HIV-1 Infection Ex Vivo Accelerates Measles Virus Infection by Upregulating Signaling Lymphocytic Activation Molecule (SLAM) in CD4+ T Cells

Abstract BackgroundA reservoir of replication-competent but latent virus is the main obstacle to a cure for HIV-infection. Much of this reservoir resides in memory CD4 T cells. We hypothesized that these cells can be reactivated with antigens from HIV and other common pathogens to reverse latency. ResultsWe obtained mononuclear cells from the peripheral blood of antiretroviral-treated patients with suppressed viremia. We tested pools of peptides and proteins derived from HIV and from other pathogens including CMV for their ability to reverse latency ex vivo by activation of memory responses. We assessed activation of the CD4 T cells by measuring the up-regulation of cell-surface CD69. We assessed HIV-expression using two assays: a real-time PCR assay for virion-associated viral RNA and a droplet digital PCR assay for cell-associated, multiply spliced viral mRNA. Reversal of latency occurred in a minority of cells from some participants, but no single antigen induced HIV-expression ex vivo consistently. When reversal of latency was induced by a specific peptide pool or protein, the extent was proportionally greater than that of T cell activation. ConclusionsIn this group of patients in whom antiretroviral therapy was started during chronic infection, the latent reservoir does not appear to consistently reside in CD4 T cells of a predominant antigen-specificity. Peptide-antigens reversed HIV-latency ex vivo with modest and variable activity. When latency was reversed by specific peptides or proteins, it was proportionally greater than the extent of T cell activation, suggesting partial enrichment of the latent reservoir in cells of specific antigen-reactivity.

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Th1/17 Polarization of CD4 T Cells Supports HIV-1 Persistence during Antiretroviral Therapy

Journal of Virology ◽

10.1128/jvi.01595-15 ◽

2015 ◽

Vol 89 (22) ◽

pp. 11284-11293 ◽

Cited By ~ 51

Author(s):

Hong Sun ◽

Dhohyung Kim ◽

Xiaodong Li ◽

Maja Kiselinova ◽

Zhengyu Ouyang ◽

...

Keyword(s):

T Cells ◽

Antiretroviral Therapy ◽

Cd4 T Cells ◽

Ex Vivo ◽

Memory Cell ◽

Viral Reservoir ◽

Target Cells ◽

Infected Cells ◽

Hiv 1

ABSTRACTThe ability to persist long term in latently infected CD4 T cells represents a characteristic feature of HIV-1 infection and the predominant barrier to efforts aiming at viral eradication and cure. Yet, increasing evidence suggests that only small subsets of CD4 T cells with specific developmental and maturational profiles are able to effectively support HIV-1 long-term persistence. Here, we analyzed how the functional polarization of CD4 T cells shapes and structures the reservoirs of HIV-1-infected cells. We found that CD4 T cells enriched for a Th1/17 polarization had elevated susceptibilities to HIV-1 infection inex vivoassays, harbored high levels of HIV-1 DNA in persons treated with antiretroviral therapy, and made a disproportionately increased contribution to the viral reservoir relative to their contribution to the CD4 T memory cell pool. Moreover, HIV-1 DNA levels in Th1/17 cells remained stable over many years of antiretroviral therapy, resulting in a progressively increasing contribution of these cells to the viral reservoir, and phylogenetic studies suggested preferential long-term persistence of identical viral sequences during prolonged antiretroviral treatment in this cell compartment. Together, these data suggest that Th1/17 CD4 T cells represent a preferred site for HIV-1 DNA long-term persistence in patients receiving antiretroviral therapy.IMPORTANCECurrent antiretroviral therapy is very effective in suppressing active HIV-1 replication but does not fully eliminate virally infected cells. The ability of HIV-1 to persist long term despite suppressive antiretroviral combination therapy represents a perplexing aspect of HIV-1 disease pathogenesis, since most HIV-1 target cells are activated, short-lived CD4 T cells. This study suggests that CD4 T helper cells with Th1/17 polarization have a preferential role as a long-term reservoir for HIV-1 infection during antiretroviral therapy, possibly because these cells may imitate some of the functional properties traditionally attributed to stem cells, such as the ability to persist for extremely long periods of time and to repopulate their own pool size through homeostatic self-renewal. These observations support the hypothesis that HIV-1 persistence is driven by small subsets of long-lasting stem cell-like CD4 T cells that may represent particularly promising targets for clinical strategies aiming at HIV-1 eradication and cure.

