Simian Immunodeficiency Virus Replication Research Articles

Lymph-Node-Based CD3+ CD20+ Cells Emerge from Membrane Exchange between T Follicular Helper Cells and B Cells and Increase Their Frequency following Simian Immunodeficiency Virus Infection.

CD4+ T follicular helper (TFH) cells are key targets for human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication and contribute to the virus reservoir under antiretroviral therapy (ART). Here, we describe a novel CD3+ CD20+ double-positive (DP) lymphocyte subset, resident in secondary lymphoid organs of humans and rhesus macaques (RMs), that appear predominantly after membrane exchange between TFH and B cells. DP lymphocytes are enriched in cells displaying a TFH phenotype (CD4+ PD1hi CXCR5hi), function (interleukin 21 positive [IL-21+]), and gene expression profile. Importantly, expression of CD40L upon brief in vitro mitogen stimulation identifies, by specific gene-expression signatures, DP cells of TFH-cell origin versus those of B-cell origin. Analysis of 56 RMs showed that DP cells (i) significantly increase following SIV infection, (ii) are reduced after 12 months of ART in comparison to pre-ART levels, and (iii) expand to a significantly higher frequency following ART interruption. Quantification of total SIV-gag DNA on sorted DP cells from chronically infected RMs showed that these cells are susceptible to SIV infection. These data reinforce earlier observations that CD20+ T cells are infected and expanded by HIV infection, while suggesting that these cells phenotypically overlap activated CD4+ TFH cells that acquire CD20 expression via trogocytosis and can be targeted as part of therapeutic strategies aimed at HIV remission. IMPORTANCE The HIV reservoir is largely composed of latently infected memory CD4+ T cells that persist during antiretroviral therapy and constitute a major barrier toward HIV eradication. In particular, CD4+ T follicular helper cells have been demonstrated as key targets for viral replication and persistence under ART. In lymph nodes from HIV-infected humans and SIV-infected rhesus macaques, we show that CD3+ CD20+ lymphocytes emerge after membrane exchange between T cells and B cells and are enriched in phenotypic, functional, and gene expression profiles found in T follicular helper cells. Furthermore, in SIV-infected rhesus macaques, these cells expand following experimental infection and after interruption of ART and harbor SIV DNA at levels similar to those found in CD4+ T cells; thus, CD3+ CD20+ lymphocytes are susceptible to SIV infection and can contribute to SIV persistence.

Natural Killer Cells Regulate Acute SIV Replication, Dissemination, and Inflammation, but Do Not Impact Independent Transmission Events.

Natural killer (NK) cells are potent effector cells of the innate immune system possessing both cytotoxic and immunoregulatory capabilities, which contribute to their crucial role in controlling human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) infections. However, despite significant evidence for NK cell modulation of HIV disease, their specific contribution to transmission and control of acute infection remains less clear. To elucidate the contribution of NK cells during acute SIV infection, we performed an acute necropsy study, where rhesus macaques (RM) were subjected to preinfection depletion of systemic NK cells using established methods of IL-15 neutralization, followed by subsequent challenge with barcoded SIVmac239X. Our study showed that depletion was highly effective, resulting in near total ablation of all NK cell subsets in blood, liver, oral, and rectal mucosae, and lymph nodes (LN) that persisted through the duration of the study. Meanwhile, frequencies and phenotypes of T cells remained virtually unchanged, indicating that our method of NK cell depletion had minimal off-target effects. Importantly, NK cell-depleted RM demonstrated an early and sustained 1 to 2 log increase in viremia over controls, but sequence analysis suggested no difference in the number of independent transmission events. Acute bulk, central memory (CM), and CCR5+ CD4+ T cell depletion was similar between experimental and control groups, while CD8+ T cell activation was higher in NK cell-depleted RM as measured by Ki67 and PD-1 expression. Using 27-plex Luminex analyses, we also found modestly increased inflammatory cytokines in NK cell-depleted RM compared to control animals. In the effort to determine the impact of NK cells on HIV/SIV transmission and acute viremia, future studies will be necessary to better harness these cells for future viral therapies. Collectively, these data suggest NK cells are important modulators of lentivirus dissemination and disease but may not have the capacity to independently eliminate individual transmission events. IMPORTANCE Natural killer (NK) cells as major effector cells of the innate immune system can contribute significantly to human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) control. However, a specific role for NK cells in blocking lentivirus transmission remains incompletely clear. In this study, we depleted NK cells prior to challenge with a barcoded SIV. Importantly, our studied showed systemic NK cell depletion was associated with a significant increase in acute viremia, but did not impact the number of independent transmission events. Collectively, these data suggest NK cells are critical modulators of early lentivirus replication but may not regulate individual transmission events at mucosal portals of entry.

