A low T regulatory cell response may contribute to both viral control and generalized immune activation in HIV controllers

HIV-infected individuals maintaining undetectable viremia in the absence of therapy (HIV controllers) often maintain high HIV-specific T cell responses, which has spurred the development of vaccines eliciting HIV-specific T cell responses. However, controllers also often have abnormally high T cell activation levels, potentially contributing to T cell dysfunction, CD4+ T cell depletion, and non-AIDS morbidity. We hypothesized that a weak T regulatory cell (Treg) response might contribute to the control of viral replication in HIV controllers, but might also contribute to generalized immune activation, contributing to CD4+ T cell loss. To address these hypotheses, we measured frequencies of activated (CD38+ HLA-DR+), regulatory (CD4+CD25+CD127(dim)), HIV-specific, and CMV-specific T cells among HIV controllers and 3 control populations: HIV-infected individuals with treatment-mediated viral suppression (ART-suppressed), untreated HIV-infected "non-controllers" with high levels of viremia, and HIV-uninfected individuals. Despite abnormally high T cell activation levels, controllers had lower Treg frequencies than HIV-uninfected controls (P = 0.014). Supporting the propensity for an unusually low Treg response to viral infection in HIV controllers, we observed unusually high CMV-specific CD4+ T cell frequencies and a strong correlation between HIV-specific CD4+ T cell responses and generalized CD8+ T cell activation levels in HIV controllers (P ≤ 0.001). These data support a model in which low frequencies of Tregs in HIV controllers may contribute to an effective adaptive immune response, but may also contribute to generalized immune activation, potentially contributing to CD4 depletion.     

Treatment-mediated alterations in HIV fitness preserve CD4+ T cell counts but have minimal effects on viral load

For most HIV-infected patients, antiretroviral therapy controls viral replication. However, in some patients drug resistance can cause therapy to fail. Nonetheless, continued therapy with a failing regimen can preserve or even lead to increases in CD4+ T cell counts. To understand the biological basis of these observations, we used mathematical models to explain observations made in patients with drug-resistant HIV treated with enfuvirtide (ENF/T-20), an HIV-1 fusion inhibitor. Due to resistance emergence, ENF was removed from the drug regimen, drug-sensitive virus regrown, and ENF was re-administered. We used our model to study the dynamics of plasma-viral RNA and CD4+ T cell levels, and the competition between drug-sensitive and resistant viruses during therapy interruption and re-administration. Focusing on resistant viruses carrying the V38A mutation in gp41, we found ENF-resistant virus to be 17±3% less fit than ENF-sensitive virus in the absence of the drug, and that the loss of resistant virus during therapy interruption was primarily due to this fitness cost. Using viral dynamic parameters estimated from these patients, we show that although re-administration of ENF cannot suppress viral load, it can, in the presence of resistant virus, increase CD4+ T cell counts, which should yield clinical benefits. This study provides a framework to investigate HIV and T cell dynamics in patients who develop drug resistance to other antiretroviral agents and may help to develop more effective strategies for treatment.     

Type I Interferon Upregulates Bak and Contributes to T Cell Loss during Human Immunodeficiency Virus (HIV) Infection

The role of Type I interferon (IFN) during pathogenic HIV and SIV infections remains unclear, with conflicting observations suggesting protective versus immunopathological effects. We therefore examined the effect of IFNα/β on T cell death and viremia in HIV infection. Ex vivo analysis of eight pro- and anti-apoptotic molecules in chronic HIV-1 infection revealed that pro-apoptotic Bak was increased in CD4+ T cells and correlated directly with sensitivity to CD95/Fas-mediated apoptosis and inversely with CD4+ T cell counts. Apoptosis sensitivity and Bak expression were primarily increased in effector memory T cells. Knockdown of Bak by RNA interference inhibited CD95/Fas-induced death of T cells from HIV-1-infected individuals. In HIV-1-infected patients, IFNα-stimulated gene expression correlated positively with ex vivo T cell Bak levels, CD95/Fas-mediated apoptosis and viremia and negatively with CD4+ T cell counts. In vitro IFNα/β stimulation enhanced Bak expression, CD95/Fas expression and CD95/Fas-mediated apoptosis in healthy donor T cells and induced death of HIV-specific CD8+ T cells from HIV-1-infected patients. HIV-1 in vitro sensitized T cells to CD95/Fas-induced apoptosis and this was Toll-like receptor (TLR)7/9- and Type I IFN-dependent. This sensitization by HIV-1 was due to an indirect effect on T cells, as it occurred in peripheral blood mononuclear cell cultures but not purified CD4+ T cells. Finally, peak IFNα levels and viral loads correlated negatively during acute SIV infection suggesting a potential antiviral effect, but positively during chronic SIV infection indicating that either the virus drives IFNα production or IFNα may facilitate loss of viral control. The above findings indicate stage-specific opposing effects of Type I IFNs during HIV-1 infection and suggest a novel mechanism by which these cytokines contribute to T cell depletion, dysregulation of cellular immunity and disease progression.     

