Estimating the infectivity of CCR5-tropic simian immunodeficiency virus SIV(mac251) in the gut

CD4+ T-cell depletion during acute human immunodeficiency virus infection occurs predominantly in the gastrointestinal mucosa. Using experimental data on SIV(mac251) viral load in blood and CD4+ T cells in the jejunum, we modeled the kinetics of CD4+ T-cell infection and death and estimated the viral infectivity. The infectivity of SIV(mac251) is higher than previously estimated for SHIV89.6P infection, but this higher infectivity is offset by a lower average peak viral load in SIV(mac251). Thus, the dynamics of target cell infection and death are remarkably similar between a CXCR4- and a CCR5-tropic infection in vivo.     

ALVAC-SIV-gag-pol-env-based vaccination and macaque major histocompatibility complex class I (A*01) delay simian immunodeficiency virus SIVmac-induced immunodeficiency.

T-cell-mediated immune effector mechanisms play an important role in the containment of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) replication after infection. Both vaccination- and infection-induced T-cell responses are dependent on the host major histocompatibility complex classes I and II (MHC-I and MHC-II) antigens. Here we report that both inherent, host-dependent immune responses to SIVmac251 infection and vaccination-induced immune responses to viral antigens were able to reduce virus replication and/or CD4+ T-cell loss. Both the presence of the MHC-I Mamu-A*01 genotype and vaccination of rhesus macaques with ALVAC-SIV-gag-pol-env (ALVAC-SIV-gpe) contributed to the restriction of SIVmac251 replication during primary infection, preservation of CD4+ T cells, and delayed disease progression following intrarectal challenge exposure of the animals to SIV(mac251 (561)). ALVAC-SIV-gpe immunization induced cytotoxic T-lymphocyte (CTL) responses cumulatively in 67% of the immunized animals. Following viral challenge, a significant secondary virus-specific CD8+ T-cell response was observed in the vaccinated macaques. In the same immunized macaques, a decrease in virus load during primary infection (P = 0.0078) and protection from CD4 loss during both acute and chronic phases of infection (P = 0.0099 and P = 0.03, respectively) were observed. A trend for enhanced survival of the vaccinated macaques was also observed. Neither boosting the ALVAC-SIV-gpe with gp120 immunizations nor administering the vaccine by the combination of mucosal and systemic immunization routes increased significantly the protective effect of the ALVAC-SIV-gpe vaccine. While assessing the role of MHC-I Mamu-A*01 alone in the restriction of viremia following challenge of nonvaccinated animals with other SIV isolates, we observed that the virus load was not significantly lower in Mamu-A*01-positive macaques following intravenous challenge with either SIV(mac251 (561)) or SIV(SME660). However, a significant delay in CD4+ T-cell loss was observed in Mamu-A*01-positive macaques in each group. Of interest, in the case of intravenous or intrarectal challenge with the chimeric SIV/HIV strains SHIV(89.6P) or SHIV(KU2), respectively, MHC-I Mamu-A*01-positive macaques did not significantly restrict primary viremia. The finding of the protective effect of the Mamu-A*01 molecule parallels the protective effect of the B*5701 HLA allele in HIV-1-infected humans and needs to be accounted for in the evaluation of vaccine efficacy against SIV challenge models.     

TRIM5α does not affect simian immunodeficiency virus SIV(mac251) replication in vaccinated or unvaccinated Indian rhesus macaques following intrarectal challenge exposure