Cytomegalovirus-vaccine-induced unconventional T cell priming and control of SIV replication is conserved between primate species

Strain 68-1 rhesus cytomegalovirus expressing simian immunodeficiency virus (SIV) antigens (RhCMV/SIV) primes MHC-E-restricted CD8+ Tcells that control SIV replication in 50%-60% of the vaccinated rhesus macaques. Whether this unconventional SIV-specific immunity and protection is unique to rhesus macaques or RhCMV or is intrinsic to CMV remains unknown. Here, using cynomolgus CMV vectors expressing SIV antigens (CyCMV/SIV) and Mauritian cynomolgus macaques, we demonstrate that the induction of MHC-E-restricted CD8+ Tcells requires matching CMV to its host species. RhCMV does not elicit MHC-E-restricted CD8+ Tcells in cynomolgus macaques. However, cynomolgus macaques vaccinated with species-matched 68-1-like CyCMV/SIV mounted MHC-E-restricted CD8+ Tcells, and half of the vaccinees stringently controlled SIV post-challenge. Protected animals manifested a vaccine-induced IL-15 transcriptomic signature that is associated with efficacy in rhesus macaques. These findings demonstrate that the ability of species-matched CMV vectors to elicit MHC-E-restricted CD8+ Tcells that are required for anti-SIV efficacy is conserved in nonhuman primates, and these data support the development of HCMV/HIV for a prophylactic HIV vaccine.

Spontaneous Control of SIV Replication Does Not Prevent T Cell Dysregulation and Bacterial Dissemination in Animals Co-Infected with M. tuberculosis.

ABSTRACTIndividuals co-infected with HIV and Mycobacterium tuberculosis (Mtb) are more likely to develop severe tuberculosis (TB) disease than HIV-naive individuals. To understand how a chronic pre-existing Simian immunodeficiency virus (SIV) infection impairs the early immune response to Mtb, we used the Mauritian cynomolgus macaque (MCM) model of SIV/Mtb co-infection. We examined the relationship between peripheral viral control and Mtb burden, Mtb dissemination, and T cell function between SIV+ spontaneous controllers, SIV+ non-controllers, and SIV-naive MCM who were challenged with a barcoded Mtb Erdman strain 6 months post-SIV infection and necropsied 6 weeks post-Mtb infection. Mycobacterial burden was highest in the SIV+ non-controllers in all assessed tissues. In lung granulomas, the frequency of TNF-α-producing CD4+ T cells was reduced in all SIV+ MCM, but IFNγ-producing CD4+ T cells were only lower in the SIV+ non-controllers. Further, while all SIV+ MCM had more PD1+ and TIGIT+ T cells in the lung granulomas relative to SIV-naive MCM, SIV+ controllers exhibited the highest frequency of cells expressing these markers. To measure the effect of SIV infection on within-host bacterial dissemination, we sequenced the molecular barcodes of Mtb present in each tissue and characterized the Mtb population complexity. While Mtb population complexity was not associated with SIV infection group, lymph nodes had increased complexity when compared with lung granulomas across all groups. These results provide evidence that SIV+ animals, independent of viral control, exhibit a dysregulated T cell immune response and enhanced dissemination of Mtb, likely contributing to the poor TB disease course across all SIV/Mtb co-infected animals.IMPORTANCE HIV and TB remain significant global health issues, despite the availability of treatments. Individuals with HIV, including those who are virally suppressed, are at an increased risk to develop and succumb to severe TB disease when compared with HIV-naive individuals. Our study aims to understand the relationship between the extent of SIV replication, mycobacterial growth, and T cell function in the tissues of co-infected Mauritian cynomolgus macaques during the first 6 weeks of Mtb infection. Here we demonstrate that increased viral replication is associated with increased bacterial burden in the tissues and impaired T cell responses, and that the immunological damage attributed to virus infection is not fully eliminated when animals spontaneously control virus replication.

