scholarly journals A Novel Assay Allows Genotyping of the Latent Reservoir for Human Immunodeficiency Virus Type 1 in the Resting CD4+ T Cells of Viremic Patients

2005 ◽  
Vol 79 (8) ◽  
pp. 5185-5202 ◽  
Author(s):  
Daphne Monie ◽  
Rachel P. Simmons ◽  
Richard E. Nettles ◽  
Tara L. Kieffer ◽  
Yan Zhou ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Latent Reservoir ◽  
Drug Resistant ◽  
Plasma Virus ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT A latent reservoir for human immunodeficiency virus type 1 (HIV-1) consisting of integrated provirus in resting memory CD4+ T cells prevents viral eradication in patients on highly active antiretroviral therapy (HAART). It is difficult to analyze the nature and dynamics of this reservoir in untreated patients and in patients failing therapy, because it is obscured by an excess of unintegrated viral DNA that constitutes the majority of viral species in resting CD4+ T cells from viremic patients. Therefore, we developed a novel culture assay that stimulates virus production from latent, integrated HIV-1 in resting CD4+ T cells in the presence of antiretroviral drugs that prevent the replication of unintegrated virus. Following activation, resting CD4+ T cells with integrated HIV-1 DNA produced virus particles for several days, with peak production at day 5. Using this assay, HIV-1 pol sequences from the resting CD4+ T cells of viremic patients were found to be genetically distinct from contemporaneous plasma virus. Despite the predominance of a relatively homogeneous population of drug-resistant viruses in the plasma of patients failing HAART, resting CD4+ T cells harbored a diverse array of wild-type and archival drug-resistant viruses that were less fit than plasma virus in the context of current therapy. These results provide the first direct evidence that resting CD4+ T cells serve as a stable reservoir for HIV-1 even in the setting of high levels of viremia. The ability to analyze archival species in viremic patients may have clinical utility in detecting drug-resistant variants not present in the plasma.

scholarly journals Characterization of Chemokine Receptor Utilization of Viruses in the Latent Reservoir for Human Immunodeficiency Virus Type 1

2000 ◽  
Vol 74 (17) ◽  
pp. 7824-7833 ◽  
Author(s):  
Theodore Pierson ◽  
Trevor L. Hoffman ◽  
Joel Blankson ◽  
Diana Finzi ◽  
Karen Chadwick ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Chemokine Receptor ◽  
Latent Reservoir ◽  
Immunodeficiency Virus ◽  
Latently Infected ◽  
Hiv 1

ABSTRACT Latently infected resting CD4+ T cells provide a long-term reservoir for human immunodeficiency virus type 1 (HIV-1) and are likely to represent the major barrier to virus eradication in patients on combination antiretroviral therapy. The mechanisms by which viruses enter the latent reservoir and the nature of the chemokine receptors involved have not been determined. To evaluate the phenotype of the virus in this compartment with respect to chemokine receptor utilization, full-length HIV-1 env genes were cloned from latently infected cells and assayed functionally. We demonstrate that the majority of the viruses in the latent reservoir utilize CCR5 during entry, although utilization of several other receptors, including CXCR4, was observed. No alternative coreceptors were shown to be involved in a systematic fashion. Although R5 viruses are present in the latent reservoir, CCR5 was not expressed at high levels on resting CD4+ T cells. To understand the mechanism by which R5 viruses enter latent reservoir, the ability of an R5 virus, HIV-1 Ba-L, to infect highly purified resting CD4+ T lymphocytes from uninfected donors was evaluated. Entry of Ba-L could be observed when virus was applied at a multiplicity approaching 1. However, infection was limited to a subset of cells expressing low levels of CCR5 and markers of immunologic memory. Naive cells could not be infected by an R5 virus even when challenged with a large inoculum. Direct cell fractionation studies showed that latent virus is present predominantly in resting memory cells but also at lower levels in resting naive cells. Taken together, these findings provide support for the hypothesis that the direct infection of naive T cells is not the major mechanism by which the latent infection of resting T cells is established.

scholarly journals Hemofiltrate CC Chemokine 1[9-74] Causes Effective Internalization of CCR5 and Is a Potent Inhibitor of R5-Tropic Human Immunodeficiency Virus Type 1 Strains in Primary T Cells and Macrophages

