Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 8 de 8
Filter
Add more filters










Database
Language
Publication year range
1.
Retrovirology ; 10: 90, 2013 Aug 16.
Article in English | MEDLINE | ID: mdl-23953889

ABSTRACT

BACKGROUND: HIV infection can be treated effectively with antiretroviral agents, but the persistence of a latent reservoir of integrated proviruses prevents eradication of HIV from infected individuals. The chromosomal environment of integrated proviruses has been proposed to influence HIV latency, but the determinants of transcriptional repression have not been fully clarified, and it is unclear whether the same molecular mechanisms drive latency in different cell culture models. RESULTS: Here we compare data from five different in vitro models of latency based on primary human T cells or a T cell line. Cells were infected in vitro and separated into fractions containing proviruses that were either expressed or silent/inducible, and integration site populations sequenced from each. We compared the locations of 6,252 expressed proviruses to those of 6,184 silent/inducible proviruses with respect to 140 forms of genomic annotation, many analyzed over chromosomal intervals of multiple lengths. A regularized logistic regression model linking proviral expression status to genomic features revealed no predictors of latency that performed better than chance, though several genomic features were significantly associated with proviral expression in individual models. Proviruses in the same chromosomal region did tend to share the same expressed or silent/inducible status if they were from the same cell culture model, but not if they were from different models. CONCLUSIONS: The silent/inducible phenotype appears to be associated with chromosomal position, but the molecular basis is not fully clarified and may differ among in vitro models of latency.


Subject(s)
CD4-Positive T-Lymphocytes/virology , HIV/physiology , Virus Integration , Virus Latency , Cells, Cultured , HIV/genetics , Humans , Proviruses/genetics , Proviruses/physiology
2.
PLoS One ; 8(8): e71879, 2013.
Article in English | MEDLINE | ID: mdl-23951263

ABSTRACT

Resting CD4+T cells infected with HIV persist in the presence of suppressive anti-viral therapy (ART) and are barriers to a cure. One potential curative approach, therapeutic vaccination, is fueled by recognition of the ability of a subset of elite controllers (EC) to control virus without therapy due to robust anti-HIV immune responses. Controllers have low levels of integrated HIV DNA and low levels of replication competent virus, suggesting a small reservoir. As our recent data indicates some reservoir cells can produce HIV proteins (termed GPR cells for Gag-positive reservoir cells), we hypothesized that a fraction of HIV-expressing resting CD4+T cells could be efficiently targeted and cleared in individuals who control HIV via anti-HIV cytotoxic T lymphocytes (CTL). To test this we examined if superinfected resting CD4+T cells from EC express HIV Gag without producing infectious virus and the susceptibility of these cells to CTL. We found that resting CD4+T cells expressed HIV Gag and were cleared by autologous CD8+T cells from EC. Importantly, we found the extent of CTL clearance in our in vitro assay correlates with in vivo reservoir size and that a population of Gag expressing resting CD4+T cells exists in vivo in patients well controlled on therapy.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , HIV-1/immunology , T-Lymphocytes, Cytotoxic/immunology , gag Gene Products, Human Immunodeficiency Virus/immunology , CD4-Positive T-Lymphocytes/virology , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/virology , Cells, Cultured , Coculture Techniques , DNA, Viral/genetics , HIV Infections/immunology , HIV Infections/virology , HIV-1/genetics , HIV-1/metabolism , Humans , Real-Time Polymerase Chain Reaction , T-Lymphocytes, Cytotoxic/virology , gag Gene Products, Human Immunodeficiency Virus/genetics , gag Gene Products, Human Immunodeficiency Virus/metabolism
3.
Virology ; 441(1): 18-21, 2013 Jun 20.
Article in English | MEDLINE | ID: mdl-23537959

ABSTRACT

Treatment resistant latent reservoirs remain a barrier to cure HIV, but the maintenance and properties of these reservoirs are not completely understood. 2-LTR circular HIV DNA has been used to assess ongoing viral replication in HAART treated patients. However, the half-life of this DNA form is still debated with conflicting in vivo and in vitro data. Prior in vitro studies have focused on cell lines or short lived activated cells in cultures of brief duration, while in vivo studies have the added complications of cell migration, division, and death. Therefore, we monitored the stability of 2-LTR circles in primary CD4+T cells in a month long culture and compared it to the stability of integrated HIV DNA and T cell receptor excision circles (TRECs), another circular DNA form that is thought to be stable. We found that 2-LTRs, along with TRECs, were stable, suggesting 2-LTRs do not necessarily indicate ongoing replication.