Correlates of preserved CD4(+) T cell homeostasis during natural, nonpathogenic simian immunodeficiency virus infection of sooty mangabeys: implications for AIDS pathogenesis

In contrast to HIV-infected humans, naturally SIV-infected sooty mangabeys (SMs) very rarely progress to AIDS. Although the mechanisms underlying this disease resistance are unknown, a consistent feature of natural SIV infection is the absence of the generalized immune activation associated with HIV infection. To define the correlates of preserved CD4(+) T cell counts in SMs, we conducted a cross-sectional immunological study of 110 naturally SIV-infected SMs. The nonpathogenic nature of the infection was confirmed by an average CD4(+) T cell count of 1,076 +/- 589/mm(3) despite chronic infection with a highly replicating virus. No correlation was found between CD4(+) T cell counts and either age (used as a surrogate marker for length of infection) or viremia. The strongest correlates of preserved CD4(+) T cell counts were a low percentage of circulating effector T cells (CD28(-)CD95(+) and/or IL-7R/CD127(-)) and a high percentage of CD4(+)CD25(+) T cells. These findings support the hypothesis that the level of immune activation is a key determinant of CD4(+) T cell counts in SIV-infected SMs. Interestingly, we identified 14 animals with CD4(+) T cell counts of <500/mm(3), of which two show severe and persistent CD4(+) T cell depletion (<50/mm(3)). Thus, significant CD4(+) T cell depletion does occasionally follow SIV infection of SMs even in the context of generally low levels of immune activation, lending support to the hypothesis of multifactorial control of CD4(+) T cell homeostasis in this model of infection. The absence of AIDS in these "CD4(low)" naturally SIV-infected SMs defines a protective role of the reduced immune activation even in the context of a significant CD4(+) T cell depletion.     

IL-15 treatment during acute simian immunodeficiency virus (SIV) infection increases viral set point and accelerates disease progression despite the induction of stronger SIV-specific CD8+ T cell responses

In this study, we examined the effect of in vivo treatment of acutely SIV-infected Mamu-A*01+ rhesus macaques with IL-15. IL-15 treatment during acute infection increased viral set point by 3 logs and accelerated the development of simian AIDS in two of six animals with one developing early minimal lesion SIV meningoencephalitis. Although IL-15 induced a 2- to 3-fold increase in SIV-specific CD8+ T cell and NK cell numbers at peak viremia and reduced lymph node (LN) SIV-infected cells, this had no impact on peak viremia and did not lower viral set point. At viral set point, however, activated SIV-specific CD8+ T cells and NK cells were reduced in the blood of IL-15-treated animals and LN SIV-infected cells were increased. Week 30 LN from IL-15-treated animals had significantly increased Gag-specific CD8+ T cell numbers, whereas total cell, lymphocyte, and CD4+ T cell numbers were reduced. IL-15 treatment significantly reduced anti-SIV Ab concentrations at week 3 and viral set point. IL-15 increased Ki-67+CD4+ T cells at week 1 of treatment and reduced blood CCR5+ and CD45RA-CD62L- CD4+ T cells. The frequency of day 7 Ki-67+CD4+ T cells strongly correlated with viral set point. These findings suggest that CD4+ T cell activation during acute infection determines subsequent viral set point and IL-15 treatment by increasing such activation elevates viral set point. Finally, IL-15-treated acutely SIV-infected primates may serve as a useful model to investigate the poorly understood mechanisms that control viral set point and disease progression in HIV infection.     

