Antigen load and viral sequence diversification determine the functional profile of HIV-1-specific CD8+ T cells

Background

Virus-specific CD8⁺ T lymphocytes play a key role in the initial reduction of peak viremia during acute viral infections, but display signs of increasing dysfunction and exhaustion under conditions of chronic antigen persistence. It has been suggested that virus-specific CD8⁺ T cells with a “polyfunctional” profile, defined by the capacity to secrete multiple cytokines or chemokines, are most competent in controlling viral replication in chronic HIV-1 infection. We used HIV-1 infection as a model of chronic persistent viral infection to investigate the process of exhaustion and dysfunction of virus-specific CD8⁺ T cell responses on the single-epitope level over time, starting in primary HIV-1 infection.

Methods and Findings

We longitudinally analyzed the polyfunctional epitope-specific CD8⁺ T cell responses of 18 patients during primary HIV-1 infection before and after therapy initiation or sequence variation in the targeted epitope. Epitope-specific CD8⁺ T cells responded with multiple effector functions to antigenic stimulation during primary HIV-1 infection, but lost their polyfunctional capacity in response to antigen and up-regulated programmed death 1 (PD-1) expression with persistent viremic infection. This exhausted phenotype significantly decreased upon removal of stimulation by antigen, either in response to antiretroviral therapy or by reduction of epitope-specific antigen load in the presence of ongoing viral replication, as a consequence of in vivo selection of cytotoxic T lymphocyte escape mutations in the respective epitopes. Monofunctionality increased in CD8⁺ T cell responses directed against conserved epitopes from 49% (95% confidence interval 27%–72%) to 76% (56%–95%) (standard deviation [SD] of the effect size 0.71), while monofunctionality remained stable or slightly decreased for responses directed against escaped epitopes from 61% (47%–75%) to 56% (42%–70%) (SD of the effect size 0.18) (p < 0.05).

Conclusion

These data suggest that persistence of antigen can be the cause, rather than the consequence, of the functional impairment of virus-specific T cell responses observed during chronic HIV-1 infection, and underscore the importance of evaluating autologous viral sequences in studies aimed at investigating the relationship between virus-specific immunity and associated pathogenesis.     

Phenotypic characterization of HIV-specific CD8+ T cells during early and chronic infant HIV-1 infection

Although CD8⁺ T cells play an important role in the containment of adult HIV-1 replication, their role in infant HIV-1 infection is not as well understood. Impaired HIV-specific CD8⁺ T cell responses may underlie the persistently high viral loads observed in infants. We examined the frequency and phenotype of infant HIV-specific CD8⁺ T cells in 7 HIV-infected antiretroviral therapy-naïve infants during the first 2 years of life, using class I HLA tetramers and IFN-γ-ELISPOT. The frequency (0.088–3.9% of CD3⁺CD8⁺ cells) and phenotype (CD27⁺CD28⁻, CD45RA^+/−, CD57^+/−, HLA-DR⁺, CD95⁺) of infant HIV-specific CD8⁺ T cells were similar to reports in adults undergoing early infection. Unlike adults, at 23–24 months post-infection a high frequency of HIV-specific CD8⁺ T cells expressed HLA-DR (mean 80%, range 68–85%) and CD95 (mean 88%, range 79–96%), suggesting sustained activation and vulnerability to apoptosis. Despite comparable expansion of HIV-specific CD8⁺ T cells of a similar phenotype to adults during early infection, infant T cells failed to contain HIV-1 replication, and remained persistently activated and vulnerable to apoptosis during chronic infection.     

Dendritic cells reveal a broad range of MHC class I epitopes for HIV-1 in persons with suppressed viral load on antiretroviral therapy

Background

HIV-1 remains sequestered during antiretroviral therapy (ART) and can resume high-level replication upon cessation of ART or development of drug resistance. Reactivity of memory CD8⁺ T lymphocytes to HIV-1 could potentially inhibit this residual viral replication, but is largely muted by ART in relation to suppression of viral antigen burden. Dendritic cells (DC) are important for MHC class I processing and presentation of peptide epitopes to memory CD8⁺ T cells, and could potentially be targeted to activate memory CD8⁺ T cells to a broad array of HIV-1 epitopes during ART.