2002 ◽  
Vol 46 (4) ◽  
pp. 982-990 ◽  
Author(s):  
Jan Münch ◽  
Ludger Ständker ◽  
Stefan Pöhlmann ◽  
Frédéric Baribaud ◽  
Armin Papkalla ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Proteolytic Processing ◽  
Cell Surface Expression ◽  
Cc Chemokine ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Proteolytic processing of the abundant plasmatic human CC chemokine 1 (HCC-1) generates a truncated form, HCC-1[9-74], which is a potent agonist of CCR1, CCR3, and CCR5; promotes calcium influx and chemotaxis of T lymphoblasts, monocytes, and eosinophils; and inhibits infection by CCR5-tropic human immunodeficiency virus type 1 (HIV-1) isolates. In the present study we demonstrate that HCC-1[9-74] interacts with the second external loop of CCR5 and inhibits replication of CCR5-tropic HIV-1 strains in both primary T cells and monocyte-derived macrophages. Low concentrations of the chemokine, however, frequently enhanced the replication of CCR5-tropic HIV-1 isolates but not the replication of X4-tropic HIV-1 isolates. Only HCC-1[9-74] and HCC-1[10-74], but not other HCC-1 length variants, displayed potent anti-HIV-1 activities. Fluorescence-activated cell sorter analysis revealed that HCC-1[9-74] caused up to 75% down-regulation of CCR5 cell surface expression, whereas RANTES (regulated on activation, normal T-cell expressed and secreted) achieved a reduction of only about 40%. Studies performed with green fluorescent protein-tagged CCR5 confirmed that both HCC-1[9-74] and RANTES, but not full-length HCC-1, mediated specific internalization of the CCR5 HIV-1 entry cofactor. Our results demonstrate that the interaction with HCC-1[9-74] causes effective intracellular sequestration of CCR5, but they also indicate that the effect of HCC-1[9-74] on viral replication is subject to marked cell donor- and HIV-1 isolate-dependent variations.

scholarly journals Nef Induces Multiple Genes Involved in Cholesterol Synthesis and Uptake in Human Immunodeficiency Virus Type 1-Infected T Cells

2005 ◽  
Vol 79 (15) ◽  
pp. 10053-10058 ◽  
Author(s):  
Angélique B. van ′t Wout ◽  
J. Victor Swain ◽  
Michael Schindler ◽  
Ushnal Rao ◽  
Melissa S. Pathmajeyan ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Cholesterol Synthesis ◽  
Cholesterol Biosynthesis ◽  
Acetate Incorporation ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Several recent reports indicate that cholesterol might play an important role in human immunodeficiency virus type 1 (HIV-1) replication. We investigated the effects of HIV-1 infection on cholesterol biosynthesis and uptake using microarrays. HIV-1 increased gene expression of cholesterol genes in both transformed T-cell lines and primary CD4+ T cells. Consistent with our microarray data, 14C-labeled mevalonate and acetate incorporation was increased in HIV-1-infected cells. Our data also demonstrate that changes in cholesterol biosynthesis and uptake are only observed in the presence of functional Nef, suggesting that increased cholesterol synthesis may contribute to Nef-mediated enhancement of virion infectivity and viral replication.

scholarly journals Differential Transmission of Human Immunodeficiency Virus Type 1 by Distinct Subsets of Effector Dendritic Cells

2002 ◽  
Vol 76 (15) ◽  
pp. 7812-7821 ◽  
Author(s):  
Rogier W. Sanders ◽  
Esther C. de Jong ◽  
Christopher E. Baldwin ◽  
Joost H. N. Schuitemaker ◽  
Martien L. Kapsenberg ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Dendritic Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Virus Transmission ◽  
Viral Envelope ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Dendritic cells (DC) support human immunodeficiency virus type 1 (HIV-1) transmission by capture of the virus particle in the mucosa and subsequent transport to the draining lymph node, where HIV-1 is presented to CD4+ Th cells. Virus transmission involves a high-affinity interaction between the DC-specific surface molecule DC-SIGN and the viral envelope glycoprotein gp120 and subsequent internalization of the virus, which remains infectious. The mechanism of viral transmission from DC to T cells is currently unknown. Sentinel immature DC (iDC) develop into Th1-promoting effector DC1 or Th2-promoting DC2, depending on the activation signals. We studied the ability of these effector DC subsets to support HIV-1 transmission in vitro. Compared with iDC, virus transmission is greatly upregulated for the DC1 subset, whereas DC2 cells are inactive. Increased transmission by DC1 correlates with increased expression of ICAM-1, and blocking studies confirm that ICAM-1 expression on DC is important for HIV transmission. The ICAM-1-LFA-1 interaction is known to be important for immunological cross talk between DC and T cells, and our results indicate that this cell-cell contact is exploited by HIV-1 for efficient transmission.