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Inhibition of HIV-1 Integration in Ex Vivo-Infected CD4 T Cells from Elite Controllers

Journal of Virology ◽

10.1128/jvi.05327-11 ◽

2011 ◽

Vol 85 (18) ◽

pp. 9646-9650 ◽

Cited By ~ 32

Author(s):

M. J. Buzon ◽

K. Seiss ◽

R. Weiss ◽

A. L. Brass ◽

E. S. Rosenberg ◽

...

Keyword(s):

T Cells ◽

Cd4 T Cells ◽

Ex Vivo ◽

Elite Controllers ◽

Hiv 1

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HIV-1–induced activation of CD4+ T cells creates new targets for HIV-1 infection in human lymphoid tissue ex vivo

Blood ◽

10.1182/blood-2007-05-088435 ◽

2008 ◽

Vol 111 (2) ◽

pp. 699-704 ◽

Cited By ~ 62

Author(s):

Angélique Biancotto ◽

Sarah J. Iglehart ◽

Christophe Vanpouille ◽

Cristian E. Condack ◽

Andrea Lisco ◽

...

Keyword(s):

T Cells ◽

Cd4 T Cells ◽

Lymphoid Tissue ◽

Ex Vivo ◽

Lymphoid Tissues ◽

Strong Positive Correlation ◽

New Targets ◽

Hla Dr ◽

Human Lymphoid Tissue ◽

Hiv 1

We demonstrate mechanisms by which HIV-1 appears to facilitate its own infection in ex vivo–infected human lymphoid tissue. In this system, HIV-1 readily infects various CD4+ T cells, but productive viral infection was supported predominantly by activated T cells expressing either CD25 or HLA-DR or both (CD25/HLA-DR) but not other activation markers: There was a strong positive correlation (r = 0.64, P = .001) between virus production and the number of CD25+/HLA-DR+ T cells. HIV-1 infection of lymphoid tissue was associated with activation of both HIV-1–infected and uninfected (bystanders) T cells. In these tissues, apoptosis was selectively increased in T cells expressing CD25/HLA-DR and p24gag but not in cells expressing either of these markers alone. In the course of HIV-1 infection, there was a significant increase in the number of activated (CD25+/HLA-DR+) T cells both infected and uninfected (bystander). By inducing T cells to express particular markers of activation that create new targets for infection, HIV-1 generates in ex vivo lymphoid tissues a vicious destructive circle of activation and infection. In vivo, such self-perpetuating cycle could contribute to HIV-1 disease.

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Chromatin maturation of the HIV-1 provirus in primary resting CD4+ T cells

10.1101/526053 ◽

2019 ◽

Author(s):

Birgitta Lindqvist ◽

Sara Svensson Akusjarvi ◽

Anders Sonnerborg ◽

Marios Dimitriou ◽

J. Peter Svensson

Keyword(s):

T Cells ◽

T Cell Activation ◽

Cd4 T Cells ◽

Cell Activation ◽

Ex Vivo ◽

Immunodeficiency Virus ◽

Latently Infected ◽

Infected Cells ◽

Active Transcription ◽

Hiv 1

Human immunodeficiency virus type 1 (HIV-1) infection is a chronic condition, where viral DNA integrates into the genome. Latently infected cells form a persistent, heterogeneous reservoir. The reservoir that reinstates an active replication comprises only cells with intact provirus that can be reactivated. We confirmed that latently infected cells from patients exhibited active transcription throughout the provirus. To find transcriptional determinants, we characterized the establishment and maintenance of viral latency during proviral chromatin maturation in cultures of primary CD4+ T-cells for four months after ex vivo HIV-1 infection. As heterochromatin (marked with H3K9me3 or H3K27me3) gradually stabilized, the provirus became less accessible with reduced activation potential. In a subset of infected cells, active marks (i.e., H3K27ac) remained detectable, even after prolonged proviral silencing. After T-cell activation, the proviral activation occurred uniquely in cells with H3K27ac-marked proviruses. Our observations suggested that, after transient proviral activation, cells were actively returned to latency.

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Gliotoxin, identified from a screen of fungal metabolites, disrupts 7SK snRNP, releases P-TEFb and reverses HIV-1 latency

10.1101/848929 ◽

2019 ◽

Cited By ~ 1

Author(s):

Mateusz Stoszko ◽

Abdullah M.S. Al-Hatmi ◽

Anton Skriba ◽

Michael Roling ◽

Enrico Ne ◽

...