TRIM5α is a natural resistance factor that binds retroviral capsid proteins and restricts virus replication. The B30.2/SPRY domain of TRIM5α is polymorphic in rhesus macaques, and some alleles are associated with reduced simian immunodeficiency virus (SIV) SIV(mac251) and SIV(smE543) replication in vivo. We determined the distribution of TRIM5α alleles by PCR and sequence analysis of the B30.2/SPRY domain in a cohort of 82 macaques. Thirty-nine of these macaques were mock vaccinated, 43 were vaccinated with either DNA-SIV/ALVAC-SIV/gp120, ALVAC-SIV/gp120, or gp120 alone, and all were exposed intrarectally to SIV(mac251) at one of three doses. We assessed whether the TRIM5α genotype of the macaques affected the replication of challenge virus by studying the number of SIV variants transmitted, the number of exposures required, the SIV(mac251) viral level in plasma and tissue, and the CD4(+) T-cell counts. Our results demonstrated that TRIM5α alleles, previously identified as restrictive for SIV(mac251) replication in vivo following intravenous exposure, did not affect SIV(mac251) replication following mucosal exposure, regardless of prior vaccination, challenge dose, or the presence of the protective major histocompatibility complex alleles (MamuA01(+), MamuB08(+), or MamuB017(+)). The TRIM5α genotype had no apparent effect on the number of transmitted variants or the number of challenge exposures necessary to infect the animals. DNA sequencing of the SIV(mac251) Gag gene of the two stocks used in our study revealed SIV(mac239)-like sequences that are predicted to be resistant to TRIM5α restriction. Thus, the TRIM5α genotype does not confound results of mucosal infection of rhesus macaques with SIV(mac251).     

Both mucosal and systemic routes of immunization with the live,attenuated NYVAC/simian immunodeficiency virus SIV(gpe) recombinant vaccine result in gag-specific CD8(+) T-cell responses in mucosal tissues of macaques

As most human immunodeficiency virus (HIV) infection occurs via mucosal surfaces, an important goal of vaccination may be the induction of virus-specific immune responses at mucosal sites to contain viral infection early on. Here we designed a study in macaques carrying the major histocompatibility complex class I Mamu-A(*)01 molecule to assess the capacity of the highly attenuated poxvirus NYVAC/simian immunodeficiency virus (SIV) SIV(gpe) vaccine candidate administered by the intranasal, intramuscular, or intrarectal route to induce mucosal immunity. All macaques, including one naive macaque, were exposed to SIV(mac251) by the intrarectal route and sacrificed 48 h after infection. The kinetics of immune response at various time points following immunization with NYVAC/SIV(gpe) and the anamnestic response to SIV(mac251) at 48 h after challenge were assessed in blood, in serial rectal and vaginal biopsy samples, and in tissues at euthanasia with an SIV(mac) Gag-specific tetramer. In addition, at euthanasia, antigen-specific cells producing gamma interferon or tumor necrosis factor alpha from the jejunum lamina propria were quantified in all macaques. Surprisingly, antigen-specific CD8(+) T cells were found in the mucosal tissues of all immunized macaques regardless of whether the vaccine was administered by a mucosal route (intranasal or intrarectal) or systemically. In addition, following mucosal SIV(mac251) challenge, antigen-specific responses were mainly confined to mucosal tissues, again regardless of the route of immunization. We conclude that immunization with a live vector vaccine results in the appearance of CD8(+) T-cell responses at mucosal sites even when the vaccine is delivered by nonmucosal routes.     

Early divergent host responses in SHIVsf162P3 and SIVmac251 infected macaques correlate with control of viremia

We previously showed intravaginal inoculation with SHIVsf162p3 results in transient viremia followed by undetectable viremia in most macaques, and some displayed subsequent immunity to superinfection with pathogenic SIVmac251. Here we compare early T cell activation, proliferation, and plasma cytokine/chemokine responses in macaques intravaginally infected with either SHIVsf162p3 or SIVmac251 to determine whether distinct differences in host responses may be associated with early viral containment. The data show SIVmac251 infection results in significantly higher levels of T cell activation, proliferation, and a mixed cytokine/chemokine "storm" in plasma in primary infection, whereas infection with SHIVsf162p3 resulted in significantly lower levels of T cell activation, proliferation, and better preservation of memory CD4+ T cells in early infection which immediately preceded control of viremia. These results support the hypothesis that early systemic immune activation, T cell proliferation, and a more prominent and broader array of cytokine/chemokine responses facilitate SIV replication, and may play a key role in persistence of infection, and the progression to AIDS. In contrast, immune unresponsiveness may be associated with eventual clearance of virus, a concept that may have key significance for therapy and vaccine design.     

