hours after vaginal transmission is likely the single most important factor in determining if vaginal exposure will result in systemic infection [31]. Thus, as we have noted [21] and as has been suggested by others [32, 33], the presence of a T-cell response with cytolytic capacity at the portal of entry is likely a desirable feature for an HIV vaccine. Studies to define the anatomic distribution of immune responses and challenge virus in immunized animals have not been undertaken until recently. Thus the relative contribution of mucosal and systemic antiviral T-cell responses to immune control of viral replication remains to be determined in SHIV-immunized animals. Additionally, as survival and apoptotic signals are also induced through the cognate antigen-T-cell receptor interaction [34], the relative resistance or sensitivity of T cells to apoptosis upon antigenic stimulation may be an important factor in an effective anti-HIV T-cell response. However analysis of pro-and anti-apoptotic molecule expression in vaccine-induced T cells has never been undertaken in a HIV vaccine model. This review will summarize our recent efforts to define the role of CD8+ T cells in live-attenuated vaccine-induced protection from vaginal SIV challenge.

Intravenous SHIV immunization increases total CD8+ T-cell counts in blood but blunts proliferative and apoptotic signals in T cells

Intravenous infection of rhesus monkeys with SHIV 89.6 results in an acute phase of high-level viral replication followed by a persistent phase of low-level, asymptomatic infection [4, 35, 36]. SHIV infection results in a significant increase in the absolute number of CD8+ T cells in blood of the macaques on this study compared to preinfection levels [22] but after SIV challenge, there are no significant changes in blood CD8+ T-cell populations of the SHIV-immunized animals [22]. The frequency of activated and dividing CD8+ T lymphocytes (Ki-67+) in blood is significantly higher in SIV controls compared with SHIV immunized macaques after vaginal SIV challenge [22]. A trend towards an increase in activated/proliferating (Ki-67+) CD8+ T cells was also apparent in lymph nodes of the SIV control animals at 14 days postchallenge but not in lymph nodes of SHIV-immunized macaques [22]. The increased immune activation present in the SIV control macaques was accompanied by increased T-cell apoptosis (caspase-3+), presumably due to activation induced cell death, whilst SHIV-immunized macaques maintained a steady low frequency of proliferating T cells in the face of SIV challenge. The relatively tranquil environment in which the anti-SIV T-cell responses of SHIV-immunized macaques encounter SIV may be a key feature of immune protection elicited by attenuated lentiviruses.

Intravenous SHIV immunization induces cytolytic memory SIV-specific T-cell responses in the vagina that persist to the day of SIV challenge

Most SHIV-immunized animals analysed had SIV-specific CD8+ T cells in the vagina that secreted cytokines or degranulated in response to SIV in vitro (Fig. 1) [21]. This vaginal T-cell response was primarily composed of a variety of monofunctional SIV-specific CD8+ T cells that expressed CD107, a marker of cytolytic activity. Anti-SIV CD4+ T cells were found at high frequency in the vagina of all animals analysed, and a variable fraction of the anti-SIV CD107+ CD4+ T cells also secreted IL-2 and/or TNF-α [21]. In the cervix, only 30% of animals had SIV-specific CD8+ T-cell responses, and only 17% had SIV-specific CD4+ T-cell response [21]. Thus the frequency of CD8+ T-cell responses in the vagina but not the cervix of SHIV-immunized monkeys is consistent with the proportion of immunized animals protected from uncontrolled viral replication after vaginal SIV challenge.

Fig. 1.