Expansion and contraction of HIV-specific CD4 T cells with short bursts of viremia, but physical loss of the majority of these cells with sustained viral replication

Chronic infection with the HIV results in poor HIV-specific CD4 T cell proliferation, but more recent analyses using intracellular cytokine staining demonstrated that IFN-gamma-producing, HIV-specific CD4 T cells can be detected for years in HIV-infected subjects. Because it is not known whether the majority of HIV-specific T cells are lost or become dysfunctional, we examined the kinetics of the T cell response over an extended period of time using a panel of 10 HLA-DR tetramers loaded with HIV p24 peptides. Tetramer+ CD4 T cells were present at a relatively high frequency during acute infection, but the size of these populations substantially contracted following suppression of viral replication. Short-term cessation of antiretroviral therapy resulted in a burst of viral replication and concomitant expansion of tetramer+ CD4 T cells, and these populations again contracted following reinitiation of therapy. The kinetics with which these cell populations contracted were characteristic of effector T cells, a conclusion that was supported by their phenotypic (CCR7-CD45RA-) and functional properties (IFN-gamma+). Continued high-level viremia resulted in the physical loss of the majority of tetramer+ CD4 T cells, and the decline of HIV p24-specific CD4 T cells occurred more rapidly and was more substantial than the reduction of total CD4 T cell numbers. We conclude that the population of HIV p24-specific CD4 T cells is initially responsive to changes in the levels of viral Ags, but that the majority of these cells are lost in a setting of chronic viremia.     

The Presence of CXCR4-Using HIV-1 Prior to Start of Antiretroviral Therapy Is an Independent Predictor of Delayed Viral Suppression

The emergence of CXCR4-using HIV variants (X4-HIV) is associated with accelerated disease progression in the absence of antiretroviral therapy. However, the effect of X4-HIV variants on the treatment response remains unclear. Here we determined whether the presence of X4-HIV variants influenced the time to undetectable viral load and CD4+ T cell reconstitution after initiation of cART in 732 patients. The presence of X4-HIV variants was determined by MT-2 assay prior to cART initiation and viral load and CD4+ T cell counts were analyzed every 3 to 6 months during a three year follow-up period. Kaplan-Meier and Cox proportional hazard analyses were performed to compare time to viral suppression and the absolute CD4+ T cell counts and increases in CD4+ T cell counts during follow-up were compared for patients with and without X4-HIV at start of cART. Patients harboring X4-HIV variants at baseline showed a delay in time to achieve viral suppression below the viral load detection limit. This delay in viral suppression was independently associated with high viral load and the presence of X4-HIV variants. Furthermore, the absolute CD4+ T cell counts were significantly lower in patients harboring X4-HIV variants at all time points during follow-up. However, no differences were observed in the increase in absolute CD4+ T cell numbers after treatment initiation, indicating that the reconstitution of CD4+ T cells is independent of the presence of X4-HIV variants. The emergence of X4-HIV has been associated with an accelerated CD4+ T cell decline during the natural course of infection and therefore, patients who develop X4-HIV variants may benefit from earlier treatment initiation in order to obtain faster reconstitution of the CD4+ T cell population to normal levels.     

Relationship between pre-existing viral reservoirs and the re-emergence of plasma viremia after discontinuation of highly active anti-retroviral therapy

We examined the pathogenic significance of the latent viral reservoir in the resting CD4+ T cell compartment of HIV-1-infected individuals as well as its involvement in the rebound of plasma viremia after discontinuation of highly active anti-retroviral therapy (HAART). Using heteroduplex mobility and tracking assays, we show that the detectable pool of latently infected, resting CD4+ T cells does not account entirely for the early rebounding plasma HIV in infected individuals in whom HAART has been discontinued. In the majority of patients examined, the rebounding plasma virus was genetically distinct from both the cell-associated HIV RNA and the replication-competent virus within the detectable pool of latently infected, resting CD4 + T cells. These results indicate the existence of other persistent HIV reservoirs that could prompt rapid emergence of plasma viremia after cessation of HAART and underscore the necessity to develop therapies directed toward such populations of infected cells.     