Rapid progression is associated with lymphoid follicle dysfunction in SIV-infected infant rhesus macaques

HIV-infected infants are at an increased risk of progressing rapidly to AIDS in the first weeks of life. Here, we evaluated immunological and virological parameters in 25 SIV-infected infant rhesus macaques to understand the factors influencing a rapid disease outcome. Infant macaques were infected with SIVmac251 and monitored for 10 to 17 weeks post-infection. SIV-infected infants were divided into either typical (TypP) or rapid (RP) progressor groups based on levels of plasma anti-SIV antibody and viral load, with RP infants having low SIV-specific antibodies and high viral loads. Following SIV infection, 11 out of 25 infant macaques exhibited an RP phenotype. Interestingly, TypP had lower levels of total CD4 T cells, similar reductions in CD4/CD8 ratios and elevated activation of CD8 T cells, as measured by the levels of HLA-DR, compared to RP. Differences between the two groups were identified in other immune cell populations, including a failure to expand activated memory (CD21-CD27+) B cells in peripheral blood in RP infant macaques, as well as reduced levels of germinal center (GC) B cells and T follicular helper (Tfh) cells in spleens (4- and 10-weeks post-SIV). Reduced B cell proliferation in splenic germinal GCs was associated with increased SIV+ cell density and follicular type 1 interferon (IFN)-induced immune activation. Further analyses determined that at 2-weeks post SIV infection TypP infants exhibited elevated levels of the GC-inducing chemokine CXCL13 in plasma, as well as significantly lower levels of viral envelope diversity compared to RP infants. Our findings provide evidence that early viral and immunologic events following SIV infection contributes to impairment of B cells, Tfh cells and germinal center formation, ultimately impeding the development of SIV-specific antibody responses in rapidly progressing infant macaques.     

Cytokine expression, natural killer cell activation, and phenotypic changes in lymphoid cells from rhesus macaques during acute infection with pathogenic simian immunodeficiency virus