Fig. 1

Open in a new tab

Schematic representation of the anatomic location and relative number of T cells in tissues on the day of vaginal simian immunodeficiency virus (SIV) challenge. simian/human immunodeficiency virus (SHIV) immunized animals (blue text) had an increased absolute number of CD8+ T cells in blood. Only rarely were these cells activated/dividing (Ki67+) and the proportion of CD8+ T cells expressing the activation marker CD38 was significantly lower in SHIV-immunized animals than in naϊve control animals prior to challenge. Further, in the SHIV-immunized animals few CD4+ T cells were activated/dividing (Ki67+). As depicted here, by 24 h after vaginal SIV challenge, the SHIV immunized animals treated with the CD8+ T-cell depleting Mab (red text) had no detectable CD8+ T cells and the number of activated CD4+ T cells (Ki67+) was low. Prior to SIV infection, the CD8+ and CD4+ T cells in blood and lymph nodes of unimmunized SIV naϊve control animals (black text) were rarely activated/proliferating (Ki67+), but 50% of T cells in normal rhesus macaques are CD38+. SHIV immunization induces populations of SIV-specific central memory, effector memory and cytotoxic CD8+ and CD4+ T cells in the vaginal mucosa, local and systemic lymph nodes. Although the same T-cell populations were present in the tissues of the CD8 Mab treated animals at the time of SIV challenge, by 24 h postchallenge, they were depleted of all CD8+ T cells (as depicted here). Note that a large population of SIV-specific CD4+ T cells remains in the mucosa and other tissues after CD8+ T-cell depletion. SIV-specific T cells were not present in the blood or tissues of unimmunized SIV naϊve control animals. Black arrows indicate the pathway of SIV dissemination after vaginal inoculation. The term ‘Total T cells’ denotes all T cells in freshly collected tissues and in blood. The term ‘SIV-specific T cells’ denotes cells responding to SIV peptide stimulation in vitro.

Intravenous SHIV immunization induced SIV-specific memory T-cell responses in peripheral blood and systemic lymphoid tissues

Based on the intracellular cytokine staining analyses, most animals have detectable anti-SIV CD8+ T-cell responses in peripheral blood mononuclear cells and genital lymph nodes at the end of the immunization period (Fig. 1); but less than half of the animals had a SIV-specific T cells in the axillary lymph nodes and mesenteric lymph nodes [21]. SIV-specific CD4+ T-cell responses were also found in at least one tissue of all SHIV-immunized animals [21].

Outcome of the vaginal challenge with SIVmac239

On day 7 postchallenge, about half of the unimmunized control monkeys and 16% of SHIV-immunized monkeys were plasma vRNA positive [22]. At 14 days postchallenge, all controls and 30% of immunized monkeys were plasma vRNA+. At 7 and 14 days postchallenge, the mean plasma vRNA level of the unimmunized control monkeys was significantly higher compared with the SHIV-immunized monkeys [22]. Strikingly, CD8+ T-cell depletion on the day of vaginal SIVmac239 challenge eliminates the protective effect of the SHIV immunization and CD8+ T-cell depleted animals had the highest plasma vRNA+ levels at 7 and 14 days postchallenge of any animal group [22].

Simian immunodeficiency virus env RNA levels, a specific marker of challenge virus replication, in the tissues of the unimmunized control animals were significantly higher than the SHIV immunized animals at day 7 and 14 postchallenge (Fig. 2). The vRNA levels in most tissues of the SHIV-immunized CD8+ T-cell depleted animals were similar to the vRNA levels in tissues of the unimmunized animals (C.J. Miller, manuscript in preparation). However it was striking that the vRNA levels in cervix and vagina of SHIV-immunized, CD8+ T-cell depleted animals were significantly higher than the levels in the genital tract of unimmunized animals (C.J. Miller, manuscript in preparation). Apparently in the absence of CD8+ T cells, the SIV-specific CD4+ T cells in the vaginal mucosa that are elicited by SHIV immunization [21] support higher levels of SIV replication than the population of CD4+ T-cells resident in the genital tract of naϊve rhesus macaques.

Fig. 2.

Fig. 2

Open in a new tab

The level of simian immunodeficiency virus (SIV) envelope RNA (/µg tissue RNA) in tissues of simian/human immunodeficiency virus SHIV-immunized and naϊve control animals after SIV challenge. Tissue samples from the vagina, cervix, inguinal lymph node, oburator lymph node, iliac lymph node, axillary lymph node, mesenteric lymph node, spleen and colon are included in the analysis. The left panel is 7 days postchallenge and the right panel is 14 days postchallenge. The P-values are the result of comparing mean tissues vRNA levels in an unpaired t-test.

