[Show abstract][Hide abstract] ABSTRACT: CD16-positive monocytes that produce pro-inflammatory cytokines expand during chronic infection with the Human Immunodeficiency Virus Type 1 (HIV). The HIV-infected long lived macrophage may contribute to the establishment and maintenance of the HIV reservoir. We found that the CD14++CD16+ and CD14+CD16++ monocyte subsets are significantly expanded during infection of Rhesus macaques with pathogenic SIV mac251 but not during infection of sooty mangabeys with the non-pathogenic isolate SIVSM. In vitro glucocorticoid (GC) treatment of peripheral blood mononuclear cells (PBMC) from un-infected or SIV mac251-infected Rhesus macaques and HIV-infected patients treated or not with antiretroviral therapy (ART), resulted in a significant decrease in the frequency of both CD16-positive monocyte subsets. Short-term in vivo treatment with high doses of GC of chronically SIVmac251 infected macaques, resulted in a significant decrease in the CD14+CD16++ population and, to a lesser extent, in the CD14++CD16+ monocytes, as well as, a significant decrease in the number of macrophage in tissues. Surprisingly, treatment of SIVmac251 infected macaques with ART significantly increased the CD14++CD16+ population and the addition of GC resulted only in a significant decrease in the CD14+CD16++ subset. No difference in SIV DNA levels in blood, lymph nodes, gut, and spleen was found between the groups treated with ART or ART plus GC. Thus, it appears that high doses of GC treatment in the absence of ART could affect both CD16-positive populations in vivo. Whether the efficacy of this treatment at higher doses to decrease virus level outweighs its risks remains to be determined.
AIDS research and human retroviruses 01/2014; 31(1). DOI:10.1089/AID.2013.0220 · 2.33 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have previously shown that macaques vaccinated with DNA vectors expressing SIVmac239 antigens developed potent immune responses able to reduce viremia upon high-dose SIVmac251 challenge. To further improve vaccine-induced immunity and protection, we combined the SIVmac239 DNA vaccine with protein immunization using inactivated SIVmac239 viral particles as protein source. Twenty-six weeks after the last vaccination, the animals were challenged intrarectally at weekly intervals with a titrated dose of the heterologous SIVsmE660. Two of DNA-protein coimmunized macaques did not become infected after 14 challenges, but all controls were infected by 11 challenges. Vaccinated macaques showed modest protection from SIVsmE660 acquisition compared with naïve controls (P = 0.050; stratified for TRIM5α genotype). Vaccinees had significantly lower peak (1.6 log, P = 0.0048) and chronic phase viremia (P = 0.044), with 73% of the vaccinees suppressing viral replication to levels below assay detection during the 40-wk follow-up. Vaccine-induced immune responses associated significantly with virus control: binding antibody titers and the presence of rectal IgG to SIVsmE660 Env correlated with delayed SIVsmE660 acquisition; SIV-specific cytotoxic T cells, prechallenge CD4(+) effector memory, and postchallenge CD8(+) transitional memory cells correlated with control of viremia. Thus, SIVmac239 DNA and protein-based vaccine protocols were able to achieve high, persistent, broad, and effective cellular and humoral immune responses able to delay heterologous SIVsmE660 infection and to provide long-term control of viremia. These studies support a role of DNA and protein-based vaccines for development of an efficacious HIV/AIDS vaccine.
Proceedings of the National Academy of Sciences 01/2013; 110(8). DOI:10.1073/pnas.1215393110 · 9.67 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Human immunodeficiency virus (HIV) and Simian immunodeficiency virus (SIV) disease progression is associated with multifocal damage to the gastrointestinal tract epithelial barrier that correlates with microbial translocation and persistent pathological immune activation, but the underlying mechanisms remain unclear. Investigating alterations in mucosal immunity during SIV infection, we found that damage to the colonic epithelial barrier was associated with loss of multiple lineages of interleukin (IL)-17-producing lymphocytes, cells that microarray analysis showed expressed genes important for enterocyte homeostasis, including IL-22. IL-22-producing lymphocytes were also lost after SIV infection. Potentially explaining coordinate loss of these distinct populations, we also observed loss of CD103+ dendritic cells (DCs) after SIV infection, which associated with the loss of IL-17- and IL-22-producing lymphocytes. CD103+ DCs expressed genes associated with promotion of IL-17/IL-22+ cells, and coculture of CD103+ DCs and naïve T cells led to increased IL17A and RORc expression in differentiating T cells. These results reveal complex interactions between mucosal immune cell subsets providing potential mechanistic insights into mechanisms of mucosal immune dysregulation during HIV/SIV infection, and offer hints for development of novel therapeutic strategies to address this aspect of AIDS virus pathogenesis.