Keyword(s):

Gene Expression ◽

Mass Spectrometry ◽

T Cells ◽

Cd4 T Cells ◽

Ex Vivo ◽

Reversal Agents ◽

Latently Infected ◽

Infected Cells ◽

Therapeutic Compounds ◽

Hiv 1

AbstractA leading pharmacological strategy towards HIV cure requires “shock” or activation of HIV gene expression in latently infected cells with Latency Reversal Agents (LRAs) followed by their subsequent clearance. In a screen for novel LRAs we used fungal secondary metabolites (extrolites) as a source of bio-active molecules. Using orthogonal mass spectrometry (MS) coupled to latency reversal bioassays, we identified gliotoxin (GTX) as a novel LRA. GTX significantly induced HIV-1 gene expression in latent ex vivo infected primary cells and in CD4+ T cells from all aviremic HIV-1+ participants. RNA sequencing identified 7SK RNA, the scaffold of the P-TEFb inhibitory 7SK snRNP complex to be significantly reduced upon GTX treatment of independent donor CD4+T cells. GTX disrupted 7SK snRNP, releasing active P-TEFb, which then phosphorylated RNA Pol II CTD, inducing HIV transcription. Our data highlight the power of combining a medium throughput bioassay, mycology and orthogonal mass spectrometry to identify novel potentially therapeutic compounds.

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Relationships Between HIV-Mediated Chemokine Coreceptor Signaling, Cofilin Hyperactivation, Viral Tropism Switch and HIV-Mediated CD4 Depletion

Current HIV Research ◽

10.2174/1570162x17666191106112018 ◽

2020 ◽

Vol 17 (6) ◽

pp. 388-396

Author(s):

Sijia He ◽

Yuntao Wu

Keyword(s):

Immune Deficiency ◽

T Cells ◽

Virus Infection ◽

Hiv Infection ◽

Cd4 T Cells ◽

Cd4 T Cell ◽

Viral Tropism ◽

High Pathogenicity ◽

Hiv 1

: HIV infection causes CD4 depletion and immune deficiency. The virus infects CD4 T cells through binding to CD4 and one of the chemokine coreceptors, CXCR4 (X4) or CCR5 (R5). It has also been known that HIV tropism switch, from R5 to X4, is associated with rapid CD4 depletion, suggesting a key role of viral factors in driving CD4 depletion. However, the virological driver for HIV-mediated CD4 depletion has not been fully elucidated. We hypothesized that HIV-mediated chemokine coreceptor signaling, particularly chronic signaling through CXCR4, plays a major role in CD4 dysfunction and depletion; we also hypothesized that there is an R5X4 signaling (R5X4sig) viral subspecies, evolving from the natural replication course of R5-utilizing viruses, that is responsible for CD4 T cell depletion in R5 virus infection. To gain traction for our hypothesis, in this review, we discuss a recent finding from Cui and co-authors who described the rapid tropism switch and high pathogenicity of an HIV-1 R5 virus, CRF01_AE. We speculate that CRF01_AE may be the hypothetical R5X4sig viral species that is rapidly evolving towards the X4 phenotype. We also attempt to discuss the intricate relationships between HIV-mediated chemokine coreceptor signaling, viral tropism switch and HIV-mediated CD4 depletion, in hopes of providing a deeper understanding of HIV pathogenesis in blood CD4 T cells.

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Ex vivo activation of CD4+ T-cells from donors on suppressive ART can lead to sustained production of infectious HIV-1 from a subset of infected cells

PLoS Pathogens ◽

10.1371/journal.ppat.1006230 ◽

2017 ◽

Vol 13 (2) ◽

pp. e1006230 ◽

Cited By ~ 22

Author(s):

John K. Bui ◽

Elias K. Halvas ◽

Elizabeth Fyne ◽

Michele D. Sobolewski ◽

Dianna Koontz ◽

...