SHIV-immunized macaques maintain polyfunctional anti-SIV CD8+ T cells in blood and tissues postchallenge

We detected SIV-specific CD8+ T-cell responses in peripheral blood mononuclear cells of 30% of SIV control animals at 14 days postchallenge, but none of the control animals had SIV-specific T cells in blood at day 7 postchallenge. These antiviral responses were characterized by monofunctional T-cells secreting IFN-γ or TNF-α [22]. In contrast, at day 0, 7 and 14 postchallenge, 58–70% of the SHIV-immunized monkeys had SIV-specific CD8+ T-cell responses in peripheral blood mononuclear cells [22]. In the CD8 depleted group, SIV-specific CD8+ T-cell responses in peripheral blood mononuclear cells could not be detected due to the effective elimination of the circulating CD8+ T cells.

Detectable SIV-specific CD8+ T-cell responses in the lymph nodes of unimmunized monkeys were rare at 7 and 14 days postchallenge. However prior to SIV challenge, live-attenuated SHIV89.6-immunization induced anti-SIV CD8+ T-cell responses in the genital lymph nodes of most animals (Fig. 1) [21]. Although genital tract antiviral effector T-cell responses were common, less than half of the SHIV-immunized animals had a SIV-specific T-cell response in axillary and mesenteric lymph nodes on the day of SIV challenge (Fig. 1) [21]. After challenge, there was minimal expansion of the anti-SIV Gag-specific CD8+ T cells in the lymph nodes of immunized macaques and most of the SIV-specific CD8+ T cells were monofunctional for IFN-γ or TNF-α (Fig. 3) [22]. Amongst the lymph nodes tested after SIV challenge, the most consistent anti-SIV CD8+ T-cell responses were in the genital lymph nodes of the SHIV-immunized animals.

Fig. 3.

Fig. 3

Open in a new tab

Schematic representation of the anatomic location and changes in the types and levels of T cells in tissues 14 days after vaginal simian immunodeficiency virus (SIV) challenge. Figure organization is similar to Fig. 1. After SIV challenge, simian/human immunodeficiency virus (SHIV) immunized animals (blue text) had steady levels of CD8+ and CD4+ T cells in blood and lymph nodes and the proportion of these cells that were activated or proliferating (Ki67+ or CD38+) did not change relative to the day of challenge. The SHIV immunized animals treated with the anti-CD8 T-cell depleting Mab (red text) had a very high frequency of proliferating, apoptotic and activated CD8+ T cells (Ki67+, caspase-3+ and CD38+). Further the CD4+ T cells in these animals were apparently prone to apoptosis (caspase-3+). Similar levels of enhanced T-cell proliferation and apoptosis (turnover) were found in the tissues of the unimmunized control animals (black text) at 14 days postchallenge. A significant expansion of the SIV-specific CD8+ T cells in the vaginal mucosa, and a trend towards an increased number of SIV-specific CD8+ T cells was found in the genital lymph nodes of SHIV-immunized animals at 14 days postchallenge. Unexpectedly, there was no evidence of expanded T cell populations, total or SIV specific, in the distant systemic lymphoid tissues of SHIV-immunized animals after vaginal SIV challenge. SIV-specific CD8+ T cells were still rare in the tissues of the CD8 Mab treated animals 14 days after SIV challenge, and as was the case with the control animals, SIV-specific CD8+ T cells were most readily detected in the vaginal mucosa.