Principal Findings

We show for the first time that HIV-1 peptide-loaded, CD40L-matured DC from HIV-1 infected persons on ART induce IFN gamma production by CD8⁺ T cells specific for a much broader range and magnitude of Gag and Nef epitopes than do peptides without DC. The DC also reveal novel, MHC class I restricted, Gag and Nef epitopes that are able to induce polyfunctional T cells producing various combinations of IFN gamma, interleukin 2, tumor necrosis factor alpha, macrophage inhibitory protein 1 beta and the cytotoxic de-granulation molecule CD107a.

Significance

There is an underlying, broad antigenic spectrum of anti-HIV-1, memory CD8⁺ T cell reactivity in persons on ART that is revealed by DC. This supports the use of DC-based immunotherapy for HIV-1 infection.     

Early and prolonged antiretroviral therapy is associated with an HIV-1-specific T-cell profile comparable to that of long-term non-progressors

Background

Intervention with antiretroviral treatment (ART) and control of viral replication at the time of HIV-1 seroconversion may curtail cumulative immunological damage. We have therefore hypothesized that ART maintenance over a very prolonged period in HIV-1 seroconverters could induce an immuno-virological status similar to that of HIV-1 long-term non-progressors (LTNPs).

Methodology/Principal Findings

We have investigated a cohort of 20 HIV-1 seroconverters on long-term ART (LTTS) and compared it to one of 15 LTNPs. Residual viral replication and reservoirs in peripheral blood, as measured by cell-associated HIV-1 RNA and DNA, respectively, were demonstrated to be similarly low in both cohorts. These two virologically matched cohorts were then comprehensively analysed by polychromatic flow cytometry for HIV-1-specific CD4⁺ and CD8⁺ T-cell functional profile in terms of cytokine production and cytotoxic capacity using IFN-γ, IL-2, TNF-α production and perforin expression, respectively. Comparable levels of highly polyfunctional HIV-1-specific CD4⁺ and CD8⁺ T-cells were found in LTTS and LTNPs, with low perforin expression on HIV-1-specific CD8⁺ T-cells, consistent with a polyfunctional/non-cytotoxic profile in a context of low viral burden.

Conclusions

Our results indicate that prolonged ART initiated at the time of HIV-1 seroconversion is associated with immuno-virological features which resemble those of LTNPs, strengthening the recent emphasis on the positive impact of early treatment initiation and paving the way for further interventions to promote virological control after treatment interruption.     

Ligand-independent exhaustion of killer immunoglobulin-like receptor-positive CD8+ T cells in human immunodeficiency virus type 1 infection

Virus-specific CD8⁺ T cells play a central role in the control of viral infections, including human immunodeficiency virus type 1 (HIV-1) infection. However, despite the presence of strong and broad HIV-specific CD8⁺ T-cell responses in chronic HIV-1 infection, these cells progressively lose critical effector functions and fail to clear the infection. Mounting evidence suggests that the upregulation of several inhibitory regulatory receptors on the surface of CD8⁺ T cells during HIV-1 infection may contribute directly to the impairment of T-cell function. Here, we investigated the role of killer immunoglobulin receptors (KIR), which are expressed on NK cells and on CD8⁺ T cells, in regulating CD8⁺ T-cell function in HIV-1 infection. KIR expression was progressively upregulated on CD8⁺ T cells during HIV-1 infection and correlated with the level of viral replication. Expression of KIR was associated with a profound inhibition of cytokine secretion, degranulation, proliferation, and activation by CD8⁺ T cells following stimulation with T-cell receptor (TCR)-dependent stimuli. In contrast, KIR⁺ CD8⁺ T cells responded potently to TCR-independent stimulation, demonstrating that these cells are functionally competent. KIR-associated suppression of CD8⁺ T-cell function was independent of ligand engagement, suggesting that these regulatory receptors may constitutively repress TCR activation. This ligand-independent repression of TCR activation of KIR⁺ CD8⁺ T cells may represent a significant barrier to therapeutic interventions aimed at improving the quality of the HIV-specific CD8⁺ T-cell response in infected individuals.     