scholarly journals Human Immunodeficiency Virus Type 1 Evades T-Helper Responses by Exploiting Antibodies That Suppress Antigen Processing

2004 ◽  
Vol 78 (14) ◽  
pp. 7645-7652 ◽  
Author(s):  
Peter C. Chien ◽  
Sandra Cohen ◽  
Michael Tuen ◽  
James Arthos ◽  
Pei-de Chen ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Antigen Processing ◽  
T Helper ◽  
Peptide Epitopes ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT T-helper responses are important for controlling chronic viral infections, yet T-helper responses specific to human immunodeficiency virus type 1 (HIV-1), particularly to envelope glycoproteins, are lacking in the vast majority of HIV-infected individuals. It was previously shown that the presence of antibodies to the CD4-binding domain (CD4bd) of HIV-1 glycoprotein 120 (gp120) prevents T-helper responses to gp120, but their suppressive mechanisms were undefined (C. E. Hioe et al., J. Virol. 75:10950-10957, 2001). The present study demonstrates that gp120, when complexed to anti-CD4bd antibodies, becomes more resistant to proteolysis by lysosomal enzymes from antigen-presenting cells such that peptide epitopes are not released and presented efficiently by major histocompatibility complex class II molecules to gp120-specific CD4 T cells. Antibodies to other gp120 regions do not confer this effect. Thus, HIV may evade anti-viral T-helper responses by inducing and exploiting antibodies that conceal the virus envelope antigens from T cells.

scholarly journals Role of Cell Surface Glycosaminoglycans of Human T Cells in Human Immunodeficiency Virus Type-1 (HIV-1) Infection

1996 ◽  
Vol 40 (11) ◽  
pp. 827-835 ◽  
Author(s):  
Yukako Ohshiro ◽  
Tsutomu Murakami ◽  
Kazuhiro Matsuda ◽  
Kiyoshi Nishioka ◽  
Keiichi Yoshida ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Cell Surface ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Human T Cells ◽  
Immunodeficiency Virus ◽  
Hiv 1

scholarly journals LFA-1 Is a Key Determinant for Preferential Infection of Memory CD4+ T Cells by Human Immunodeficiency Virus Type 1

2005 ◽  
Vol 79 (21) ◽  
pp. 13714-13724 ◽  
Author(s):  
Mélanie R. Tardif ◽  
Michel J. Tremblay
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Cell Subset ◽  
T Cell Subsets ◽  
Virus Infectivity ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Memory CD4+ T cells are considered a stable latent reservoir for human immunodeficiency virus type 1 (HIV-1) and a barrier to eradication of this retroviral infection in patients under therapy. It has been shown that memory CD4+ T cells are preferentially infected with HIV-1, but the exact mechanism(s) responsible for this higher susceptibility remains obscure. Previous findings indicate that incorporation of host-derived intercellular adhesion molecule 1 (ICAM-1) in HIV-1 increases virus infectivity. To measure the putative involvement of virus-anchored ICAM-1 in the preferential infection of memory cells by HIV-1, quiescent and activated naive and memory T-cell subsets were exposed to isogenic virions either lacking or bearing ICAM-1. Memory CD4+ T cells were found to be more susceptible than naive CD4+ T cells to infection with ICAM-1-bearing virions, as exemplified by a more important virus replication, an increase in integrated viral DNA copies, and a more efficient entry process. Interactions between virus-associated host ICAM-1 and cell surface LFA-1 under a cluster formation seem to be responsible for the preferential HIV-1 infection of the memory cell subset. Altogether, these data shed light on a potential mechanism by which HIV-1 preferentially targets long-lived memory CD4+ T cells.

scholarly journals High Potency of Indolyl Aryl Sulfone Nonnucleoside Inhibitors towards Drug-Resistant Human Immunodeficiency Virus Type 1 Reverse Transcriptase Mutants Is Due to Selective Targeting of Different Mechanistic Forms of the Enzyme

2005 ◽  
Vol 49 (11) ◽  
pp. 4546-4554 ◽  
Author(s):  
Reynel Cancio ◽  
Romano Silvestri ◽  
Rino Ragno ◽  
Marino Artico ◽  
Gabriella De Martino ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Mechanism Of Action ◽  
Drug Resistant ◽  
Wild Type ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Indolyl aryl sulfone (IAS) nonnucleoside inhibitors have been shown to potently inhibit the growth of wild-type and drug-resistant human immunodeficiency virus type 1 (HIV-1), but their exact mechanism of action has not been elucidated yet. Here, we describe the mechanism of inhibition of HIV-1 reverse transcriptase (RT) by selected IAS derivatives. Our results showed that, depending on the substitutions introduced in the IAS common pharmacophore, these compounds can be made selective for different enzyme-substrate complexes. Moreover, we showed that the molecular basis for this selectivity was a different association rate of the drug to a particular enzymatic form along the reaction pathway. By comparing the activities of the different compounds against wild-type RT and the nonnucleoside reverse transcriptase inhibitor-resistant mutant Lys103Asn, it was possible to hypothesize, on the basis of their mechanism of action, a rationale for the design of drugs which could overcome the steric barrier imposed by the Lys103Asn mutation.