Keyword(s):

T Cells ◽

Cd4 T Cells ◽

Ex Vivo ◽

Infected Cells ◽

Hiv 1

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Optimizing CD4+ T-Cells Transduction Protocol for Gene Therapy of HIV-1 Infected Patients

Blood ◽

10.1182/blood.v126.23.4429.4429 ◽

2015 ◽

Vol 126 (23) ◽

pp. 4429-4429

Author(s):

Amani Ouedrani ◽

Lounes Djerroudi ◽

Isabelle Hmitou ◽

Marina Cavazzana ◽

Fabien Touzot

Keyword(s):

Gene Therapy ◽

T Cells ◽

Cd4 T Cells ◽

Conflicts Of Interest ◽

Lentiviral Vector ◽

Ex Vivo ◽

Effector Memory ◽

Target Cells ◽

Hematopoietic Stem ◽

Hiv 1

Abstract Gene therapy represents an alternative and promising strategy that could provide a path to a curative therapy for HIV-1 infection. One approach involves the introduction of protective gene into a cell, thereby conferring protection against HIV. We plan to conduct an open label phase I/II gene therapy trial for HIV-1 infected patients presenting with lymphoma. The patients will received autologous hematopoietic stem cells transplantation with gene modified CD34+ cells and CD4+ T-cells. CD34+ and CD4+ will be ex vivo transduced by the LVsh5/C46 lentiviral vector (Cal-1, Calimmune, Inc. Tucson, USA). LVsh5/C46 is a SIN lentiviral vector that inhibits two crucial steps of CD4+ T cell infection by the HIV virus: (i) attachment of the virus to its target by downregulation of CCR5 via a short hairpin RNA, (ii) fusion of the virus to the target cell through expression of the C46 inhibitor. We developed a transduction process for CD4+ T-cells using the TransAct™ reagent (Miltenyi Biotec, Bergisch Gladbach , Germany) for CD4+ T-cells activation. Compared to previously published T-cells transduction protocols, the use of Miltenyi TransAct™ permits an equivalent efficacy of transduction - evaluated by measurement of vector copy number through quantitative PCR - without major phenotypic modification. Indeed, CD4+ T-cells ex vivo transduced after activation with the TransAct™ reagent display very few changes in their surface marker with conservation of naive (CCR7+CD62L+CD45RA+), central memory (CCR7+CD62L+CD45RA-) and effector memory (CCR7-CD62L-CD45RA-) subsets in superimposable proportions as initially. Moreover, expression of CD25 remains below 15-25% of cells suggesting a more "gentle " activation of the transduced CD4+ T-cells. Our transduction process had no significant impact in TCRβ repertoire diversity as evaluated by high-throughput sequencing and analyzis of diversity through the Gini-Simpson index or the Shannon index. Finally, transduced CD4 + T-cells retained the ability to to be primed towards the TH1, TH2 and TH17 pathways suggesting that the transduction protocol used did not alter the functional properties of the target cells. Disclosures No relevant conflicts of interest to declare.

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Inconsistent reversal of HIV-1 latency ex vivo by antigens of HIV-1, CMV, and other infectious agents

10.21203/rs.3.rs-35386/v2 ◽

2020 ◽

Author(s):

Thomas Vollbrecht ◽

Aaron O. Angerstein ◽

Bryson Menke ◽

Nikesh M. Kumar ◽

Michelli Faria Oliveira ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Cd4 T Cells ◽

Cell Activation ◽

Ex Vivo ◽

Latent Reservoir ◽

Antigen Specificity ◽

Pcr Assay ◽

Hiv 1

Abstract BackgroundA reservoir of replication-competent but latent virus is the main obstacle to a cure for HIV-1 infection. Much of this reservoir resides in memory CD4 T cells. We hypothesized that these cells can be reactivated with antigens from HIV-1 and other common pathogens to reverse latency. ResultsWe obtained mononuclear cells from the peripheral blood of antiretroviral-treated patients with suppressed viremia. We tested pools of peptides and proteins derived from HIV-1 and from other pathogens including CMV for their ability to reverse latency ex vivo by activation of memory responses. We assessed activation of the CD4 T cells by measuring the up-regulation of cell-surface CD69. We assessed HIV-1 expression using two assays: a real-time PCR assay for virion-associated viral RNA and a droplet digital PCR assay for cell-associated, multiply spliced viral mRNA. Reversal of latency occurred in a minority of cells from some participants, but no single antigen induced HIV-1 expression ex vivo consistently. When reversal of latency was induced by a specific peptide pool or protein, the extent was proportionally greater than that of T cell activation. ConclusionsIn this group of patients in whom antiretroviral therapy was started during chronic infection, the latent reservoir does not appear to consistently reside in CD4 T cells of a predominant antigen-specificity. Peptide-antigens reversed HIV-1 latency ex vivo with modest and variable activity. When latency was reversed by specific peptides or proteins, it was proportionally greater than the extent of T cell activation, suggesting partial enrichment of the latent reservoir in cells of specific antigen-reactivity.

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