Maintenance of large numbers of virus-specific CD8+ T cells in HIV-infected progressors and long-term nonprogressors

The virus-specific CD8+ T cell responses of 21 HIV-infected patients were studied including a unique cohort of long-term nonprogressors with low levels of plasma viral RNA and strong proliferative responses to HIV Ags. HIV-specific CD8+ T cell responses were studied by a combination of standard cytotoxic T cell (CTL) assays, MHC tetramers, and TCR repertoire analysis. The frequencies of CD8+ T cells specific to the majority of HIV gene products were measured by flow cytometric detection of intracellular IFN-gamma in response to HIV-vaccinia recombinant-infected autologous B cells. Very high frequencies (0.8-18.0%) of circulating CD8+ T cells were found to be HIV specific. High frequencies of HIV-specific CD8+ T cells were not limited to long-term nonprogressors with restriction of plasma virus. No correlation was found between the frequency of HIV-specific CD8+ T cells and levels of plasma viremia. In each case, the vast majority of cells (up to 17.2%) responded to gag-pol. Repertoire analysis showed these large numbers of Ag-specific cells were scattered throughout the repertoire and in the majority of cases not contained within large monoclonal expansions. These data demonstrate that high numbers of HIV-specific CD8+ T cells exist even in patients with high-level viremia and progressive disease. Further, they suggest that other qualitative parameters of the CD8+ T cell response may differentiate some patients with very low levels of plasma virus and nonprogressive disease.     

Acute Plasma Biomarkers of T Cell Activation Set-Point Levels and of Disease Progression in HIV-1 Infection

T cell activation levels, viral load and CD4⁺ T cell counts at early stages of HIV-1 infection are predictive of the rate of progression towards AIDS. We evaluated whether the inflammatory profile during primary HIV-1 infection is predictive of the virological and immunological set-points and of disease progression. We quantified 28 plasma proteins during acute and post-acute HIV-1 infection in individuals with known disease progression profiles. Forty-six untreated patients, enrolled during primary HIV-1 infection, were categorized into rapid progressors, progressors and slow progressors according to their spontaneous progression profile over 42 months of follow-up. Already during primary infection, rapid progressors showed a higher number of increased plasma proteins than progressors or slow progressors. The plasma levels of TGF-β1 and IL-18 in primary HIV-1 infection were both positively associated with T cell activation level at set-point (6 months after acute infection) and together able to predict 74% of the T cell activation variation at set-point. Plasma IP-10 was positively and negatively associated with, respectively, T cell activation and CD4⁺ T cell counts at set-point and capable to predict 30% of the CD4⁺ T cell count variation at set-point. Moreover, plasma IP-10 levels during primary infection were predictive of rapid progression. In primary infection, IP-10 was an even better predictor of rapid disease progression than viremia or CD4⁺ T cell levels at this time point. The superior predictive capacity of IP-10 was confirmed in an independent group of 88 HIV-1 infected individuals. Altogether, this study shows that the inflammatory profile in primary HIV-1 infection is associated with T cell activation levels and CD4⁺ T cell counts at set-point. Plasma IP-10 levels were of strong predictive value for rapid disease progression. The data suggest IP-10 being an earlier marker of disease progression than CD4⁺ T cell counts or viremia levels.     

Interleukin-7 (IL-7): immune function, involvement in the pathogenesis of HIV infection and therapeutic potential

Interleukin 7 (IL-7), which is constitutively produced particularly by stromal cells from the bone marrow and thymus, plays a crucial role in T cell homeostasis. This cytokine is implicated in thymopoiesis since it sustains thymocyte proliferation and survival. It regulates peripheral naive T cell survival by modulating the expression of the anti-apoptotic molecule Bcl-2, and sustains peripheral T cell expansion in response to antigenic stimulation. Infection by the human immunodeficiency virus (HIV) leads to severe T lymphopenia and general immune dysfunction. Increased IL-7 plasma levels are generally observed in HIV-infected patients. The existence of an inverse correlation between IL-7 plasma levels and the CD4+ T cell count suggests that a direct feedback mechanism is working to restore peripheral T cell counts in lymphopenic patients. Here, IL-7 plasma levels are a good predictive marker of CD4+ T cell restoration during therapy. Combinations of antiretroviral treatments considerably slow disease progression. They drastically decrease the viral load and, in most patients, significantly increase peripheral CD4+ T cell counts. However, despite their usual ability to reduce viral replication, such treatments often fail to reverse lymphopenia and do not restore specific antiviral immune responses. IL-7, based therapy, combined with efficient antiretroviral treatment, may be beneficial to HIV-infected patients by promoting thymic output, sustaining naive T cell counts and increasing memory T cell activation.     