After challenge, the expanded CD8+ T-cell responses in the genital tract of SHIV-immunized macaques are more polyfunctional and cytotoxic than in control monkeys

Some SIV control animals had SIV-specific CD8+ T-cell responses in the genital tract as early as 7 days postchallenge and these responses were associated with high levels of viral replication. The SIV-specific CD8+ T-cell responses in control animals declined in strength by day 14 postchallenge [22]. Approximately 70% of SHIV-immunized macaques had SIV-specific CD8+ T-cell responses in the vagina at 7 and 14 days postchallenge and the number of responding T cells were significantly increased in SHIV-immunized animals (P = 0.045, one-tailed unpaired t-test) by day 14 postchallenge [22]. After SIV challenge, the proportion of SHIV-immunized macaques with SIV-specific T-cell responses in cervix also increased [22]. Thus, an increase in genital tract SIV-specific T cells occurred in SHIV-immunized monkeys after SIV challenge, however, expansion of SIV-specific T cells was not observed in other tissues.

Notably, at all time points examined, the number of functions exhibited by SIV-specific CD8+ T cells in the cervix and vagina of the SHIV-immunized animals was markedly higher than control animals. Whilst all four functions assessed were represented in the SIV-specific CD8+ T cells in the vaginal mucosa of the SHIV-immunized macaques, cytotoxic/degranulating (CD107+) SIV-specific CD8+ T cells were very common after SIV challenge. In the SIV control monkeys, IFN-γ-secreting CD8+ cells predominated, and degranulation was not evident in the SIV-specific T-cell population in the vagina until 14 days postchallenge [22]. In fact, at 7 days postchallenge, SHIV-immunized macaques had significantly higher number of cytotoxic/degranualting (CD107+) cells in the vaginal mucosa compared with SIV control monkeys (P = 0.0084). In the cervix, the SIV-specific CD8+ T-cell responses of immunized macaques consisted of a few degranulating (CD107+) cells amongst a large population of IFN-γ+ T cells. In contrast, amongst the SIV-specific CD8+ T cells of the SIV control macaques, TNF-α was the predominant function in the cervix [22].

Attenuated SHIV89.6 infection induces SIV-specific CD8+ T cells with reduced apoptotic susceptibility

We have previously shown that an increased expression of survival signals in SIV-specific, polyfunctional CD8+ T cells is associated with better control of SIVmac239 replication in SHIV-immunized monkeys [20]. In peripheral blood mononuclear cells, SHIV-immunized and SIV control macaques had similar ratios of CD8+ T cells expressing survival (Bcl-2) and pro-apoptotic (caspase-3) molecules on the day of challenge [22]. Within the Bcl-2+ fraction of CD8+ T cells, we also determined the ratio of CD8+ T cells that were negative or positive for caspase-3 [22]. However, at 14 days postchallenge, there was a significant increase in the relative expression levels of pro-survival and pro-apoptotic mediators (Bcl-2+ caspase-3−/Bcl-2+ caspase-3+) in blood CD8+ T cells of the SIV controls compared with SHIV-immunized animals at 7 days postchallenge [22]. In peripheral blood mononuclear cells at days 0 and 7 postchallenge, SHIV-immunized macaques had a similar balance between long-lived and short-lived CD8+ T cells (Bcl-2+ caspase-3−/Bcl-2+ caspase-3+ CD8+ T-cell ratio) to the SIV control monkeys at day 0. But, at day 14 postchallenge, the SHIV-immunized monkeys had significantly fewer long-lived CD8+ T cells compared with SIV controls at 14 days postchallenge [22].

In the genital lymph node, the ratio of Bcl-2+ caspase-3− to Bcl-2+ caspase-3+ CD8+ T cells was significantly lower in the SIV control animals compared with SHIV-immunized animals at 14 days postchallenge [22]. Moreover, at 7 and 14 days postchallenge, SIV controls had significantly lower ratios compared with the SHIV-immunized monkeys in CD8+ T cells from the vagina [22] and cervix [22]. Thus, during the acute postchallenge period, the balance of pro-survival to pro-apoptotic signals in CD8+ T cells in the genital tract was significantly biased towards survival in the SHIV-immunized monkeys. However, this pro-survival bias was not seen in the peripheral blood mononuclear cells of the immunized monkeys.