Mutation in the Disordered Linker Region of Capsid Disrupts Viral Kinetics of a Neuropathogenic SIV in Rhesus Macaques.

ABSTRACTTRIM5α polymorphism in rhesus macaques (RM) limits the genetic pool of animals in which we can perform simian immunodeficiency virus (SIV) studies without first screening animals for permissive TRIM5α genotypes. We have previously shown that polymorphisms in the TRIM5α B30.2/SPRY domain impact the level of SIVsmm viremia in RM and that amino acid substitutions (P37S/R98S) in the capsid N-terminal domain (CA-NTD) enables the virus to overcome restriction in RMs with the restrictive homozygous TRIM5αTFP/TFP genotype. Since this genotype also negatively impacted the development of central nervous system (CNS) lesions in animals infected with the parental source of CL757, we sought to generate a TRIM5αTFP/TFP-resistant clone, SIV-804E-CL757-P37S/R98S (CL757-SS), using a similar strategy. Unexpectedly, viral replication of CL757-SS was impaired in RMs with either the permissive TRIM5αTFP/Q or the restrictive TRIM5αTFP/TFP genotype. Analysis of the virus which emerged in the latter animals led to the discovery of a preexisting mutation relative to other SIVs. This P146T substitution in a conserved disordered linker region in the C-terminal domain of capsid (CA-CTD) has been shown to inhibit proper formation of HIV-1 capsid particles. Restoration of this residue to proline in the context of the TRIM5α-SS escape mutations not only restored viral replication, but also enhanced the infectivity of our previously reported neurotropic clone, even in RMs with permissive TRIM5α genotypes.IMPORTANCE SIV infection of rhesus macaques has become a valuable model for the development of AIDS vaccines and antiretroviral therapies. Polymorphisms in the rhesus macaque TRIM5α gene can affect SIV replication, making it necessary to genetically screen macaques for TRIM5α alleles that are permissive for SIV replication. This limits the pool of animals that can be used in a study, thereby making the acquisition of animals needed to fulfill study parameters difficult. We have constructed a viral clone that induces neuroAIDS in rhesus macaques regardless of their TRIM5α genotype, while also highlighting the important role the disordered linker domain plays in viral infectivity.

SIV Evolutionary Dynamics in Cynomolgus Macaques during SIV-Mycobacterium tuberculosis Co-Infection.

Co-infection with Mycobacterium tuberculosis (Mtb) and human immunodeficiency virus (HIV) is a worldwide public health concern, leading to worse clinical outcomes caused by both pathogens. We used a non-human primate model of simian immunodeficiency virus (SIV)-Mtb co-infection, in which latent Mtb infection was established prior to SIVmac251 infection. The evolutionary dynamics of SIV env was evaluated from samples in plasma, lymph nodes, and lungs (including granulomas) of SIV-Mtb co-infected and SIV only control animals. While the diversity of the challenge virus was low and overall viral diversity remained relatively low over 6–9 weeks, changes in viral diversity and divergence were observed, including evidence for tissue compartmentalization. Overall, viral diversity was highest in SIV-Mtb animals that did not develop clinical Mtb reactivation compared to animals with Mtb reactivation. Among lung granulomas, viral diversity was positively correlated with the frequency of CD4+ T cells and negatively correlated with the frequency of CD8+ T cells. SIV diversity was highest in the thoracic lymph nodes compared to other sites, suggesting that lymphatic drainage from the lungs in co-infected animals provides an advantageous environment for SIV replication. This is the first assessment of SIV diversity across tissue compartments during SIV-Mtb co-infection after established Mtb latency.

A Stretch of Unpaired Purines in the Leader Region of Simian Immunodeficiency Virus (SIV) Genomic RNA is Critical for its Packaging into Virions