GB Virus C Infection Is Associated with Altered Lymphocyte Subset Distribution and Reduced T Cell Activation and Proliferation in HIV-Infected Individuals

GBV-C infection is associated with prolonged survival and with reduced T cell activation in HIV-infected subjects not receiving combination antiretroviral therapy (cART). The relationship between GBV-C and T cell activation in HIV-infected subjects was examined. HIV-infected subjects on cART with non-detectable HIV viral load (VL) or cART naïve subjects were studied. GBV-C VL and HIV VL were determined. Cell surface markers of activation (CD38⁺/HLA-DR⁺), proliferation (Ki-67+), and HIV entry co-receptor expression (CCR5+ and CXCR4+) on total CD4+ and CD8+ T cells, and on naïve, central memory (CM), effector memory (EM), and effector CD4+ and CD8+ subpopulations were measured by flow cytometry. In subjects with suppressed HIV VL, GBV-C was consistently associated with reduced activation in naïve, CM, EM, and effector CD4+ cells. GBV-C was associated with reduced CD4+ and CD8+ T cell surface expression of activation and proliferation markers, independent of HIV VL classification. GBV-C was also associated with higher proportions of naïve CD4+ and CD8+ T cells, and with lower proportions of EM CD4+ and CD8+ T cells. In conclusion, GBV-C infection was associated with reduced activation of CD4+ and CD8+ T cells in both HIV viremic and HIV RNA suppressed patients. Those with GBV-C infection demonstrated an increased proportion of naive T cells and a reduction in T cell activation and proliferation independent of HIV VL classification, including those with suppressed HIV VL on cART. Since HIV pathogenesis is thought to be accelerated by T cell activation, these results may contribute to prolonged survival among HIV infected individuals co-infected with GBV-C. Furthermore, since cART therapy does not reduce T cell activation to levels seen in HIV-uninfected people, GBV-C infection may be beneficial for HIV-related diseases in those effectively treated with anti-HIV therapy.     

T cell-specific siRNA delivery suppresses HIV-1 infection in humanized mice

Evaluation of the therapeutic potential of RNAi for HIV infection has been hampered by the challenges of siRNA delivery and lack of suitable animal models. Using a delivery method for T cells, we show that siRNA treatment can dramatically suppress HIV infection. A CD7-specific single-chain antibody was conjugated to oligo-9-arginine peptide (scFvCD7-9R) for T cell-specific siRNA delivery in NOD/SCIDIL2rgamma-/- mice reconstituted with human lymphocytes (Hu-PBL) or CD34+ hematopoietic stem cells (Hu-HSC). In HIV-infected Hu-PBL mice, treatment with anti-CCR5 (viral coreceptor) and antiviral siRNAs complexed to scFvCD7-9R controlled viral replication and prevented the disease-associated CD4 T cell loss. This treatment also suppressed endogenous virus and restored CD4 T cell counts in mice reconstituted with HIV+ peripheral blood mononuclear cells. Moreover, scFvCD7-9R could deliver antiviral siRNAs to naive T cells in Hu-HSC mice and effectively suppress viremia in infected mice. Thus, siRNA therapy for HIV infection appears to be feasible in a preclinical animal model.     

After 18 months of antiretroviral therapy,total HIV DNA decreases more pronouncedly in patients infected by CRF01_AE than in those infected by subtype B and CRF07_BC

Whether the amount of HIV DNA is associated with the subtype of HIV‐1 after antiretroviral therapy (ART) has not been reported. In the present study, the amount of HIV DNA and RNA and CD4+T counts in blood and semen prior to and after 18 months of ART were compared in 48 patients infected by CRF01_AE, subtype B or CRF07_BC of HIV‐1. Viral RNA was suppressed and CD4 cell count recovery achieved in all patients. The level of HIV DNA were similar before ART; however, patients with CRF01_AE had less HIV DNA after ART than those with subtype B and CRF07_BC infection. According to prediction of co‐receptor usage by Geno2Pheno and PSSM in combination, more than 35.6% of clones for CRF01_AE were predicted as CXCR4‐using before ART, whereas less than 6% of those for subtype B and CRF07_BC were predicted as CXCR4‐using. After 18 months of ART, no CXCR4‐using clones were predicted in any of the subtypes. Despite more HIV RNA and fewer CD4 + T cells in patients with CRF01_AE before therapy, no significant differences (P > 0.05) in viral RNA or CD4 cell counts were observed between the subtypes after 18 months of ART. Thus, 18 months of antiretroviral therapy was more efficient in patients with CRF01_AE. Considering that successful ART dramatically reduces the viral load in both blood and semen, risks of sexual transmission of HIV were reduced, contributing to prevention of rapid spread of HIV among men who have sex with men in the region.