scholarly journals Activation of the CD95 (APO-1/Fas) system in T cells from human immunodeficiency virus type-1-infected children

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1741-1746 ◽  
Author(s):  
CB Baumler ◽  
T Bohler ◽  
I Herr ◽  
A Benner ◽  
PH Krammer ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
T Cell ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Cell Depletion ◽  
T Cell Depletion ◽  
Immunodeficiency Virus ◽  
Hiv 1

Abstract Increased apoptosis of CD4+ T cells is considered to be involved in CD4+ T-cell depletion in human immunodeficiency virus type-1 (HIV-1)- infected individuals progressing toward acquired immunodeficiency syndrome (AIDS). We have recently shown that CD95 (APO-1/Fas) expression is strongly increased in T cells of HIV-1-infected children. In this report we provide further evidence for a deregulated CD95 system in AIDS. CD95 expression in HIV-1+ children is not restricted to previously activated CD45RO+ T cells but is also increased on freshly isolated naive CD45RA+ T cells. In addition, specific CD95-mediated apoptosis is enhanced in both CD4+ and CD8+ T cells. Furthermore, levels of CD95 ligand mRNA are profoundly increased. Specific T-cell receptor/CD3-triggered apoptosis in HIV-1+ children is more enhanced in CD8+ than in CD4+ T cells. Accelerated activation induced cell death of T cells could partially be inhibited by blocking anti-CD95 antibody fragments. These data suggest an involvement of the CD95 receptor/ligand system in T-cell depletion and apoptosis in AIDS and may open new avenues of rational intervention strategies.

scholarly journals Intracellular Substrates for the Primer-Unblocking Reaction by Human Immunodeficiency Virus Type 1 Reverse Transcriptase: Detection and Quantitation in Extracts from Quiescent- and Activated-Lymphocyte Subpopulations

2005 ◽  
Vol 49 (5) ◽  
pp. 1761-1769 ◽  
Author(s):  
Anthony J. Smith ◽  
Peter R. Meyer ◽  
Deshratn Asthana ◽  
Margarita R. Ashman ◽  
Walter A. Scott
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
Reverse Transcriptase ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Wild Type ◽  
Cell Extracts ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Treatment of human immunodeficiency virus type 1 (HIV-1)-infected patients with 3′-azido-3′-deoxythymidine (AZT) selects for mutant forms of viral reverse transcriptase (RT) with increased ability to remove chain-terminating nucleotides from blocked DNA chains. We tested various cell extracts for the presence of endogenous acceptor substrates for this reaction. Cell extracts incubated with HIV-1 RT and [32P]ddAMP-terminated DNA primer/template gave rise to 32P-labeled adenosine 2′,3′-dideoxyadenosine 5′,5′′′−P1,P4-tetraphosphate (Ap4ddA), ddATP, Gp4ddA, and Ap3ddA, corresponding to the transfer of [32P]ddAMP to ATP, PPi, GTP, and ADP, respectively. Incubation with [32P]AZT monophosphate (AZTMP)-terminated primer/template gave rise to the analogous 32P-labeled AZT derivatives. Based on the rates of formation of the specific excision products, ATP and PPi levels were determined: ATP was present at 1.3 to 2.2 mM in H9 cells, macrophages, and unstimulated CD4+ or CD8+ T cells, while PPi was present at 7 to 15 μM. Under these conditions, the ATP-dependent reaction predominated, and excision by the AZT-resistant mutant RT was more efficient than wild type RT. Activated CD4+ or CD8+ T cells contained 1.4 to 2.7 mM ATP and 55 to 79 μM PPi. These cellular PPi concentrations are lower than previously reported; nonetheless, the PPi-dependent reaction predominated in extracts from activated T cells, and excision by mutant and wild-type RT occurred with similar efficiency. While PPi-dependent excision may contribute to AZT resistance in vivo, it is likely that selection of AZT-resistant mutants occurs primarily in an environment where the ATP-dependent reaction predominates.

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