Discussion

This review summarizes a recent series of studies in which we show that CD8+ T cells mediate the protection from uncontrolled viral replication and disease progression after vaginal SIV challenge in rhesus macaques immunized with an attenuated lentivirus, SHIV 89.6. The SIV-specific CD8+ T-cell response in vaginal mucosa, at the time of challenge and immediately after SIV challenge, consisted largely of effector CD8+ T cells with cytolytic potential. These effector CD8+T cells where likely critical to the observed protection as CD8+ T-cell depletion of vaccinated animals at the time of SIV challenge resulted in a total abrogation of control of viral replication. Importantly, there was no evidence of widespread expansion or proliferation of SIV-specific CD8+ T cells in the blood or tissues of SHIV-immunized animals after vaginal SIV challenge. In fact the only significant expansion of SIV-specific T cells in these animals after vaginal SIV challenge occurred in the vagina. Thus the presence of SIV-specific CD8+ T cells in the vagina on the day of vaginal SIV challenge and a modest expansion of effector T cells is sufficient to stop viral dissemination and uncontrolled SIV replication postchallenge.

At least two potentially overlapping mechanisms can be invoked to explain how expansion of SIV-specific T cells was limited to the site of mucosal SIV challenge in SHIV-immunized animals. First, the antiviral T-cell response in the vagina of SHIV-immunized animals may have been so effective at eliminating the initial round of infected cells in the first few days after challenge that insufficient antigen was produced to stimulate expansion of antiviral T cells in distal sites. Alternatively, SHIV infection may have induced regulatory T cells that suppress T-cell proliferation in most tissues thereby minimizing entry of SIV-specific T cells into tissues except in the genital tract the site of viral replication. In mice, this type of regulatory T-cell response has been reported to direct antiviral T cells away from lymph nodes and to the sites of herpes virus replication in the genital tract [37]. In any case, the data from the studies reviewed here indicate that, with minimal systemic activation or proliferation, anti-SIV CD8+ T cells in the vaginal mucosa mediate control of viral replication after vaginal SIV challenge in SHIV 89.6 immunized rhesus macaques.

The role of CD8+ T cells in SHIV-mediated protection seems clear. Immunization with SHIV89.6 induced and maintained antiviral CD4+ and CD8+ T cells in the vaginal mucosa of female rhesus macaques [21] and the proportion of immunized monkeys with these CD8+ T-cell responses (60%) correlates with the proportion of immunized monkeys protected from uncontrolled SIV replication (60%) in this model [4, 5]. Further SHIV-immunized animals maintained and increased anti-SIV CD8+ T-cell responses in the genital lymph node and vagina after pathogenic SIV challenge. Finally, immunized monkeys depleted of CD8α+ lymphocytes on the day of challenge were unable to control SIV replication after challenge, confirming a critical role for CD8+ T cells in SHIV-induced protection from vaginal SIV challenge.

The quality of the CD8+ T-cell response, including the cytokine profile and survival capacity of CD8+ T cells, appears to be an important component of an effective immune response against SIV. Significantly, protection was associated with the presence of CD8+ T cells with lytic function (CD107+) in the genital tract. Although SIV specific CD8+ T cells were detected in the genital tract of both groups at day 14 postchallenge, polyfunctional and degranulating CD8+ T cells were limited to the immunized monkeys. These findings suggest that SHIV89.6-induced SIV Gag-specific CD8+ T cells with lytic function at the portal of entry are critical to SHIV89.6 mediated protection from vaginal SIV challenge.