Simian immunodeficiency virus (SIV) is an important lentivirus used as a non-human primate model to study HIV replication, and pathogenesis of human AIDS, as well as a potential vector for human gene therapy. This study investigated the role of single-stranded purines (ssPurines) as potential genomic RNA (gRNA) packaging determinants in SIV replication. Similar ssPurines have been implicated as important motifs for gRNA packaging in many retroviruses like, HIV-1, MPMV, and MMTV by serving as Gag binding sites during virion assembly. In examining the secondary structure of the SIV 5′ leader region, as recently deduced using SHAPE methodology, we identified four specific stretches of ssPurines (I-IV) in the region that harbors major packaging determinants of SIV. The significance of these ssPurine motifs were investigated by mutational analysis coupled with a biologically relevant single round of replication assay. These analyses revealed that while ssPurine II was essential, the others (ssPurines I, III, & IV) did not significantly contribute to SIV gRNA packaging. Any mutation in the ssPurine II, such as its deletion or substitution, or other mutations that caused base pairing of ssPurine II loop resulted in near abrogation of RNA packaging, further substantiating the crucial role of ssPurine II and its looped conformation in SIV gRNA packaging. Structure prediction analysis of these mutants further corroborated the biological results and further revealed that the unpaired nature of ssPurine II is critical for its function during SIV RNA packaging perhaps by enabling it to function as a specific binding site for SIV Gag.

Two Human Monoclonal HLA-Reactive Antibodies Cross-React with Mamu-B*008, a Rhesus Macaque MHC Allotype Associated with Control of Simian Immunodeficiency Virus Replication.

MHC class I molecules play an important role in adaptive immune responses against intracellular pathogens. These molecules are highly polymorphic, and many allotypes have been characterized. In a transplantation setting, a mismatch between MHC allotypes may initiate an alloimmune response. Rhesus macaques (Macaca mulatta, Mamu) are valuable as a preclinical model species in transplantation research as well as to evaluate the safety and efficacy of vaccine candidates. In both lines of research, the availability of nonhuman primate MHC-reactive mAbs may enable in vitro monitoring and detection of presence of particular Mamu molecules. In this study, we screened a collection of thoroughly characterized HLA class I-specific human mAbs for cross-reactivity with rhesus macaque MHC class I allotypes. Two mAbs, OK4F9 and OK4F10, recognize an epitope that is defined by isoleucine (I) at amino acid position 142 that is present on the Indian rhesus macaque Mamu-B*008:01 allotype, which is an allotype known to be associated with elite control of SIV replication. The reactive pattern of a third mAb, MUS4H4, is more complex and includes an epitope shared on Mamu-A2*05:01 and -B*001:01-encoded Ags. This is the first description, to our knowledge, of human HLA-reactive mAbs that can recognize Mamu allotypes, and these can be useful tools for in vitro monitoring the presence of the relevant allelic products. Moreover, OK4F9 and OK4F10 can be powerful mAbs for application in SIV-related research.

Selective Disruption of SERINC5 Antagonism by Nef Impairs SIV Replication in Primary CD4+ T Cells.

The Nef proteins of HIV-1 and SIV enhance viral infectivity by preventing the incorporation of the multipass transmembrane protein serine incorporator 5 (SERINC5), and to a lesser extent SERINC3, into virions. In addition to counteracting SERINCs, SIV Nef also downmodulates several transmembrane proteins from the surface of virus-infected cells, including simian tetherin, CD4 and MHC class I (MHC I) molecules. From a systematic analysis of alanine substitutions throughout the SIVmac239 Nef protein, we identified residues that are required to counteract SERINC5. This information was used to engineer an infectious molecular clone of SIV (SIVmac239nef AV), which differs by two amino acids in the N-terminal domain of Nef that make the virus sensitive to SERINC5 while retaining other activities of Nef. SIVmac239nef AV downmodulates CD3, CD4, MHC I and simian tetherin, but cannot counteract SERINC5. In primary rhesus macaque CD4+ T cells, SIVmac239nef AV exhibits impaired infectivity and replication compared to wild-type SIVmac239. These results demonstrate that SERINC5 antagonism can be separated from other Nef functions and reveal the impact of SERINC5 on lentiviral replication.Importance: SERINC5, a multipass transmembrane protein, is incorporated into retroviral particles during assembly. This leads to a reduction of particle infectivity by inhibiting virus fusion with the target cell membrane. The Nef proteins of HIV-1 and SIV enhance viral infectivity by preventing the incorporation of SERINC5 into virions. However, the relevance of this restriction factor in viral replication has not been elucidated. Here we report a systematic mapping of Nef residues required for SERINC5 antagonism. Counter screens for three other functions of Nef helped identify two residues in the N-terminal domain of Nef, which when mutated make Nef selectively susceptible to SERINC5. Since Nef is multi-functional, genetic separation of SERINC5 antagonism from its other functions affords comparison of the replication of isogenic viruses that are or are not sensitive to SERINC5. Such a strategy revealed the impact of SERINC5 on SIV replication in primary rhesus macaque CD4+ T-cells.