Whole Genome Deep Sequencing of HIV-1 Reveals the Impact of Early Minor Variants Upon Immune Recognition During Acute Infection

Deep sequencing technologies have the potential to transform the study of highly variable viral pathogens by providing a rapid and cost-effective approach to sensitively characterize rapidly evolving viral quasispecies. Here, we report on a high-throughput whole HIV-1 genome deep sequencing platform that combines 454 pyrosequencing with novel assembly and variant detection algorithms. In one subject we combined these genetic data with detailed immunological analyses to comprehensively evaluate viral evolution and immune escape during the acute phase of HIV-1 infection. The majority of early, low frequency mutations represented viral adaptation to host CD8+ T cell responses, evidence of strong immune selection pressure occurring during the early decline from peak viremia. CD8+ T cell responses capable of recognizing these low frequency escape variants coincided with the selection and evolution of more effective secondary HLA-anchor escape mutations. Frequent, and in some cases rapid, reversion of transmitted mutations was also observed across the viral genome. When located within restricted CD8 epitopes these low frequency reverting mutations were sufficient to prime de novo responses to these epitopes, again illustrating the capacity of the immune response to recognize and respond to low frequency variants. More importantly, rapid viral escape from the most immunodominant CD8+ T cell responses coincided with plateauing of the initial viral load decline in this subject, suggestive of a potential link between maintenance of effective, dominant CD8 responses and the degree of early viremia reduction. We conclude that the early control of HIV-1 replication by immunodominant CD8+ T cell responses may be substantially influenced by rapid, low frequency viral adaptations not detected by conventional sequencing approaches, which warrants further investigation. These data support the critical need for vaccine-induced CD8+ T cell responses to target more highly constrained regions of the virus in order to ensure the maintenance of immunodominant CD8 responses and the sustained decline of early viremia.     

Minocycline inhibition of monocyte activation correlates with neuronal protection in SIV neuroAIDS

Background

Minocycline is a tetracycline antibiotic that has been proposed as a potential conjunctive therapy for HIV-1 associated cognitive disorders. Precise mechanism(s) of minocycline''s functions are not well defined.

Methods

Fourteen rhesus macaques were SIV infected and neuronal metabolites measured by proton magnetic resonance spectroscopy (¹H MRS). Seven received minocycline (4 mg/kg) daily starting at day 28 post-infection (pi). Monocyte expansion and activation were assessed by flow cytometry, cell traffic to lymph nodes, CD16 regulation, viral replication, and cytokine production were studied.

Results

Minocycline treatment decreased plasma virus and pro-inflammatory CD14+CD16+ and CD14^loCD16+ monocytes, and reduced their expression of CD11b, CD163, CD64, CCR2 and HLA-DR. There was reduced recruitment of monocyte/macrophages and productively infected cells in axillary lymph nodes. There was an inverse correlation between brain NAA/Cr (neuronal injury) and circulating CD14+CD16+ and CD14^loCD16+ monocytes. Minocycline treatment in vitro reduced SIV replication CD16 expression on activated CD14+CD16+ monocytes, and IL-6 production by monocytes following LPS stimulation.

Conclusion

Neuroprotective effects of minocycline are due in part to reduction of activated monocytes, monocyte traffic. Mechanisms for these effects include CD16 regulation, reduced viral replication, and inhibited immune activation.     

HIV-1 Residual Viremia Correlates with Persistent T-Cell Activation in Poor Immunological Responders to Combination Antiretroviral Therapy

Background

The clinical significance and cellular sources of residual human immunodeficiency virus type 1 (HIV-1) production despite suppressive combination antiretroviral therapy (cART) remain unclear and the effect of low-level viremia on T-cell homeostasis is still debated.

Methodology/Principal Findings

We characterized the recently produced residual viruses in the plasma and short-lived blood monocytes of 23 patients with various immunological responses to sustained suppressive cART. We quantified the residual HIV-1 in the plasma below 50 copies/ml, and in the CD14^high CD16⁻ and CD16⁺ monocyte subsets sorted by flow cytometry, and predicted coreceptor usage by genotyping V3 env sequences.We detected residual viremia in the plasma of 8 of 10 patients with poor CD4⁺ T-cell reconstitution in response to cART and in only 5 of 13 patients with good CD4⁺ T-cell reconstitution. CXCR4-using viruses were frequent among the recently produced viruses in the plasma and in the main CD14^high CD16⁻ monocyte subset. Finally, the residual viremia was correlated with persistent CD4⁺ and CD8⁺ T-cell activation in patients with poor immune reconstitution.