[Show abstract][Hide abstract] ABSTRACT: Viral protein X (Vpx) of SIV has been reported to be important for establishing infection in vivo. Vpx has several different activities in vitro, promoting preintegration complex import into the nucleus in quiescent lymphocytes and overcoming a block in reverse transcription in macrophages. Vpx interacts with the DDB1-CUL4-DCAF1 E3 ligase complex, which may or may not be required for the ascribed functions. The goal of the current study was to determine whether these activities of Vpx are important in vivo.
An infectious, pathogenic clone of SIVmne was used to examine correlations between Vpx functions in vitro and in vivo. Three previously described HIV-2 Vpx mutants that were shown to be important for nuclear import of the preintegration complex in quiescent lymphocytes were constructed in SIVmne: A vpx-deleted virus, a truncation of Vpx at amino acid 102 that deletes the C-terminal proline-rich domain (X(102)), and a mutant with tyrosines 66, 69, and 71 changed to alanine (X(y-a)). All mutant viruses replicated similarly to wild type SIVmne027 in primary pigtail macaque PBMCs, and were only slightly retarded in CEMx174 cells. However, all the vpx mutant viruses were defective for replication in both human and pigtail monocyte-derived macrophages. PCR assays demonstrated that the efficiency of reverse transcription and the levels of viral integration in macrophages were substantially reduced for the vpx mutant viruses. In vitro, the X(y-a) mutant, but not the X(102) mutant lost interaction with DCAF1. The wild type SIVmne027 and the three vpx mutant SIVs were inoculated by the intra-rectal route into pigtail macaques. Peak levels of plasma viremia of the vpx mutant SIVs were variable, but consistently lower than that observed in macaques infected with wild type SIVmne. In situ hybridization for SIV demonstrated that compared to wild type SIVmne infected macaques five of the six animals inoculated with the vpx mutant SIVs had only low levels of SIV-expressing cells in the rectum, most intestinal epithelial tissues, spleen, and mesenteric and peripheral nodes.
This work demonstrates that the activities of Vpx to overcome restrictions in culture in vitro are also likely to be important for establishment of infection in vivo and suggest that both the nuclear localization and DCAF1-interaction functions of Vpx are critical in vivo.
[Show abstract][Hide abstract] ABSTRACT: APOBEC3G (A3G) is a host cytidine deaminase that inhibits retroviruses. HIV and related primate lentiviruses encode Vif, which counteracts A3G by inducing its degradation. This Vif-mediated A3G inhibition is species specific, suggesting that the A3G-Vif interaction has evolved as primate lentiviruses have adapted to their hosts. We examined the evolutionary dynamics of the A3G-Vif interaction within four African green monkey (AGM) subspecies, which are each naturally infected with a distinct simian immunodeficiency virus (SIV). We identified single amino acid changes within A3G in two AGM subspecies that render it resistant to Vif proteins, except for Vif from the viruses that naturally infect these subspecies. Moreover, experimental infection of AGMs shows that Vif can rapidly adapt to these arising Vif-resistant A3G genotypes. These data suggest that despite being generally nonpathogenic in its natural host, SIV infection selects for Vif-resistant forms of A3G in AGM populations, driving Vif counterevolution and functional divergence.
[Show abstract][Hide abstract] ABSTRACT: The simian immunodeficiency viruses (SIVs) are a diverse group of viruses that naturally infect a wide range of African primates, including African green monkeys (AGMs) and sooty mangabey monkeys (SMs). Although natural infection is widespread in feral populations of AGMs and SMs, this infection generally does not result in immunodeficiency. However, experimental inoculation of Asian macaques results in an immunodeficiency syndrome remarkably similar to human AIDS. Thus, natural nonprogressive SIV infections appear to represent an evolutionary adaptation between these animals and their primate lentiviruses. Curiously, these animals maintain robust virus replication but have evolved strategies to avoid disease progression. Adaptations observed in these primates include phenotypic changes to CD4(+) T cells, limited chronic immune activation, and altered mucosal immunity. It is probable that these animals have achieved a unique balance between T-cell renewal and proliferation and loss through activation-induced apoptosis, and virus-induced cell death. A clearer understanding of the mechanisms underlying the lack of disease progression in natural hosts for SIV infection should therefore yield insights into the pathogenesis of AIDS and may inform vaccine design.