In vivo protective mechanisms of neutralizing antibodies against simian immunodeficiency virus replicatio

Identifying protective adaptive immune responses against human immunodeficiency virus type 1 (HIV-1), mainly comprising CD8+ cytotoxic T lymphocyte (CTL) and neutralizing antibody (NAb) responses, is crucial for understanding in vivo mechanisms of viral persistence and developing prophylactic/intervention strategies. In HIV-1 and pathogenic simian immunodeficiency virus (SIV) infections, CTL responses play the canonical role in primary viral replication control, whereas NAb responses are impaired. This NAb impairment in early infection conversely highlights the necessity of elucidating anti-HIV/SIV antibody defense/induction mechanisms, and one approach to analyze the impact of NAbs on HIV/SIV infection is passive immunization. We have analyzed a simian AIDS model of highly pathogenic SIVmac239-infected rhesus macaques, and characterized that a single acute-phase passive infusion of SIV-specific polyclonal NAbs drives a synergistic qualitative boosting of virus-specific T-cell responses, resulting in sustained SIV replication control. This in vivo functional augmentation of virus-specific T cells by NAbs in the SIV model provides insights into the design of protective immunity against HIV-1 infection.

Innate, non-cytolytic CD8+ T cell-mediated suppression of HIV replication by MHC-independent inhibition of virus transcription.

MHC-I-restricted, virus-specific cytotoxic CD8+ T cells (CTLs) may control human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) replication via the recognition and killing of productively infected CD4+ T cells. Several studies in SIV-infected macaques suggest that CD8+ T cells may also decrease virus production by suppressing viral transcription. Here, we show that non-HIV-specific, TCR-activated non-cytolytic CD8+ T cells suppress HIV transcription via a virus- and MHC-independent immunoregulatory mechanism that modulates CD4+ T cell proliferation and activation. We also demonstrate that this CD8+ T cell-mediated effect promotes the survival of infected CD4+ T cells harboring integrated, inducible virus. Finally, we used RNA sequencing and secretome analyses to identify candidate cellular pathways that are involved in the virus-silencing mediated by these CD8+ T cells. This study characterizes a previously undescribed mechanism of immune-mediated HIV silencing that may be involved in the establishment and maintenance of the reservoir under antiretroviral therapy and therefore represent a major obstacle to HIV eradication.

SIV and Mycobacterium tuberculosis synergy within the granuloma accelerates the reactivation pattern of latent tuberculosis.

Human immunodeficiency virus infection is the most common risk factor for severe forms of tuberculosis (TB), regardless of CD4 T cell count. Using a well-characterized cynomolgus macaque model of human TB, we compared radiographic, immunologic and microbiologic characteristics of early (subclinical) reactivation of latent M. tuberculosis (Mtb) infection among animals subsequently infected with simian immunodeficiency virus (SIV) or who underwent anti-CD4 depletion by a depletion antibody. CD4 depleted animals had significantly fewer CD4 T cells within granulomas compared to Mtb/SIV co-infected and Mtb-only control animals. After 2 months of treatment, subclinical reactivation occurred at similar rates among CD4 depleted (5 of 7 animals) and SIV infected animals (4 of 8 animals). However, SIV-induced reactivation was associated with more dissemination of lung granulomas that were permissive to Mtb growth resulting in greater bacterial burden within granulomas compared to CD4 depleted reactivators. Granulomas from Mtb/SIV animals displayed a more robust T cell activation profile (IFN-α, IFN-γ, TNF, IL-17, IL-2, IL-10, IL-4 and granzyme B) compared to CD4 depleted animals and controls though these effectors did not protect against reactivation or dissemination, but instead may be related to increased viral and/or Mtb antigens. SIV replication within the granuloma was associated with reactivation, greater overall Mtb growth and reduced Mtb killing resulting in greater overall Mtb burden. These data support that SIV disrupts protective immune responses against latent Mtb infection beyond the loss of CD4 T cells, and that synergy between SIV and Mtb occurs within granulomas.