The goal of most conventional HIV vaccine strategies is to elicit a strong anamnestic CD8+ T-cell response after HIV challenge/exposure. However, in the studies reviewed here, the anamnestic anti-SIV T cell response was limited to the vaginal mucosa of the SHIV-immunized animals. The expansion of SIV-specific T-cell responses in the vagina of SHIV-immunized monkeys occurred without bystander T-cell proliferation in the lymph nodes as evidenced by constant levels of Ki-67 expression throughout the postchallenge period. In contrast Ki-67 expression increased in the CD8+ T cells in all tissues of SIV controls postchallenge, consistent with expansion or recruitment of specific CD8+ T cells to the initial site of viral dissemination after vaginal SIV inoculation [30]. Consistent with the conclusion that there was minimal systemic expansion and turnover of T cells, the levels of pro-apoptotic and anti-apoptotic molecules in T cells of SHIV-immunized animals did not change after SIV challenge. The lack of T-cell proliferation and apoptosis in the immunized monkeys supports the conclusion that an anamnestic T-cell response after vaginal SIV challenge was restricted to the genital tract of these monkeys. The increased number of circulating CD8+ T cells and the large proportion of IL-2 secreting SIV-specific CD8+ T cells in immunized animals prior to challenge suggests that SHIV infection led to a large pool of memory CD8+ T cells that replenished vaginal cytotoxic effector T cells at an adequate rate to control viral replication without producing bystander T-cell activation and proliferation in systemic tissues.

This studies reviewed here demonstrate that CD8+ T cells mediate the protection from vaginal SIVmac challenge in rhesus macaques previously immunized with SHIV89.6 [4, 5, 21]. Effective anti-SIV CD8+ T-cell responses were characterized by cells with multiple functions, but especially by the presence of T cells in the vagina that had cytolytic potential. Further, the CD8+ T cells in an effective response have increased survival capacity and do not dramatically proliferate upon antigenic stimulation. Thus, T-cell based vaccine strategies that can elicit mucosal effector CD8+ T-cell populations without inducing strong T-cell expansion in systemic tissues upon exposure to HIV have the greatest potential for mimicking the success of live-attenuated lentiviral vaccines.

Acknowledgements

This work was supported by Public Health Service grants U51RR00169 from the National Center for Research Resources; P01 AI066314 and R01 AI44480 from the National Institute of Allergy and Infectious Diseases and a gift from the James B. Pendleton charitable trust.

Footnotes

Conflict of interest statement

No conflict of interest was declared.