Cold Spring Harbor Perspectives in Medicine 01/2012; 2(1):a007153. DOI:10.1101/cshperspect.a007153 · 9.47 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: 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).
Journal of Virology 09/2011; 85(23):12399-409. DOI:10.1128/JVI.05707-11 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A simian immunodeficiency virus (SIV) vaccine coexpressing granulocyte-macrophage colony stimulating factor (GM-CSF) prevented infection in 71% of macaques that received 12 rectal challenges. The SIVsmE660 challenge had the tropism of incident human immunodeficiency virus (HIV) infections and a similar genetic distance from the SIV239 vaccine as intraclade HIV isolates. The heterologous prime-boost vaccine regimen used recombinant DNA for priming and recombinant modified vaccinia Ankara for boosting. Co-expression of GM-CSF in the DNA prime enhanced the avidity of elicited immunoglobulin G for SIV envelope glycoproteins, the titers of neutralizing antibody for easy-to-neutralize SIV isolates, and antibody-dependent cellular cytotoxicity. Impressively, the co-expressed GM-CSF increased vaccine-induced prevention of infection from 25% in the non-GM-CSF co-expressing vaccine group to 71% in the GM-CSF co-expressing vaccine group. The prevention of infection showed a strong correlation with the avidity of the elicited Env-specific antibody for the Env of the SIVsmE660 challenge virus (r = 0.9; P < .0001).
The Journal of Infectious Diseases 07/2011; 204(1):164-73. DOI:10.1093/infdis/jir199 · 6.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Three Indian rhesus macaques, Ad-SIV primed/protein boosted and exposed twice to high-dose mucosal SIV(mac251) challenges, exhibited elite control of viremia over 6.5 years. They were negative for host factors associated with control of SIV infection. After a third intrarectal challenge with SIV(smE660), all controlled viremia, with one (macaque #5) maintaining undetectable viremia in blood. Acquisition was not blocked, but virus was contained in the jejunum and draining lymph nodes. Polyfunctional memory T cell responses and high-titered neutralizing and non-neutralizing serum and mucosal antibodies were present before and maintained post-challenge. The level of protection seen for animal #5 was predicted from analyses of gene transcription in jejunum 2 weeks post-challenge. Macaques #7 and #9, exhibiting lower pre-challenge cellular and humoral immunity, partially controlled the SIV(smE660) challenge. Initial vaccine-induced control by macaque #5 extended to the SIV(smE660) challenge due to multiple immune mechanisms that were boosted and augmented by cryptic SIV exposure.
[Show abstract][Hide abstract] ABSTRACT: Homologous SIVsm stocks were generated by passage of a macaque isolate of SIVsm (SIVsm/E660) in human CEM x 174 cells or macaque peripheral blood mononuclear cells. Macaques were immunized with whole inactivated SIV vaccine consisting of virus generated by transfection of CEM x 174 cells with the SIVsmH4 clone and were challenged with either cell-free stock. Only vaccinees challenged with virus generated in human cells were protected from infection. This confirms the species-specificity of whole inactivated vaccine-mediated protection.
Journal of Medical Primatology 01/2011; 23(2-3):75-82. DOI:10.1111/j.1600-0684.1994.tb00105.x · 0.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Patas monkeys were not reported to carry species‐specific simian immunodeficiency virus (SIV), but cross‐species transmission of SIVagm to patas monkeys occurred in the wild. We report that patas monkeys share immunophenotypic features with natural hosts of SIV; that is, low levels of CD4+ T cells and low CCR5 expression on CD4+ T cells. In 1 patas monkey with undetectable levels of CD4+ T cells, experimental exposure to SIVagm did not result in infection. The other experimentally infected patas monkeys showed an infection pattern similar to SIV infection in natural hosts. Thus, down‐regulation of CD4 and CCR5 expression on CD4+ T cells may effectively control human immunodeficiency virus acquisition and result in SIV extinction.