Antiretroviral and cytotoxic activities of Tityus obscurus synthetic peptide.

New drugs are constantly in demand, and nature's biodiversity is a rich source of new compounds for therapeutic applications. Synthetic peptides based on the transcriptome analysis of scorpion venoms of Tityus obscurus, Opisthacanthus cayaporum, and Hadrurus gertschi were assayed for their cytotoxic and antiretroviral activity. The Tityus obscurus scorpion-derived synthetic peptide (FFGTLFKLGSKLIPGVMKLFSKKKER), in concentrations ranging from 6.24 to 0.39 μM, proved to be the most active one against simian immunodeficiency virus (SIV) replication in the HUT-78 cell line and in primary human leukocytes, with the lowest toxicity for these cells. The immune cellular response evaluated in primary human leukocytes treated with the most promising peptide and challenged with SIV infection exhibited production of cytokines such as interleukin (IL)-4, IL-6, IL-8, IL-10, and interferon-γ, which could be involved in cell defense mechanisms to overcome viral infection through proinflammatory and anti-inflammatory pathways, similar to those evoked for triggering the mechanisms exerted by antiviral restriction factors.

Therapeutic vaccine-mediated Gag-specific CD8+ T-cell induction under anti-retroviral therapy augments anti-virus efficacy of CD8+ cells in simian immunodeficiency virus-infected macaques

Anti-retroviral therapy (ART) can inhibit HIV proliferation but not achieve virus eradication from HIV-infected individuals. Under ART-based HIV control, virus-specific CD8+ T-cell responses are often reduced. Here, we investigated the impact of therapeutic vaccination inducing virus-specific CD8+ T-cell responses under ART on viral control in a macaque AIDS model. Twelve rhesus macaques received ART from week 12 to 32 after simian immunodeficiency virus (SIV) infection. Six of them were vaccinated with Sendai virus vectors expressing SIV Gag and Vif at weeks 26 and 32, and Gag/Vif-specific CD8+ T-cell responses were enhanced and became predominant. All macaques controlled viremia during ART but showed viremia rebound after ART cessation. Analysis of in vitro CD8+ cell ability to suppress replication of autologous lymphocytes-derived SIVs found augmentation of anti-SIV efficacy of CD8+ cells after vaccination. In the vaccinated animals, the anti-SIV efficacy of CD8+ cells at week 34 was correlated positively with Gag-specific CD8+ T-cell frequencies and inversely with rebound viral loads at week 34. These results indicate that Gag-specific CD8+ T-cell induction by therapeutic vaccination can augment anti-virus efficacy of CD8+ cells, which may be insufficient for functional cure but contribute to more stable viral control under ART.

The colonic mucosa-associated microbiome in SIV infection: shift towards Bacteroidetes coincides with mucosal CD4+ T cell depletion and enterocyte damage

The intesinal microbiome is considered important in human immunodeficiency virus (HIV) pathogenesis and therefore represents a potential therapeutic target to improve the patients’ health status. Longitudinal alterations in the colonic mucosa-associated microbiome during simian immunodeficiency virus (SIV) infection were investigated using a 16S rRNA amplicon approach on the Illumina sequencing platform and bioinformatics analyses. Following SIV infection of six animals, no alterations in microbial composition were observed before the viral load peaked in the colon. At the time of acute mucosal SIV replication, the phylum Bacteroidetes including the Bacteroidia class as well as the phylum Firmicutes and its families Ruminococcaceae and Eubacteriaceae became more abundant. Enrichment of Bacteroidetes was maintained until the chronic phase of SIV infection. The shift towards Bacteroidetes in the mucosa-associated microbiome was associated with the extent of SIV infection-induced mucosal CD4+ T cell depletion and correlated with increasing rates of enterocyte damage. These observations suggest that Bacteroidetes strains increase during virus-induced mucosal immune destruction. As Bacteroidetes belong to the lipopolysaccharide- and short chain fatty acids-producing bacteria, their rapid enrichment may contribute to inflammatory tissue damage and metabolic alterations in SIV/HIV infection. These aspects should be considered in future studies on therapeutic interventions.