We studied the innate and adaptive immune system of rhesus macaques infected with the virulent simian immunodeficiency virus isolate SIVmac251 by evaluating natural killer (NK) cell activity, cytokine levels in plasma, humoral and virological parameters, and changes in the activation markers CD25 (interleukin 2R [IL-2R] α chain), CD69 (early activation marker), and CD154 (CD40 ligand) in lymphoid cells. We found that infection with SIVmac251 induced the sequential production of interferon-α/β (IFN-α/β), IL-18, and IL-12. IFN-γ, IL-4, and granulocyte-macrophage colony-stimulating factor were undetected in plasma by the assays used. NK cell activity peaked at 1 to 2 weeks postinfection and paralleled changes in viral loads. Maximum expression of CD69 on CD3⁻CD16⁺ lymphocytes correlated with NK cytotoxicity during this period. CD25 expression, which is associated with proliferation, was static or slightly down-regulated in CD4⁺ T cells from both peripheral blood (PB) and lymph nodes (LN). CD69, which is normally present in LN CD4⁺ T cells and absent in peripheral blood leukocyte (PBL) CD4⁺ T cells, was down-regulated in LN CD4⁺ T cells and up-regulated in PBL CD4⁺ T cells immediately after infection. CD8⁺ T cells increased CD69 but not CD25 expression, indicating the activation of this cellular subset in PB and LN. Finally, CD154 was transiently up-regulated in PBL CD4⁺ T cells but not in LN CD4⁺ T cells. Levels of antibodies to SIV Gag and Env did not correlate with the level of activation of CD154, a critical costimulatory molecule for T-cell-dependent immunity. In summary, we present the first documented evidence that the innate immune system of rhesus macaques recognizes SIV infection by sequential production of proinflammatory cytokines and transient activation of NK cytotoxic activity. Additionally, pathogenic SIV induces drastic changes in the level of activation markers on T cells from different anatomic compartments. These changes involve activation in the absence of proliferation, indicating that activation-induced cell death may cause some of the reported increase in lymphocyte turnover during SIV infection.The immune system of higher vertebrates consists of innate and adaptive components. Innate immunity exhibits immediate recognition and response without prior sensitization. Cells of the innate immune system (i.e., monocytes/macrophages, natural killer [NK] cells, and polymorphonuclear leukocytes) recognize pathogen-associated molecular patterns and activate events such as phagocytosis, induction of the synthesis of antimicrobial peptides, expression of inflammatory and effector cytokines and chemokines, induction of nitric oxide synthase in macrophages, and expression of costimulatory molecules on antigen-presenting cells. The adaptive immune system uses somatically generated antigen receptors that are clonally distributed on T and B lymphocytes. Generally, adaptive immune recognition in the absence of innate immune recognition results in inactivation of lymphocytes that express receptors involved in the identification events (20). Thus, innate immune responses have critical consequences in adaptive immune responses.Little is known of the contribution of the innate immune system during infection with the human immunodeficiency virus (HIV). Based on similarities of biologic and genetic features, simian immunodeficiency virus (SIV) infection of rhesus macaques provides the best animal model of HIV infection and AIDS. Accordingly, this animal model is critical for the elucidation of mechanisms of pathogenesis and for the development of vaccines and antiviral therapies (12). As with almost all viral infections, the innate immune system is thought to be the first component of the immune system that recognizes SIV infection. However, few studies have methodically analyzed the changes induced in cell phenotype and cytokine levels by SIV infection. Recent studies have demonstrated that SIV infection results in a generalized increase in lymphocyte turnover (23) and that the primary site for viral replication is activated memory CD4⁺ T cells that are present in the intestinal lamina propia (46). Although cellular changes are not that dramatic at this early stage in peripheral lymphoid tissue, peripheral blood (PB) and lymph nodes (LN) still reflect the pathologic changes induced by the viral infection and are readily available for longitudinal studies.To analyze changes in the activation state of cells from the innate and adaptive immune system after SIV infection, we evaluated NK activity, cytokine levels in plasma, and changes in activation markers on lymphoid cells of rhesus macaques after infection with pathogenic SIVmac251. We found the sequential appearance in plasma of interferon-α/β (IFN-α/β) interleukin-18 (IL-18) and IL-12, whereas IL-4, IFN-γ and granulocyte-macrophage colony-stimulating factor (GM-CSF) remained undetectable. We also found transient activation of NK cells during the peak of viral replication, and this activation was not predictive of disease progression. Finally, we observed that after SIV infection, both CD4⁺ and CD8⁺ T cells became activated in the absence of markers for proliferation, suggesting that the increased turnover of these cells reflects activation-induced cell death rather than differential compartmentalization.     

Interleukin-15 but not interleukin-7 abrogates vaccine-induced decrease in virus level in simian immunodeficiency virus mac251-infected macaques

The loss of CD4(+) T cells and the impairment of CD8(+) T cell function in HIV infection suggest that pharmacological treatment with IL-7 and IL-15, cytokines that increase the homeostatic proliferation of T cells and improve effector function, may be beneficial. However, these cytokines could also have a detrimental effect in HIV-1-infected individuals, because both cytokines increase HIV replication in vitro. We assessed the impact of IL-7 and IL-15 treatment on viral replication and the immunogenicity of live poxvirus vaccines in SIV(mac251)-infected macaques (Macaca mulatta). Neither cytokine augmented the frequency of vaccine-expanded CD4(+) or CD8(+) memory T cells, clonal recruitment to the SIV-specific CD8(+) T cell pool, or CD8(+) T cell function. Vaccination alone transiently decreased the viral set point following antiretroviral therapy suspension. IL-15 induced massive proliferation of CD4(+) effector T cells and abrogated the ability of vaccination to decrease set point viremia. In contrast, IL-7 neither augmented nor decreased the vaccine effect and was associated with a decrease in TGF-beta expression. These results underscore the importance of testing immunomodulatory approaches in vivo to assess potential risks and benefits for HIV-1-infected individuals.     