Subject(s)
CD4-Positive T-Lymphocytes/virology , DNA, Circular/genetics , DNA, Viral/genetics , Genomic Instability , HIV Long Terminal Repeat/genetics , HIV-2/genetics , Cells, Cultured , Humans , Time Factors
4.
AIDS ; 26(18): 2295-306, 2012 Nov 28.
Article in English | MEDLINE | ID: mdl-23014521

ABSTRACT

OBJECTIVES: Interest in targeting HIV reservoirs is fueling trials that may decrease reservoir size and/or induce viral replication. Therefore, we aimed to develop strategies to sensitively measure changes in these parameters in patients on and off antiretroviral therapy (ART). Achieving these goals may help evaluate the effects of future clinical trials. DESIGN: To determine the relationship between measurements of total and integrated HIV DNA and their role as markers of reservoir size and ongoing replication, these parameters were measured during the first year of ART, during long-term effective ART, and during a clinical trial aimed at targeting reservoirs. METHODS: Total and integrated HIV DNA were measured in patient samples using quantitative PCR techniques. CD4(+)T cell counts and plasma viremia were also monitored. RESULTS: Unintegrated HIV DNA became undetectable during the first year of ART. Total and integrated HIV DNA levels were generally equal in well controlled patients on ART, and low-level plasma viremia correlated best with integration measures. Finally, patients who controlled plasma viremia (<400 copies/ml) during interferon-α monotherapy exhibited a decrease in the level of integrated but not total HIV DNA and a rise in the ratio of total to integrated HIV DNA over time. CONCLUSION: Our findings suggest that appearance of unintegrated HIV DNA reflects residual HIV expression and de-novo reverse transcription, providing insight into the mechanism by which interferon-α reduces the HIV reservoir. We conclude that concurrent measurements of total and integrated HIV DNA provide information regarding reservoir size and ongoing replication in trials targeting HIV.


Subject(s)
Anti-HIV Agents/pharmacology , DNA, Viral/drug effects , HIV Infections/drug therapy , HIV-1/drug effects , HIV-1/genetics , Interferon-alpha/pharmacology , Adult , Anti-HIV Agents/therapeutic use , Biomarkers/metabolism , CD4 Lymphocyte Count , CD4-Positive T-Lymphocytes/drug effects , Clinical Trials as Topic , Disease Reservoirs/virology , Female , Humans , Interferon-alpha/therapeutic use , Male , Middle Aged , Polymerase Chain Reaction , Viral Load/drug effects , Viremia/virology , Virus Integration/drug effects , Virus Replication/drug effects
5.
PLoS Pathog ; 8(7): e1002818, 2012.
Article in English | MEDLINE | ID: mdl-22911005

ABSTRACT

Despite the effectiveness of highly active antiretroviral therapy (HAART) in treating individuals infected with HIV, HAART is not a cure. A latent reservoir, composed mainly of resting CD4+T cells, drives viral rebound once therapy is stopped. Understanding the formation and maintenance of latently infected cells could provide clues to eradicating this reservoir. However, there have been discrepancies regarding the susceptibility of resting cells to HIV infection in vitro and in vivo. As we have previously shown that resting CD4+T cells are susceptible to HIV integration, we asked whether these cells were capable of producing viral proteins and if so, why resting cells were incapable of supporting productive infection. To answer this question, we spinoculated resting CD4+T cells with or without prior stimulation, and measured integration, transcription, and translation of viral proteins. We found that resting cells were capable of producing HIV Gag without supporting spreading infection. This block corresponded with low HIV envelope levels both at the level of protein and RNA and was not an artifact of spinoculation. The defect was reversed upon stimulation with IL-7 or CD3/28 beads. Thus, a population of latent cells can produce viral proteins without resulting in spreading infection. These results have implications for therapies targeting the latent reservoir and suggest that some latent cells could be cleared by a robust immune response.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/virology , HIV-1/physiology , Virus Latency , env Gene Products, Human Immunodeficiency Virus/biosynthesis , gag Gene Products, Human Immunodeficiency Virus/biosynthesis , CD28 Antigens/metabolism , CD3 Complex/metabolism , Cells, Cultured , Gene Expression Regulation, Viral , HIV Infections/immunology , HIV-1/immunology , HIV-1/metabolism , Humans , Interleukin-17/metabolism , Interleukin-7/immunology , Macrophage Inflammatory Proteins/immunology , Virus Replication
6.
Virology ; 411(2): 344-54, 2011 Mar 15.
Article in English | MEDLINE | ID: mdl-21284992