SIV and CD4 depletion distinctly reactivate latent Mycobacterium. tuberculosis infection

Abstract Human immunodeficiency virus infection is the most common risk factor for severe forms of tuberculosis (TB) regardless of CD4 T cell count. Using a well-characterized cynomolgus macaque model, we compared reactivation of latent M. tuberculosis (Mtb) infection induced by simian immunodeficiency virus (SIV) or anti-CD4 antibody (αCD4). Reactivation, as defined by the appearance of a new lung lesion by PET CT, occurred in 5 of 7 αCD4 treated animals compared to 4 of 8 SIV infected animals within 2 months of treatment. αCD4 animals had significantly fewer CD4 T cells within granulomas compared to SIV/Mtb infected animals. However, compared to αCD4 groups, SIV infection caused more dissemination of lung granulomas and higher granuloma bacterial burdens. Within the granulomas, SIV replication was associated with greater Mtb growth and reduced Mtb killing. Granulomas from SIV/Mtb animals displayed a distinctly different profile of T cell cytokine and granzyme B expression compared to αCD4 animals and controls. PET CT imaging prior to treatment could predict reactivation in αCD4-treated animals, but not in SIV/Mtb animals. These data suggest that SIV infection disrupts protective immune responses against Mtb infection beyond the loss of CD4 T cells, and that synergy between SIV and Mtb occurs within granulomas.

CD8β Depletion Does Not Prevent Control of Viral Replication or Protection from Challenge in Macaques Chronically Infected with a Live Attenuated Simian Immunodeficiency Virus.

We evaluated the contribution of CD8αβ+ T cells to control of live-attenuated simian immunodeficiency virus (LASIV) replication during chronic infection and subsequent protection from pathogenic SIV challenge. Unlike previous reports with a CD8α-specific depleting monoclonal antibody (mAb), the CD8β-specific mAb CD8β255R1 selectively depleted CD8αβ+ T cells without also depleting non-CD8+ T cell populations that express CD8α, such as natural killer (NK) cells and γδ T cells. Following infusion with CD8β255R1, plasma viremia transiently increased coincident with declining peripheral CD8αβ+ T cells. Interestingly, plasma viremia returned to predepletion levels even when peripheral CD8αβ+ T cells did not. Although depletion of CD8αβ+ T cells in the lymph node (LN) was incomplete, frequencies of these cells were 3-fold lower (P = 0.006) in animals that received CD8β255R1 than in those that received control IgG. It is possible that these residual SIV-specific CD8αβ+ T cells may have contributed to suppression of viremia during chronic infection. We also determined whether infusion of CD8β255R1 in the LASIV-vaccinated animals increased their susceptibility to infection following intravenous challenge with pathogenic SIVmac239. We found that 7/8 animals infused with CD8β255R1, and 3/4 animals infused with the control IgG, were resistant to SIVmac239 infection. These results suggest that infusion with CD8β255R1 did not eliminate the protection afforded to LASIV vaccination. This provides a comprehensive description of the impact of CD8β255R1 infusion on the immunological composition in cynomolgus macaques, compared to an isotype-matched control IgG, while showing that the control of LASIV viremia and protection from challenge can occur even after CD8β255R1 administration.IMPORTANCE Studies of SIV-infected macaques that deplete CD8+ T cells in vivo with monoclonal antibodies have provided compelling evidence for their direct antiviral role. These studies utilized CD8α-specific mAbs that target both the major (CD8αβ+) and minor (CD8αα+) populations of CD8+ T cells but additionally deplete non-CD8+ T cell populations that express CD8α, such as NK cells and γδ T cells. In the current study, we administered the CD8β-specific depleting mAb CD8β255R1 to cynomolgus macaques chronically infected with a LASIV to selectively deplete CD8αβ+ T cells without removing CD8αα+ lymphocytes. We evaluated the impact on control of virus replication and protection from pathogenic SIVmac239 challenge. These results underscore the utility of CD8β255R1 for studying the direct contribution of CD8αβ+ T cells in various disease states.

The Tat inhibitor didehydro-cortistatin A suppresses SIV replication and reactivation.