Conclusions

Low-level viremia could result from the release of archived viruses from cellular reservoirs and/or from ongoing virus replication in some patients. The compartmentalization of the viruses between the plasma and the blood monocytes suggests at least two origins of residual virus production during effective cART. CXCR4-using viruses might be produced preferentially in patients on cART. Our results also suggest that low-level HIV-1 production in some patients may contribute to persistent immune dysfunction despite cART.     

The dual impact of HIV-1 infection and aging on naïve CD4 T-cells: additive and distinct patterns of impairment

HIV-1-infected adults over the age of 50 years progress to AIDS more rapidly than adults in their twenties or thirties. In addition, HIV-1-infected individuals receiving antiretroviral therapy (ART) present with clinical diseases, such as various cancers and liver disease, more commonly seen in older uninfected adults. These observations suggest that HIV-1 infection in older persons can have detrimental immunological effects that are not completely reversed by ART. As naïve T-cells are critically important in responses to neoantigens, we first analyzed two subsets (CD45RA⁺CD31⁺ and CD45RA⁺CD31^-) within the naïve CD4⁺ T-cell compartment in young (20–32 years old) and older (39–58 years old), ART-naïve, HIV-1 seropositive individuals within 1–3 years of infection and in age-matched seronegative controls. HIV-1 infection in the young cohort was associated with lower absolute numbers of, and shorter telomere lengths within, both CD45RA⁺CD31⁺CD4⁺ and CD45RA⁺CD31^-CD4⁺ T-cell subsets in comparison to age-matched seronegative controls, changes that resembled seronegative individuals who were decades older. Longitudinal analysis provided evidence of thymic emigration and reconstitution of CD45RA⁺CD31⁺CD4⁺ T-cells two years post-ART, but minimal reconstitution of the CD45RA⁺CD31^-CD4⁺ subset, which could impair de novo immune responses. For both ART-naïve and ART-treated HIV-1-infected adults, a renewable pool of thymic emigrants is necessary to maintain CD4⁺ T-cell homeostasis. Overall, these results offer a partial explanation both for the faster disease progression of older adults and the observation that viral responders to ART present with clinical diseases associated with older adults.     

Sialoadhesin expressed on IFN-induced monocytes binds HIV-1 and enhances infectivity

Background

HIV-1 infection dysregulates the immune system and alters gene expression in circulating monocytes. Differential gene expression analysis of CD14⁺ monocytes from subjects infected with HIV-1 revealed increased expression of sialoadhesin (Sn, CD169, Siglec 1), a cell adhesion molecule first described in a subset of macrophages activated in chronic inflammatory diseases.

Methodology/Principal Findings

We analyzed sialoadhesin expression on CD14⁺ monocytes by flow cytometry and found significantly higher expression in subjects with elevated viral loads compared to subjects with undetectable viral loads. In cultured CD14⁺ monocytes isolated from healthy individuals, sialoadhesin expression was induced by interferon-α and interferon-γ but not tumor necrosis factor-α. Using a stringent binding assay, sialoadhesin-expressing monocytes adsorbed HIV-1 through interaction with the sialic acid residues on the viral envelope glycoprotein gp120. Furthermore, monocytes expressing sialoadhesin facilitated HIV-1 trans infection of permissive cells, which occurred in the absence of monocyte self-infection.

Conclusions/Significance

Increased sialoadhesin expression on CD14⁺ monocytes occurred in response to HIV-1 infection with maximum expression associated with high viral load. We show that interferons induce sialoadhesin in primary CD14⁺ monocytes, which is consistent with an antiviral response during viremia. Our findings suggest that circulating sialoadhesin-expressing monocytes are capable of binding HIV-1 and effectively delivering virus to target cells thereby enhancing the distribution of HIV-1. Sialoadhesin could disseminate HIV-1 to viral reservoirs during monocyte immunosurveillance or migration to sites of inflammation and then facilitate HIV-1 infection of permissive cells.     