     

Limited HIV Infection of Central Memory and Stem Cell Memory CD4+ T Cells Is Associated with Lack of Progression in Viremic Individuals

A rare subset of HIV-infected individuals, designated viremic non-progressors (VNP), remain asymptomatic and maintain normal levels of CD4+ T-cells despite persistently high viremia. To identify mechanisms potentially responsible for the VNP phenotype, we compared VNPs (average >9 years of HIV infection) to HIV-infected individuals who have similar CD4+ T-cell counts and viral load, but who are likely to progress if left untreated (“putative progressors”, PP), thus avoiding the confounding effect of differences related to substantial CD4+ T cell depletion. We found that VNPs, compared to PPs, had preserved levels of CD4+ stem cell memory cells (T_SCM (p<0.0001), which was associated with decreased HIV infection of these cells in VNPs (r = −0.649, p = 0.019). In addition, VNPs had decreased HIV infection in CD4+ central memory (T_CM) cells (p = 0.035), and the total number of T_CM cells was associated with increased proliferation of memory CD4+ T cells (r = 0.733, p = 0.01). Our results suggest that, in HIV-infected VNPs, decreased infection of CD4+ T_CM and T_SCM, cells are involved in preservation of CD4+ T cell homeostasis and lack of disease progression despite high viremia.     

HIV-specific CD8+ T cell responses to HXB2 Gag and Nef peptide pools in Chinese HIV/AIDS patients

HXB2 is primarily used as a template strain in developing HIV vaccines in Europe and the US. However,it is not yet known whether the strain can induce strong HIV-specific CD8+ T cell responses in Chinese HIV/AIDS patients. In the present study,two groups of subjects were investigated:9 AIDS patients and 7 long-term nonprogressors (LTNPs). HIV-specific CD8+ T cell responses were examined in all patients through the ELISPOT assay. CD4+ T cell counts,CD8+ T cell counts,viral load and HIV subtype of each patien...     

Impaired monocyte maturation in response to CpG oligodeoxynucleotide is related to viral RNA levels in human immunodeficiency virus disease and is at least partially mediated by deficiencies in alpha/beta interferon responsiveness and production

The biological activity of CpG oligodeoxynucleotide 2216 (ODN2216), a Toll-like receptor 9 agonist, was investigated with monocytes from human immunodeficiency virus (HIV)-negative and HIV-positive (HIV+) donors. Exposure of peripheral blood mononuclear cells to CpG ODN2216 led to decreased expression of the monocyte marker CD14 and increased expression of the dendritic cell marker CD83, as well as increased expression of HLA-DR, CD40, CD80, and CD86 among the monocytes. Several features of the CpG ODN-induced maturation were diminished in monocytes from HIV+ donors, and these deficiencies were related to increased viremia but not to CD4 cell counts. Alpha interferon (IFN-alpha) was implicated as at least a partial mediator of the CpG ODN-induced monocyte maturation. Reduced production of IFN-alpha in response to CpG ODN and reduced frequencies of plasmacytoid dendritic cells, the principal IFN-alpha-producing cell type in peripheral blood, were observed in peripheral blood mononuclear cells from HIV+ donors. These deficiencies also were related to levels of plasma HIV RNA. Responses of monocytes from HIV+ donors to direct stimulation with IFN-alpha also were partially impaired. Thus, reduced production of IFN-alpha and reduced IFN-alpha responsiveness may contribute to diminished functional responses to CpG ODN in HIV disease. Application of CpG ODNs in HIV disease for adjuvant or immunoregulatory purposes may be particularly useful for HIV+ donors without high-level viremia.