The HIV-1 transactivation protein (Tat) binds the HIV mRNA transactivation responsive element (TAR), regulating transcription and reactivation from latency. Drugs against Tat are unfortunately not clinically available. We reported that didehydro-cortistatin A (dCA) inhibits HIV-1 Tat activity. In human CD4+ T cells isolated from aviremic individuals and in the humanized mouse model of latency, combining dCA with antiretroviral therapy accelerates HIV-1 suppression and delays viral rebound upon treatment interruption. This drug class is amenable to block-and-lock functional cure approaches, aimed at a durable state of latency. Simian immunodeficiency virus (SIV) infection of rhesus macaques (RhMs) is the best-characterized model for AIDS research. Here, we demonstrate, using in vitro and cell-based assays, that dCA directly binds to SIV Tat's basic domain. dCA specifically inhibits SIV Tat binding to TAR, but not a Tat-Rev fusion protein, which activates transcription when Rev binds to its cognate RNA binding site replacing the apical region of TAR. Tat-TAR inhibition results in loss of RNA polymerase II recruitment to the SIV promoter. Importantly, dCA potently inhibits SIV reactivation from latently infected Hut78 cells and from primary CD4+ T cells explanted from SIVmac239-infected RhMs. In sum, dCA's remarkable breadth of activity encourages SIV-infected RhM use for dCA preclinical evaluation.-Mediouni, S., Kessing, C. F., Jablonski, J. A., Thenin-Houssier, S., Clementz, M., Kovach, M. D., Mousseau, G., de Vera, I.M.S., Li, C., Kojetin, D. J., Evans, D. T., Valente, S. T. The Tat inhibitor didehydro-cortistatin A suppresses SIV replication and reactivation.

CD8+ Cytotoxic-T-Lymphocyte Breadth Could Facilitate Early Immune Detection of Immunodeficiency Virus-Derived Epitopes with Limited Expression Levels.

Cytotoxic-T-lymphocyte (CTL) responses are important to control the replication of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV). Accumulating evidence suggests that the ability of a few immunodominant T-cell populations to detect and kill HIV/SIV-infected cells is important in individuals with a protective major histocompatibility complex class I (MHC-I) allele. On the other hand, immunization with live(-attenuated) viruses may be effective against superinfection of virulent viral strains regardless of the host's MHC-I haplotypes, although the underlying mechanisms have not been fully documented. In this article, we propose a hypothesis that the early detection of infected cells in superinfected individuals may be partly facilitated by recognition of diverse CTL epitopes with limited expression levels. We further explain the hypothesis using simple mathematics that was written based on previous in vitro viral suppression assay results and by considering the physical contact of infected cells with CTLs.

Polymorphisms in Rhesus Macaque Tetherin Are Associated with Differences in Acute Viremia in Simian Immunodeficiency Virus Δnef-Infected Animals.

Tetherin (BST-2 or CD317) is an interferon-inducible transmembrane protein that inhibits virus release from infected cells. To determine the extent of sequence variation and the impact of polymorphisms in rhesus macaque tetherin on simian immunodeficiency virus (SIV) infection, tetherin alleles were sequenced from 146 rhesus macaques, including 68 animals infected with wild-type SIVmac239 and 47 animals infected with SIVmac239Δnef Since Nef is the viral gene product of SIV that counteracts restriction by tetherin, these groups afford a comparison of the effects of tetherin polymorphisms on SIV strains that are, and are not, resistant to tetherin. We identified 15 alleles of rhesus macaque tetherin with dimorphic residues at 9 positions. The relationship between these alleles and plasma viral loads was compared during acute infection, prior to the onset of adaptive immunity. Acute viremia did not differ significantly among the wild-type SIV-infected animals; however, differences in acute viral loads were associated with polymorphisms in tetherin among the animals infected with SIVΔnef In particular, polymorphisms at positions 43 and 111 (P43 and H111) were associated with lower acute-phase viral loads for SIVΔnef infection. These observations reveal extensive polymorphism in rhesus macaque tetherin, maintained perhaps as a consequence of variability in the selective pressure of diverse viral pathogens, and identify tetherin alleles that may have an inherently greater capacity to restrict SIV replication in the absence of Nef.IMPORTANCE As a consequence of ongoing evolutionary conflict with viral pathogens, tetherin has accumulated numerous species-specific differences that represent important barriers to the transmission of viruses between species. This study reveals extensive polymorphism in rhesus macaque tetherin and identifies specific alleles that are associated with lower viral loads during the first few weeks after infection with nef-deleted SIV. These observations suggest that the variable selective pressure of viral pathogens, in addition to driving the diversification of tetherin among species, also operates within certain species to maintain sequence variation in tetherin.