Naïve and Memory CD4 T Cells Differ in Their Susceptibilities to Human Immunodeficiency Virus Type 1 Infection following CD28 Costimulation: Implications for Transmission and Pathogenesis

In vitro evidence suggests that memory CD4⁺ cells are preferentially infected by human immunodeficiency virus type 1 (HIV-1), yet studies of HIV-1-infected individuals have failed to detect preferential memory cell depletion. To explore this paradox, we stimulated CD45RA⁺ CD4⁺ (naïve) and CD45RO⁺ CD4⁺ (memory) cells with antibodies to CD3 and CD28 and infected them with either CCR5-dependent (R5) or CXCR4-dependent (X4) HIV-1 isolates. Naïve CD4⁺ cells supported less X4 HIV replication than their memory counterparts. However, naïve cells were susceptible to R5 viral infection, while memory cells remained resistant to infection and viral replication. As with the unseparated cells, mixing the naïve and memory cells prior to infection resulted in cells resistant to R5 infection and highly susceptible to X4 infection. While both naïve and memory CD4⁺ subsets downregulated CCR5 expression in response to CD28 costimulation, only the memory cells produced high levels of the β-chemokines RANTES, MIP-1α, and MIP-1β upon stimulation. Neutralization of these β-chemokines rendered memory CD4⁺ cells highly sensitive to infection with R5 HIV-1 isolates, indicating that downregulation of CCR5 is not sufficient to mediate complete protection from CCR5 strains of HIV-1. These results indicate that susceptibility to R5 HIV-1 isolates is determined not only by the level of CCR5 expression but also by the balance of CCR5 expression and β-chemokine production. Furthermore, our results suggest a model of HIV-1 transmission and pathogenesis in which naïve rather than memory CD4⁺ T cells serve as the targets for early rounds of HIV-1 replication.     

Discovery of a Small Molecule Agonist of Phosphatidylinositol 3-Kinase p110α That Reactivates Latent HIV-1

Combination antiretroviral therapy (cART) can effectively suppress HIV-1 replication, but the latent viral reservoir in resting memory CD4⁺ T cells is impervious to cART and represents a major barrier to curing HIV-1 infection. Reactivation of latent HIV-1 represents a possible strategy for elimination of this reservoir. In this study we describe the discovery of 1,2,9,10-tetramethoxy-7H-dibenzo[de,g]quinolin-7-one (57704) which reactivates latent HIV-1 in several cell-line models of latency (J89GFP, U1 and ACH-2). 57704 also increased HIV-1 expression in 3 of 4 CD8⁺-depleted blood mononuclear cell preparations isolated from HIV-1-infected individuals on suppressive cART. In contrast, vorinostat increased HIV-1 expression in only 1 of the 4 donors tested. Importantly, 57704 does not induce global T cell activation. Mechanistic studies revealed that 57704 reactivates latent HIV-1 via the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway. 57704 was found to be an agonist of PI3K with specificity to the p110α isoform, but not the p110β, δ or γ isoforms. Taken together, our work suggests that 57704 could serve as a scaffold for the development of more potent activators of latent HIV-1. Furthermore, it highlights the involvement of the PI3K/Akt pathway in the maintenance of HIV-1 latency.     

Magnitude and complexity of rectal mucosa HIV-1-specific CD8+ T-cell responses during chronic infection reflect clinical status

Background

The intestinal mucosa displays robust virus replication and pronounced CD4+ T-cell loss during acute human immunodeficiency virus type 1 (HIV-1) infection. The ability of HIV-specific CD8+ T-cells to modulate disease course has prompted intensive study, yet the significance of virus-specific CD8+ T-cells in mucosal sites remains unclear.

Methods and Findings

We evaluated five distinct effector functions of HIVgag-specific CD8+ T-cells in rectal mucosa and blood, individually and in combination, in relationship to clinical status and antiretroviral therapy (ART). In subjects not on ART, the percentage of rectal Gag-specific CD8+ T-cells capable of 3, 4 or 5 simultaneous effector functions was significantly related to blood CD4 count and inversely related to plasma viral load (PVL) (p<0.05). Polyfunctional rectal CD8+ T-cells expressed higher levels of MIP-1β and CD107a on a per cell basis than mono- or bifunctional cells. The production of TNFα, IFN-γ, and CD107a by Gag-specific rectal CD8+ T-cells each correlated inversely (p<0.05) with PVL, and MIP-1β expression revealed a similar trend. CD107a and IFN-γ production were positively related to blood CD4 count (p<0.05), with MIP-1β showing a similar trend. IL-2 production by rectal CD8+ T-cells was highly variable and generally low, and showed no relationship to viral load or blood CD4 count.