Memory CD4+ T-lymphocyte loss and dysfunction during primary simian immunodeficiency virus infection

It has long been appreciated that CD4+ T lymphocytes are dysfunctional in human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV)-infected individuals, and it has recently been shown that HIV/SIV infections are associated with a dramatic early destruction of memory CD4+ T lymphocytes. However, the relative contributions of CD4+ T-lymphocyte dysfunction and loss to immune dysregulation during primary HIV/SIV infection have not been fully elucidated. In the current study, we evaluated CD4+ T lymphocytes and their functional repertoire during primary SIVmac251 infection in rhesus monkeys. We show that the extent of loss of memory CD4+ T lymphocytes and staphylococcal enterotoxin B-stimulated cytokine production by total CD4+ T lymphocytes during primary SIVmac251 infection is tightly linked in a cohort of six rhesus monkeys to set point plasma viral RNA levels, with greater loss and dysfunction being associated with higher steady-state viral replication. Moreover, in exploring the mechanism underlying this phenomenon, we demonstrate that the loss of functional CD4+ T lymphocytes during primary SIVmac251 infection is associated with both a selective depletion of memory CD4+ T cells and a loss of the functional capacity of the memory CD4+ T lymphocytes that escape viral destruction.     

Timing of Immune Escape Linked to Success or Failure of Vaccination

Successful vaccination against HIV should limit viral replication sufficiently to prevent the emergence of viral immune escape mutations. Broadly directed immunity is likely to be required to limit opportunities for immune escape variants to flourish. We studied the emergence of an SIV Gag cytotoxic T cell immune escape variant in pigtail macaques expressing the Mane-A*10 MHC I allele using a quantitative RT-PCR to measure viral loads of escape and wild type variants. Animals receiving whole Gag expressing vaccines completely controlled an SIV_mac251 challenge, had broader CTL responses and exhibited minimal CTL escape. In contrast, animals vaccinated with only a single CTL epitope and challenged with the same SIV_mac251 stock had high levels of viral replication and rapid CTL escape. Unvaccinated naïve animals exhibited a slower emergence of immune escape variants. Thus narrowly directed vaccination against a single epitope resulted in rapid immune escape and viral levels equivalent to that of naïve unvaccinated animals. These results emphasize the importance of inducing broadly directed HIV-specific immunity that effectively quashes early viral replication and limits the generation of immune escape variants. This has important implications for the selection of HIV vaccines for expanded human trials.     

Immune modulation through 4-1BB enhances SIV vaccine protection in non-human primates against SIVmac251 challenge

Costimulatory molecules play a central role in the development of cellular immunity. Understanding how costimulatory pathways can be directed to positively influence the immune response may be critical for the generation of an effective HIV vaccine. Here, we evaluated the ability of intravenous administration of a blocking monoclonal antibody (mAb) directed against the negative costimulatory molecule CTLA-4, and an agonist mAb directed against the positive costimulatory molecule 4-1BB, either alone or in combination, to augment intramuscular SIV DNA immunizations. We then tested the ability these of these responses to impact a high-dose SIVmac251 challenge. Following immunization, the groups infused with the anti-4-1BB mAb exhibited enhanced IFN-γ responses compared to the DNA vaccine only group. Interestingly, although CTLA-4 blockade alone did not enhance IFN-γ responses it did increase the proliferative capacity of the CD4⁺ and CD8⁺ T cells. The combination of both mAbs enhanced the magnitude of the polyfunctional CD8⁺ T cell response. Following challenge, the group that received both mAbs exhibited a significant, ∼2.0 log, decrease in plasma viral load compared to the naïve group the included complete suppression of viral load in some animals. Furthermore, the use of the CTLA-4 blocking antibody resulted in significantly higher viral loads during chronic infection compared to animals that received the 4-1BB mAb, likely due to the higher CD4⁺ T cell proliferative responses which were driven by this adjuvant following immunization. These novel studies show that these adjuvants induce differential modulation of immune responses, which have dramatically different consequences for control of SIV replication, suggesting important implications for HIV vaccine development.     