References

  • 1.Steinbrook R. One step forward, two steps back-will there ever be an AIDS vaccine? N Engl J Med. 2007;357:2653–2655. doi: 10.1056/NEJMp0708117. [DOI] [PubMed] [Google Scholar]
  • 2.Lohman BL, McChesney MB, Miller CJ, et al. A partially attenuated simian immunodeficiency virus induces host immunity that correlates to resistance to pathogenic virus challenge. J Virol. 1994;68:7021–7029. doi: 10.1128/jvi.68.11.7021-7029.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Wyand MS, Manson KH, Garcia-Moll M, Montefiori D, Desrosiers RC. Vaccine protection by a triple deletion mutant of simian immunodeficiency virus. J Virol. 1996;70:3714–3733. doi: 10.1128/jvi.70.6.3724-3733.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 4.Abel K, Compton L, Rourke T, et al. 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. J Virol. 2003;77:3099–3118. doi: 10.1128/JVI.77.5.3099-3118.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Miller CJ, McChesney MB, Lu X, et al. Rhesus macaques previously infected with simian/human immunodeficiency virus are protected from vaginal challenge with pathogenic SIVmac239. J Virol. 1997;71:1911–1921. doi: 10.1128/jvi.71.3.1911-1921.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Reimann KA, Li JT, Voss G, et al. An env gene derived from a primary HIV-1 isolate confers high in vivo replicative capacity to a chimeric simian/human immunodeficiency virus in rhesus monkeys. J Virol. 1996;70:3198–3206. doi: 10.1128/jvi.70.5.3198-3206.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Koup RA, Safrit JT, Cao Y, et al. Temporal association of cellular immune responses with the initial control of viremia in primary human immunodeficiency virus type 1 syndrome. J Virol. 1994;68:4650–4655. doi: 10.1128/jvi.68.7.4650-4655.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Borrow P, Lewicki H, Hahn BH, Shaw GM, Oldstone MB. Virus-specific CD8+ cytotoxic T-lymphocyte activity associated with control of viremia in primary human immunodeficiency virus type 1 infection. J Virol. 1994;68:6103–6110. doi: 10.1128/jvi.68.9.6103-6110.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Egan MA, Kuroda MJ, Voss G, et al. Use of major histocompatibility complex class I/peptide/beta2M tetramers to quantitate CD8(+) cytotoxic T lymphocytes specific for dominant and nondominant viral epitopes in simian-human immunodeficiency virus-infected rhesus monkeys. J Virol. 1999;73:5466–5472. doi: 10.1128/jvi.73.7.5466-5472.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Schmitz JE, Kuroda MJ, Santra S, et al. Control of viremia in simian immunodeficiency virus infection by CD8+ lymphocytes. Science. 1999;283:857–860. doi: 10.1126/science.283.5403.857. [DOI] [PubMed] [Google Scholar]
  • 11.Schmitz JE, Johnson RP, McClure HM, et al. Effect of CD8+ lymphocyte depletion on virus containment after simian immunodeficiency virus SIVmac251 challenge of live attenuated SIVmac239delta3-vaccinated rhesus macaques. J Virol. 2005;79:8131–8141. doi: 10.1128/JVI.79.13.8131-8141.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Jin X, Bauer DE, Tuttleton SE, et al. Dramatic rise in plasma viremia after CD8(+) T cell depletion in simian immunodeficiency virus-infected macaques. J Exp Med. 1999;189:991–998. doi: 10.1084/jem.189.6.991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Priddy FH, Brown D, Kublin J, et al. Safety and immunogenicity of a replication-incompetent adenovirus type 5 HIV-1 clade B gag/pol/nef vaccine in healthy adults. Clin Infect Dis. 2008;46:1769–1781. doi: 10.1086/587993. [DOI] [PubMed] [Google Scholar]
  • 14.Pantaleo G. HIV-1 T-cell vaccines: evaluating the next step. Lancet Infect Dis. 2008;8:82–83. doi: 10.1016/S1473-3099(07)70266-9. [DOI] [PubMed] [Google Scholar]
  • 15.Haynes BF, Shattock RJ. Critical issues in mucosal immunity for HIV-1 vaccine development. J Allergy Clin Immunol. 2008;122:3–9. doi: 10.1016/j.jaci.2008.03.036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Moore JP, Klasse PJ, Dolan MJ, Ahuja SK. AIDS/: a step into darkness or light? Science. 2008;320:753–755. doi: 10.1126/science.1154258. [DOI] [PubMed] [Google Scholar]
  • 17.Walker BD, Burton DR. Toward an AIDS vaccine. Science. 2008;320:760–764. doi: 10.1126/science.1152622. [DOI] [PubMed] [Google Scholar]