ABSTRACT

The main impediment to a cure for HIV is the existence of long-lasting treatment resistant viral reservoirs. In this review, we discuss what is currently known about reservoirs, including their formation and maintenance, while focusing on latently infected CD4+ T cells. In addition, we compare several different in vivo and in vitro models of latency. We comment on how each model may reflect the properties of reservoirs in vivo, especially with regard to cell phenotype, since recent studies demonstrate that multiple CD4+ T cell subsets contribute to HIV reservoirs and that with HAART and disease progression the relative contribution of different subsets may change. Finally, we focus on the direct infection of resting CD4+ T cells as a source of reservoir formation and as a model of latency, since recent results help explain the misconception that resting CD4+ T cells appeared to be resistant to HIV in vitro.


Subject(s)
HIV Infections/virology , HIV/growth & development , HIV/pathogenicity , Virus Latency , CD4-Positive T-Lymphocytes/virology , Humans , T-Lymphocyte Subsets/virology
7.
J Virol ; 85(1): 644-8, 2011 Jan.
Article in English | MEDLINE | ID: mdl-20980513

ABSTRACT

Naïve CD4(4) T cells are resistant to both HIV R5 env and vesicular stomatitis virus G protein (VSV-G)-mediated fusion. However, viral particles carrying both HIV R5 env and VSV-G infect naïve cells by an unexplained mechanism. We show that VSV-G-pseudotyped virus cannot fuse to unstimulated cells because the viral particles cannot be endocytosed. However, virions carrying both HIV R5 env and VSV-G can fuse because CD4 binding allows viral uptake. Our findings reveal a unique mechanism by which R5 HIV env and VSV-G cooperate to allow entry to naïve CD4(+) T cells, providing a tool to target naïve CD4(+) T cells with R5 HIV to study HIV coreceptor signaling and latency.


Subject(s)
CD4-Positive T-Lymphocytes/virology , Membrane Fusion , Membrane Glycoproteins/metabolism , Viral Envelope Proteins/metabolism , Virion/metabolism , Virus Internalization , env Gene Products, Human Immunodeficiency Virus/metabolism , HIV-1/genetics , HIV-1/metabolism , Humans , Membrane Glycoproteins/genetics , Receptors, CCR5/metabolism , Vesicular stomatitis Indiana virus/genetics , Vesicular stomatitis Indiana virus/metabolism , Vesicular stomatitis Indiana virus/pathogenicity , Viral Envelope Proteins/genetics , Virion/genetics , env Gene Products, Human Immunodeficiency Virus/genetics
8.
J Virol ; 83(9): 4528-37, 2009 May.
Article in English | MEDLINE | ID: mdl-19211752

ABSTRACT

Resting CD4(+) T cells restrict human immunodeficiency virus (HIV) infection at or before reverse transcription, resulting in slower kinetics of reverse transcription. In a previous study, we showed that, despite this restriction at reverse transcription, HIV integration occurs in resting CD4(+) T cells, albeit with slower kinetics. In that study, the resting T cells were a mixture of memory and naïve cells. Here we asked whether the more quiescent naïve cell subset could be directly infected by HIV and, if so, whether the level of integration in naïve cells was comparable to that in memory cells. We found that HIV integrates in the naïve subset of resting CD4(+) T cells without prior activation of the cells. The level of integration (proviruses/cell) in naïve cells was lower than that in memory cells. This difference between naïve and memory cells was observed whether we inoculated the cells with R5 or X4 HIV and could not be explained solely by differences in coreceptor expression. The presence of endogenous dendritic cells did not change the number of proviruses/cell in memory or naïve cells, and deoxynucleoside pools were equally limiting. Our results instead indicate the existence of a novel restriction point in naïve T cells at viral fusion that results in reduced levels of fusion to naïve CD4(+) T cells. We conclude that HIV can integrate into both naïve and memory cells directly. Our data further support our hypothesis that integrated proviral infection of resting T cells can be established without T-cell activation.


Subject(s)
CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/virology , HIV/immunology , Immunity, Innate/immunology , Immunologic Memory/immunology , Virus Integration/immunology , Cell Line , Cell Separation , Genome/genetics , HIV/genetics , Transcription, Genetic/genetics
SELECTION OF CITATIONS
SEARCH DETAIL
...