CD8+ T cell dynamics during primary simian immunodeficiency virus infection in macaques: relationship of effector cell differentiation with the extent of viral replication

Immunological and virological events that occur during the earliest stages of HIV-1 infection are now considered to have a major impact on subsequent disease progression. We observed changes in the frequencies of CD8(bright) T cells expressing different chemokine receptors in the peripheral blood and lymph nodes of rhesus macaques during the acute phase of the pathogenic SIVmac251 infection; the frequency of CD8(bright) T cells expressing CXCR4 decreased, while the frequency of those expressing CCR5 increased. These reciprocal changes in chemokine receptor expression were associated with changes in the proportion of cycling (Ki67(+)) CD8(bright) T cells, and with the pattern of CD8(bright) T cell differentiation as defined by expression of CCR7 and CD45RA. In contrast, during the primary phase of the attenuated SIVmac251Deltanef infection, no major change was observed. Whereas during the acute phase of the infection with pathogenic SIV (2 wk postinfection) no correlate of disease protection was identified, once the viral load set points were established (2 mo postinfection), we found that the levels of cycling and of CCR5- and CXCR4-positive CD8(bright) T cells were correlated with the extent of viral replication and therefore with SIV-infection outcome. Our data reveal that, during primary SIV infection, despite intense CD8 T cell activation and an increase in CCR5 expression, which are considered as essential for optimal effector function of CD8(+) T cells, these changes are associated with a poor prognosis for disease progression to AIDS.     

Impairment of Gag-Specific CD8+ T-Cell Function in Mucosal and Systemic Compartments of Simian Immunodeficiency Virus mac251- and Simian-Human Immunodeficiency Virus KU2-Infected Macaques

The identification of several simian immunodeficiency virus mac251 (SIV(mac251)) cytotoxic T-lymphocyte epitopes recognized by CD8(+) T cells of infected rhesus macaques carrying the Mamu-A*01 molecule and the use of peptide-major histocompatibility complex tetrameric complexes enable the study of the frequency, breadth, functionality, and distribution of virus-specific CD8(+) T cells in the body. To begin to address these issues, we have performed a pilot study to measure the virus-specific CD8(+) and CD4(+) T-cell response in the blood, lymph nodes, spleen, and gastrointestinal lymphoid tissues of eight Mamu-A*01-positive macaques, six of those infected with SIV(mac251) and two infected with the pathogenic simian-human immunodeficiency virus KU2. We focused on the analysis of the response to peptide p11C, C-M (Gag 181), since it was predominant in most tissues of all macaques. Five macaques restricted viral replication effectively, whereas the remaining three failed to control viremia and experienced a progressive loss of CD4(+) T cells. The frequency of the Gag 181 (p11C, C-->M) immunodominant response varied among different tissues of the same animal and in the same tissues from different animals. We found that the functionality of this virus-specific CD8(+) T-cell population could not be assumed based on the ability to specifically bind to the Gag 181 tetramer, particularly in the mucosal tissues of some of the macaques infected by SIV(mac251) that were progressing to disease. Overall, the functionality of CD8(+) tetramer-binding T cells in tissues assessed by either measurement of cytolytic activity or the ability of these cells to produce gamma interferon or tumor necrosis factor alpha was low and was even lower in the mucosal tissue than in blood or spleen of some SIV(mac251)-infected animals that failed to control viremia. The data obtained in this pilot study lead to the hypothesis that disease progression may be associated with loss of virus-specific CD8(+) T-cell function.     