Conclusions

The polyfunctionality of rectal Gag-specific CD8+ T-cells appears to be related to blood CD4 count and inversely related to PVL. The extent to which these associations reflect causality remains to be determined; nevertheless, our data suggest a potentially important role for mucosal T-cells in limiting virus replication during chronic infection.     

Differential expression of CD163 on monocyte subsets in healthy and HIV-1 infected individuals

CD163, a haptoglobin-hemoglobin (Hp-Hb) scavenger receptor, expressed by monocytes and macrophages, is important in resolution of inflammation. Age-related non-AIDS co-morbidities in HIV-infected individuals, particularly dementia and cardiovascular disease, result in part from effects of HIV-1 infection on monocyte and macrophage biology. CD163 co-expression on CD14+CD16++ monocytes has been proposed as a useful biomarker for HIV-1 disease progression and the presence of HIV associated dementia. Here we investigated CD163 expression on monocyte subsets ex vivo, on cultured macrophages, and soluble in plasma, in the setting of HIV-1 infection. Whole blood immunophenotyping revealed CD163 expression on CD14++CD16- monocytes but not on CD14+CD16++ monocytes (P = 0.004), supported by CD163 mRNA levels. Incubation with M-CSF induced CD163 protein expression on CD14+CD16++ monocytes to the same extent as CD14++CD16− monocytes. CD163 expression on CD14++CD16+ monocytes from HIV-infected subjects was significantly higher than from uninfected individuals, with a trend towards increased expression on CD14++CD16− monocytes (P = 0.019 and 0.069 respectively), which is accounted for by HIV-1 therapy including protease inhibitors. Shedding of CD163 was shown to predominantly occur from the CD14++CD16− subset after Ficoll isolation and LPS stimulation. Soluble CD163 concentration in plasma from HIV-1 infected donors was similar to HIV-1 uninfected donors. Monocyte CD163 expression in HIV-1 infected patients showed a complicated relationship with classical measures of disease progression. Our findings clarify technical issues regarding CD163 expression on monocyte subsets and further elucidates its role in HIV-associated inflammation by demonstrating that CD163 is readily lost from CD14++CD16− monocytes and induced in pro-inflammatory CD14+CD16++ monocytes by M-CSF. Our data show that all monocyte subsets are potentially capable of differentiating into CD163-expressing anti-inflammatory macrophages given appropriate stimuli. Levels of CD163 expression on monocytes may be a potential biomarker reflecting efforts by the immune system to resolve immune activation and inflammation in HIV-infected individuals.     

Are Long-Term Non-Progressors Very Slow Progressors? Insights from the Chelsea and Westminster HIV Cohort, 1988–2010

Define and identify long-term non-progressors (LTNP) and HIV controllers (HIC), and estimate time until disease progression. LTNP are HIV-1⁺ patients who maintain stable CD4⁺ T-cell counts, with no history of opportunistic infection or antiretroviral therapy (ART). HIC are a subset of LTNP who additionally have undetectable viraemia. These individuals may provide insights for prophylactic and therapeutic development. Records of HIV-1⁺ individuals attending Chelsea and Westminster Hospital (1988–2010), were analysed. LTNP were defined as: HIV-1⁺ for >7 years; ART-naïve; no history of opportunistic infection and normal, stable CD4⁺ T-cell counts. MIXED procedure in SAS using random intercept model identified long-term stable CD4⁺ T-cell counts. Survival analysis estimated time since diagnosis until disease progression. Subjects exhibiting long-term stable CD4⁺ T-cell counts with history below the normal range (<450 cells/µl blood) were compared to LTNP whose CD4⁺ T-cell count always remained normal. Within these two groups subjects with HIV-1 RNA load below limit of detection (BLD) were identified. Of 14,227 patients, 1,204 were diagnosed HIV-1⁺ over 7 years ago and were ART-naïve. Estimated time until disease progression for the 20% (239) whose CD4⁺ T-cell counts remained within the normal range, was 6.2 years (IQR: 2.0 to 9.6); significantly longer than 4.0 years (IQR: 1.0 to 7.3) for patients with historical CD4⁺ T-cell count below normal (Logrank chi-squared = 21.26; p<0.001). Within a subpopulation of 312 asymptomatic patients, 50 exhibited long-term stable CD4⁺ T-cell counts. Of these, 13 were LTNP, one of whom met HIC criteria. Of the remaining 37 patients with long-term stable low CD4⁺ T-cell counts, 3 controlled HIV-1 RNA load BLD. Individuals with stable, normal CD4⁺ T-cell counts progressed less rapidly than those with low CD4⁺ T-cell counts. Few LTNP and HIC identified in this and other studies, endorse the need for universal definitions to facilitate comparison.     