The Journal of Infectious Diseases 11/2010; 202 Suppl 3(S3):S371-6. DOI:10.1086/655970 · 6.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: APOBEC3F (A3F) is a member of the family of cytidine deaminases that is often coexpressed with APOBEC3G (A3G) in cells susceptible
to HIV infection. A3F has been shown to have strong antiviral activity in transient-expression studies, and together with
A3G, it is considered the most potent cytidine deaminase targeting HIV. Previous analyses suggested that the antiviral properties
of A3F can be dissociated from its catalytic deaminase activity. We were able to confirm the deaminase-independent antiviral
activity of exogenously expressed A3F; however, we also noted that exogenous expression was associated with very high A3F
mRNA and protein levels. In analogy to our previous study of A3G, we produced stable HeLa cell lines constitutively expressing
wild-type or deaminase-defective A3F at levels that were more in line with the levels of endogenous A3F in H9 cells. A3F expressed
in stable HeLa cells was packaged into Vif-deficient viral particles with an efficiency similar to that of A3G and was properly
targeted to the viral nucleoprotein complex. Surprisingly, however, neither wild-type nor deaminase-defective A3F inhibited
HIV-1 infectivity. These results imply that the antiviral activity of endogenous A3F is negligible compared to that of A3G.
Journal of Virology 11/2010; 84(21):11067-75. DOI:10.1128/JVI.01249-10 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The mechanisms underlying the AIDS resistance of natural hosts for simian immunodeficiency virus (SIV) remain unknown. Recently,
it was proposed that natural SIV hosts avoid disease because their plasmacytoid dendritic cells (pDCs) are intrinsically unable
to produce alpha interferon (IFN-α) in response to SIV RNA stimulation. However, here we show that (i) acute SIV infections
of natural hosts are associated with a rapid and robust type I IFN response in vivo, (ii) pDCs are the principal in vivo producers of IFN-α/β at peak acute infection in lymphatic tissues, and (iii) natural SIV hosts downregulate these responses
in early chronic infection. In contrast, persistently high type I IFN responses are observed during pathogenic SIV infection
of rhesus macaques.
Journal of Virology 08/2010; 84(15):7886-91. DOI:10.1128/JVI.02612-09 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: HIV vaccine development has been hampered by issues such as undefined correlates of protection and extensive diversity of HIV. We addressed these issues using a previously established SIV-macaque model in which SIV mutants with deletions of multiple gp120 N-glycans function as potent live attenuated vaccines to induce near-sterile immunity against the parental pathogenic SIVmac239. In this study, we investigated the protective efficacy of these mutants against a highly pathogenic heterologous SIVsmE543-3 delivered intravenously to rhesus macaques with diverse MHC genotypes. All 11 vaccinated macaques contained the acute-phase infection with blood viral loads below the level of detection between 4 and 10 weeks postchallenge (pc), following a transient but marginal peak of viral replication at 2 weeks in only half of the challenged animals. In the chronic phase, seven vaccinees contained viral replication for over 80 weeks pc, while four did not. Neutralizing antibodies against challenge virus were not detected. Although overall levels of SIV specific T cell responses did not correlate with containment of acute and chronic viral replication, a critical role of cellular responses in the containment of viral replication was suggested. Emergence of viruses with altered fitness due to recombination between the vaccine and challenge viruses and increased gp120 glycosylation was linked to the failure to control SIV. These results demonstrate the induction of effective protective immune responses in a significant number of animals against heterologous virus by infection with deglycosylated attenuated SIV mutants in macaques with highly diverse MHC background. These findings suggest that broad HIV cross clade protection is possible, even in hosts with diverse genetic backgrounds. In summary, results of this study indicate that deglycosylated live-attenuated vaccines may provide a platform for the elucidation of correlates of protection needed for a successful HIV vaccine against diverse isolates.
PLoS ONE 07/2010; 5(7):e11678. DOI:10.1371/journal.pone.0011678 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To define potential common features of simian immunodeficiency virus (SIV) infections in different naturally infected host species, we compared the dynamics of viral replication in 31 African green monkeys (10 sabeus, 15 vervets and seven Caribbean AGMs), 14 mandrills and three sooty mangabeys (SMs) that were experimentally infected with their species-specific viruses.