Virally induced CD4+ T cell depletion is not sufficient to induce AIDS in a natural host

Peripheral blood CD4+ T cell counts are a key measure for assessing disease progression and need for antiretroviral therapy in HIV-infected patients. More recently, studies have demonstrated a dramatic depletion of mucosal CD4+ T cells during acute infection that is maintained during chronic pathogenic HIV as well as SIV infection. A different clinical disease course is observed during the infection of natural hosts of SIV infection, such as sooty mangabeys (Cercocebus atys), which typically do not progress to AIDS. Previous studies have determined that SIV+ mangabeys generally maintain healthy levels of CD4+ T cells despite having viral replication comparable to HIV-infected patients. In this study, we identify the emergence of a multitropic (R5/X4/R8-using) SIV infection after 43 or 71 wk postinfection in two mangabeys that is associated with an extreme, persistent (>5.5 years), and generalized loss of CD4+ T cells (5-80 cells/microl of blood) in the absence of clinical signs of AIDS. This study demonstrates that generalized CD4+ T cell depletion from the blood and mucosal tissues is not sufficient to induce AIDS in this natural host species. Rather, AIDS pathogenesis appears to be the cumulative result of multiple aberrant immunologic parameters that include CD4+ T cell depletion, generalized immune activation, and depletion/dysfunction of non-CD4+ T cells. Therefore, these data provide a rationale for investigating multifaceted therapeutic strategies to prevent progression to AIDS, even following dramatic CD4 depletion, such that HIV+ humans can survive normal life spans analogous to what occurs naturally in SIV+ mangabeys.     

Systemic vaccination prevents the total destruction of mucosal CD4 T cells during acute SIV challenge

BACKGROUND: Acute human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infections are accompanied by a systemic loss of memory CD4 T cells, with mucosal sites serving as a major site for viral replication, dissemination and CD4 T cell depletion. Protecting the mucosal CD4 T cell compartment thus is critical to contain HIV, and preserve the integrity of the mucosal immune system. The primary objective of this study was to determine if systemic vaccination with DNA/rAd-5 encoding SIV-mac239-env, gag and pol could prevent the destruction of CD4 T cells in mucosal tissues. METHODS: Rhesus macaques were immunized with DNA/r-Ad-5 encoding SIV genes and compared with those immunized with sham vectors following high dose intravenous challenge with SIVmac251. SIV specific CD4 and CD8 T cell responses, cell associated viral loads and mucosal CD4 T cell dynamics were evaluated. RESULTS: Strong SIV specific immune responses were induced in mucosal tissues of vaccinated animals as compared with sham controls. These responses expanded rapidly following challenge suggesting a strong anamnestic response. Immune responses were associated with a decrease in cell associated viral loads, and a loss of fewer mucosal CD4 T cells. Approximately 25% of mucosal CD4 T cells were preserved in vaccinated animals as compared with <5% in sham controls. These results demonstrate that systemic immunization strategies can induce immune responses in mucosal tissues that can protect mucosal CD4 T cells from complete destruction following challenge. CONCLUSIONS: Preservation of mucosal CD4 T cells can contribute to maintaining immune competence in mucosal tissues and provide a substantial immune benefit to the vaccinees.     