CD8+ T cells from SIV elite controller macaques recognize Mamu-B*08-bound epitopes and select for widespread viral variation

Background

It is generally accepted that CD8⁺ T cell responses play an important role in control of immunodeficiency virus replication. The association of HLA-B27 and -B57 with control of viremia supports this conclusion. However, specific correlates of viral control in individuals expressing these alleles have been difficult to define. We recently reported that transient in vivo CD8⁺ cell depletion in simian immunodeficiency virus (SIV)-infected elite controller (EC) macaques resulted in a brief period of viral recrudescence. SIV replication was rapidly controlled with the reappearance of CD8⁺ cells, implicating that these cells actively suppress viral replication in ECs.

Methods and Findings

Here we show that three ECs in that study made at least seven robust CD8⁺ T cell responses directed against novel epitopes in Vif, Rev, and Nef restricted by the MHC class I molecule Mamu-B*08. Two of these Mamu-B*08-positive animals subsequently lost control of SIV replication. Their breakthrough virus harbored substitutions in multiple Mamu-B*08-restricted epitopes. Indeed, we found evidence for selection pressure mediated by Mamu-B*08-restricted CD8⁺ T cells in all of the newly identified epitopes in a cohort of chronically infected macaques.

Conclusions

Together, our data suggest that Mamu-B*08-restricted CD8⁺ T cell responses effectively control replication of pathogenic SIV_mac239. All seven regions encoding Mamu-B*08-restricted CD8⁺ T cell epitopes also exhibit amino acid replacements typically seen only in the presence of Mamu-B*08, suggesting that the variation we observe is indeed selected by CD8⁺ T cell responses. SIV_mac239 infection of Indian rhesus macaques expressing Mamu-B*08 may therefore provide an animal model for understanding CD8⁺ T cell-mediated control of HIV replication in humans.     

Inefficient Nef-mediated downmodulation of CD3 and MHC-I correlates with loss of CD4+T cells in natural SIV infection

Recent data suggest that Nef-mediated downmodulation of TCR-CD3 may protect SIVsmm-infected sooty mangabeys (SMs) against the loss of CD4⁺ T cells. However, the mechanisms underlying this protective effect remain unclear. To further assess the role of Nef in nonpathogenic SIV infection, we cloned nef alleles from 11 SIVsmm-infected SMs with high (>500) and 15 animals with low (<500) CD4⁺ T-cells/µl in bulk into proviral HIV-1 IRES/eGFP constructs and analyzed their effects on the phenotype, activation, and apoptosis of primary T cells. We found that not only efficient Nef-mediated downmodulation of TCR-CD3 but also of MHC-I correlated with preserved CD4⁺ T cell counts, as well as with high numbers of Ki67⁺CD4⁺ and CD8⁺CD28⁺ T cells and reduced CD95 expression by CD4⁺ T cells. Moreover, effective MHC-I downregulation correlated with low proportions of effector and high percentages of naïve and memory CD8⁺ T cells. We found that T cells infected with viruses expressing Nef alleles from the CD4low SM group expressed significantly higher levels of the CD69, interleukin (IL)-2 and programmed death (PD)-1 receptors than those expressing Nefs from the CD4high group. SIVsmm Nef alleles that were less active in downmodulating TCR-CD3 were also less potent in suppressing the activation of virally infected T cells and subsequent cell death. However, only nef alleles from a single animal with very low CD4⁺ T cell counts rendered T cells hyper-responsive to activation, similar to those of HIV-1. Our data suggest that Nef may protect the natural hosts of SIV against the loss of CD4⁺ T cells by at least two mechanisms: (i) downmodulation of TCR-CD3 to prevent activation-induced cell death and to suppress the induction of PD-1 that may impair T cell function and survival, and (ii) downmodulation of MHC-I to reduce CTL lysis of virally infected CD4⁺ T cells and/or bystander CD8⁺ T cell activation.