After infection, these SIVs replicated rapidly reaching viral loads (VLs) of 10(5)-10(9) copies/ml of plasma between days 9-14 post-infection (p.i). Set point viremia was established between days 42 and 60 p.i., with levels of approximately 10(5)-10(6) copies/ml in SM and mandrills, and lower levels (10(3)-10(5) copies/ml) in AGMs. VL during the chronic phase did not correlate with viral genome structure: SIVmnd-2 (a vpx-containing virus) and SIVmnd-1 (which does not contain vpu or vpx) replicated to similar levels in mandrills. VL was dependent on virus strain: vervets infected with three different viral strains showed different patterns of viral replication. The pattern of viral replication of SIVagm.sab, which uses both CCR5 and CXCR4 co-receptors was similar to those of the other viruses.
Our results show a common pattern of SIV replication in naturally and experimentally infected hosts. This is similar overall to that observed in pathogenic SIV infection of macaques. This result indicates that differences in clinical outcome between pathogenic and non-pathogenic infections rely on host responses rather than the characteristics of the virus itself.
Journal of Medical Primatology 09/2006; 35(4-5):194-201. DOI:10.1111/j.1600-0684.2006.00168.x · 0.82 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have previously described two isogenic molecularly cloned simian immunodeficiency virus/human immunodeficiency virus chimeric
viruses (SHIVs) that differ from one another by 9 amino acids and direct distinct clinical outcomes in inoculated rhesus monkeys.
SHIVDH12R-Clone 7, like other highly pathogenic CXCR4-tropic SHIVs, induces rapid and complete depletions of CD4+ T lymphocytes and immunodeficiency in infected animals. In contrast, macaques inoculated with SHIVDH12R-Clone 8 experience only partial and transient losses of CD4+ T cells, show prompt control of their viremia, and remain healthy for periods of time extending for up to 4 years. The contributions
of CD8+ and CD20+ lymphocytes in suppressing the replication of the attenuated SHIVDH12R-Clone 8 and maintaining a prolonged asymptomatic clinical course was assessed by treating animals with monoclonal antibodies that
deplete each lymphocyte subset at the time of virus inoculation. The absence of either CD8+ or CD20+ cells during the SHIVDH12R-Clone 8 acute infection resulted in the rapid, complete, and irreversible loss of CD4+ T cells; sustained high levels of postpeak plasma viremia; and symptomatic disease in Mamu-A*01-negative Indian rhesus monkeys.
In Mamu-A*01-positive animals, however, the aggressive, highly pathogenic phenotype was observed only in macaques depleted
of CD8+ cells; SHIVDH12R-Clone 8 was effectively controlled in Mamu-A*01-positive monkeys in the absence of B lymphocytes. Taken together, these results indicate
that both CD8+ and CD20+ B cells contribute to the control of primate lentiviral infection in Mamu-A*01-negative macaques. Furthermore, the major
histocompatibility complex genotype of an infected animal, as exemplified by the Mamu-A*01 allele in this study, has the additional
capacity to shift the balance of the composite immune response.
Journal of Virology 01/2006; 79(23):14887-98. DOI:10.1128/JVI.79.23.14887-14898.2005 · 4.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have evaluated the homologous and heterologous neutralizing antibody response in a cohort of six Macaca nemestrina infected with the cloned virus SIVsm62d that showed different levels of envelope diversification. Two progressor macaques developed AIDS by 1.5 years post-inoculation and four non-progressors were asymptomatic for 3 years of follow-up. All macaques developed high titers of neutralizing antibodies against homologous SIVsm viruses and intermediate titers against SIVsmB670. Heterologous virus neutralization of SIVmac, SIVmne, and HIV-2 was detected at much lower levels in both progressor macaques; only one of four non-progressors had evidence for broader neutralizing antibody activity. We noted changes in potential N-linked glycosylation (PNG) sites in V1/V2, C2, and V4 that were common to multiple macaques. These results support a model for viral neutralization where heterologous neutralization is, in part, driven by a strong homologous response and may be coupled to changes in PNG sites in envelope.
Journal of Medical Primatology 11/2005; 34(5-6):253-61. DOI:10.1111/j.1600-0684.2005.00123.x · 0.82 Impact Factor