Expression of CD154 by a simian immunodeficiency virus vector induces only transitory changes in rhesus macaques

Human immunodeficiency virus infection is characterized by dysregulation of antigen-presenting cell function and defects in cell-mediated immunity. Recent evidence suggests that impaired ability of CD4+ T cells to upregulate the costimulatory molecule CD154 is at the core of this dysregulation. To test the hypothesis that increased expression of CD154 on infected CD4+ T cells could modulate immune function, we constructed a replication-competent simian immunodeficiency virus (SIV) vector that expressed CD154. We found that this recombinant vector directed the expression of CD154 on the surface of infected CD4+ T cells and that expression of CD154 resulted in activation of B cells present in the same cultures. Experimental infection of rhesus macaques resulted in very low viral loads for the CD154-expressing virus and the control virus, indicating that expression of CD154 did not result in increased viral replication. Analyses of the anti-SIV immune responses and the phenotype of lymphocytes in blood and lymphoid tissues showed changes that occurred during the acute phase of infection only in animals infected with the CD154-expressing SIV, but that became indistinguishable from those seen in animals infected with the control virus at later time points. We conclude that the level of expression of CD154 in itself is not responsible for affecting the immune response to an attenuated virus. Considering that the CD154-expressing SIV vector and the virus control did not carry an active nef gene, our results suggest that, in CD4+ T cells infected with wild-type virus, Nef is the viral factor that interferes with the immune mechanisms that regulate expression of CD154.     

Depletion of CD8+ cells in sooty mangabey monkeys naturally infected with simian immunodeficiency virus reveals limited role for immune control of virus replication in a natural host species

SIV infection of sooty mangabeys (SMs), a natural host species, does not cause AIDS despite high-level virus replication. In contrast, SIV infection of nonnatural hosts such as rhesus macaques (RMs) induces an AIDS-like disease. The depletion of CD8+ T cells during SIV infection of RMs results in marked increases in plasma viremia, suggesting a key role for CD8+ T cells in controlling levels of SIV replication. To assess the role that CD8+ T cells play in determining the virologic and immunologic features of nonpathogenic SIV infection in SMs, we transiently depleted CD8+ T cells in SIV-infected and uninfected SMs using a CD8alpha-specific Ab (OKT8F) previously used in studies of SIV-infected RMs. Treatment of SMs with the OKT8F Ab resulted in the prompt and profound depletion of CD8+ T cells. However, in contrast to CD8+ cell depleted, SIV-infected RMs, only minor changes in the levels of plasma viremia were observed in most SIV-infected SMs during the period of CD8+ cell deficiency. Those SMs demonstrating greater increases in SIV replication following CD8+ cell depletion also displayed higher levels of CD4+ T cell activation and/or evidence of CMV reactivation, suggesting that an expanded target cell pool rather than the absence of CD8+ T cell control may have been primarily responsible for transient increases in viremia. These data indicate that CD8+ T cells exert a limited influence in determining the levels of SIV replication in SMs and provide additional evidence demonstrating that the absence of AIDS in SIV-infected SMs is not due to the effective control of viral replication by cellular immune responses.     

Acute loss of intestinal CD4+ T cells is not predictive of simian immunodeficiency virus virulence

The predictive value of acute gut-associated lymphoid tissue (GALT) CD4+ T cell depletion in lentiviral infections was assessed by comparing three animal models illustrative of the outcomes of SIV infection: pathogenic infection (SIVsmm infection of rhesus macaques (Rh)), persistent nonprogressive infection (SIVagm infection of African green monkeys (AGM)), and transient, controlled infection (SIVagm infection of Rh). Massive acute depletion of GALT CD4+ T cells was a common feature of acute SIV infection in all three models. The outcome of this mucosal CD4+ T cell depletion, however, differed substantially between the three models: in SIVsmm-infected Rh, the acute GALT CD4+ T cell depletion was persistent and continued with disease progression; in SIVagm, intestinal CD4+ T cells were partially restored during chronic infection in the context of normal levels of apoptosis and immune activation and absence of damage to the mucosal immunologic barrier; in SIVagm-infected Rh, complete control of viral replication resulted in restoration of the mucosal barrier and immune restoration. Therefore, our data support a revised paradigm wherein severe GALT CD4+ T cell depletion during acute pathogenic HIV and SIV infections of humans and Rh is necessary but neither sufficient nor predictive of disease progression, with levels of immune activation, proliferation and apoptosis being key factors involved in determining progression to AIDS.