  • 18.Betts MR, Nason MC, West SM, et al. HIV nonprogressors preferentially maintain highly functional HIV-specific CD8+ T cells. Blood. 2006;107:4781–4789. doi: 10.1182/blood-2005-12-4818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Almeida JR, Price DA, Papagno L, et al. Superior control of HIV-1 replication by CD8+ T cells is reflected by their avidity, polyfunctionality, and clonal turnover. J Exp Med. 2007;204:2473–2485. doi: 10.1084/jem.20070784. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Genescà M, Rourke T, Li J, et al. Live attenuated lentivirus infection elicits polyfunctional simian immunodeficiency virus Gag-specific CD8+ T cells with reduced apoptotic susceptibility in rhesus macaques that control virus replication after challenge with pathogenic SIVmac239. J Immunol. 2007;179:4732–4740. doi: 10.4049/jimmunol.179.7.4732. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Genescà M, Skinner PJ, Bost K, et al. Protective attenuated lentivirus immunization induces SIV-specific T cells in the genital tract of rhesus monkeys. Mucosal Immunol. 2008;1:219–228. doi: 10.1038/mi.2008.6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Genescà M, Skinner PJ, Hong JJ, et al. With minimal systemic T cell expansion, CD8+ T cells mediate protection from vaginal SIV challenge in rhesus macaques immunized with attenuated SHIV89.6. J Virology. 2008;82:1181–1196. doi: 10.1128/JVI.01433-08. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Johansson EL, Rudin A, Wassen L, Holmgren J. Distribution of lymphocytes and adhesion molecules in human cervix and vagina. Immunology. 1999;96:272–277. doi: 10.1046/j.1365-2567.1999.00675.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Berlin C, Berg EL, Briskin MJ, et al. α4β7 Integrin mediates lymphocyte binding to the mucosal vascular addressin MAd-CAM-1. Cell. 1993;74:185–195. doi: 10.1016/0092-8674(93)90305-a. [DOI] [PubMed] [Google Scholar]
  • 25.Miller CJ, Lu FX. Anti-HIV and -SIV immunity in the vagina. Int Rev Immunol. 2003;22:65–76. doi: 10.1080/08830180305230. [DOI] [PubMed] [Google Scholar]
  • 26.Miller CJ, McChesney M, Moore PF. Langerhans cells, macrophages and lymphocyte subsets in the cervix and vagina of rhesus macaques. Lab Invest. 1992;67:628–634. [PubMed] [Google Scholar]
  • 27.Lohman BL, Miller CJ, McChesney MB. Antiviral cytotoxic T lymphocytes in vaginal mucosa of simian immunodeficiency virus-infected rhesus macaques. J Immunol. 1995;155:5855–5860. [PMC free article] [PubMed] [Google Scholar]
  • 28.Reynolds MR, Rakasz E, Skinner PJ, et al. CD8+ T-lymphocyte response to major immunodominant epitopes after vaginal exposure to simian immunodeficiency virus: too late and too little. J Virol. 2005;79:9228–9235. doi: 10.1128/JVI.79.14.9228-9235.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Musey L, Hu Y, Eckert L, Christensen M, Karchmer T, McElrath MJ. HIV-1 induces cytotoxic T lymphocytes in the cervix of infected women. J Exp Med. 1997;185:293–303. doi: 10.1084/jem.185.2.293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Miller CJ, Li Q, Abel K, et al. Propagation and dissemination of infection after vaginal transmission of simian immunodeficiency virus. J Virol. 2005;79:9217–9227. doi: 10.1128/JVI.79.14.9217-9227.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Miller CJ, Shattock RJ. Target cells in vaginal HIV transmission. Microbes Infect. 2003;5:59–67. doi: 10.1016/s1286-4579(02)00056-4. [DOI] [PubMed] [Google Scholar]
  • 32.Appay V, Nixon DF, Donahoe SM, et al. HIV-specific CD8(+) T cells produce antiviral cytokines but are impaired in cytolytic function. J Exp Med. 2000;192:63–75. doi: 10.1084/jem.192.1.63. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33.Rollman E, Smith MZ, Brooks AG, et al. Killing kinetics of simian immunodeficiency virus-specific CD8+ T cells: implications for HIV vaccine strategies. J Immunol. 2007;179:4571–4579. doi: 10.4049/jimmunol.179.7.4571. [DOI] [PubMed] [Google Scholar]
  • 34.Cheroutre H, Madakamutil L. Mucosal effector memory T cells: the other side of the coin. Cell Mol Life Sci. 2005;62:2853–2866. doi: 10.1007/s00018-005-5232-y. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Abel K, Rourke T, Lu D, Bost K, McChesney MB, Miller CJ. Abrogation of attenuated lentivirus-induced protection in Rhesus Macaques by administration of depo-provera before intravaginal challenge with simian immunodeficiency virus mac239. J Infect Dis. 2004;190:1697–1705. doi: 10.1086/424600. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Genesca M, Li J, Fritts L, et al. Depo-Provera abrogates attenuated lentivirus-induced protection in male rhesus macaques challenged intravenously with pathogenic SIVmac239. J Med Primatol. 2007;36:266–275. doi: 10.1111/j.1600-0684.2007.00244.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Lund JM, Hsing L, Pham TT, Rudensky AY. Coordination of early protective immunity to viral infection by regulatory T cells. Science. 2008;320:1220–1224. doi: 10.1126/science.1155209. [DOI] [PMC free article] [PubMed] [Google Scholar]

RESOURCES