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Olivier Schwartz - One of the best experts on this subject based on the ideXlab platform.

  • HIV-1 Nef promotes the localization of Gag to the cell membrane and facilitates Viral cell-to-cell Transfer
    Retrovirology, 2013
    Co-Authors: Marine Malbec, Marion Sourisseau, Francoise Porrot, Fabien Blanchet, Olivier Schwartz, Florence Guivel-benhassine, Nicoletta Casartelli
    Abstract:

    Background Newly synthesized HIV-1 particles assemble at the plasma membrane of infected cells, before being released as free virions or being Transferred through direct cell-to-cell contacts to neighboring cells. Localization of HIV-1 Gag precursor at the cell membrane is necessary and sufficient to trigger Viral assembly, whereas the GagPol precursor is additionally required to generate a fully matured virion. HIV-1 Nef is an accessory protein that optimizes Viral replication through partly defined mechanisms. Whether Nef modulates Gag and/or GagPol localization and assembly at the membrane and facilitates Viral cell-to-cell Transfer has not been extensively characterized so far. Results We report that Nef increases the total amount of Gag proteins present in infected cells, and promotes Gag localization at the cell membrane. Moreover, the processing of p55 into p24 is improved in the presence of Nef. We also examined the effect of Nef during HIV-1 cell-to-cell Transfer. We show that without Nef, Viral Transfer through direct contacts between infected cells and target cells is impaired. With a nef-deleted virus, the number of HIV-1 positive target cells after a short 2h co-culture is reduced, and Viral material Transferred to uninfected cells is less matured. At later time points, this defect is associated with a reduction in the productive infection of new target cells. Conclusions Our results highlight a previously unappreciated role of Nef during the Viral replication cycle. Nef promotes HIV-1 Gag membrane localization and processing, and facilitates Viral cell-to-cell Transfer.

  • simultaneous cell to cell transmission of human immunodeficiency virus to multiple targets through polysynapses
    Journal of Virology, 2009
    Co-Authors: Dominika Rudnicka, Francoise Porrot, Jerome Feldmann, Steve Wietgrefe, Stephanie Guadagnini, Marie Christine Prevost, Jerome Estaquier, Ashley T Haase, Nathalie Solfoulon, Olivier Schwartz
    Abstract:

    Human immunodeficiency virus type 1 (HIV-1) efficiently propagates through cell-to-cell contacts, which include virological synapses (VS), filopodia, and nanotubes. Here, we quantified and characterized further these diverse modes of contact in lymphocytes. We report that Viral transmission mainly occurs across VS and through “polysynapses,” a rosette-like structure formed between one infected cell and multiple adjacent recipients. Polysynapses are characterized by simultaneous HIV clustering and Transfer at multiple membrane regions. HIV Gag proteins often adopt a ring-like supramolecular organization at sites of intercellular contacts and colocalize with CD63 tetraspanin and raft components GM1, Thy-1, and CD59. In donor cells engaged in polysynapses, there is no preferential accumulation of Gag proteins at contact sites facing the microtubule organizing center. The LFA-1 adhesion molecule, known to facilitate Viral replication, enhances formation of polysynapses. Altogether, our results reveal an underestimated mode of Viral Transfer through polysynapses. In HIV-infected individuals, these structures, by promoting concomitant infection of multiple targets in the vicinity of infected cells, may facilitate exponential Viral growth and escape from immune responses.

  • Inefficient human immunodeficiency virus replication in mobile lymphocytes.
    Journal of Virology, 2007
    Co-Authors: Marion Sourisseau, Nathalie Sol-foulon, Francoise Porrot, Fabien Blanchet, Olivier Schwartz
    Abstract:

    Cell-to-cell Viral Transfer facilitates the spread of lymphotropic retroviruses such as human immunodeficiency virus (HIV) and human T-cell leukemia virus (HTLV), likely through the formation of "virological synapses" between donor and target cells. Regarding HIV replication, the importance of cell contacts has been demonstrated, but this phenomenon remains only partly characterized. In order to alter cell-to-cell HIV transmission, we have maintained cultures under continuous gentle shaking and followed Viral replication in this experimental system. In lymphoid cell lines, as well as in primary lymphocytes, Viral replication was dramatically reduced in shaken cultures. To document this phenomenon, we have developed an assay to assess the relative contributions of free and cell-associated virions in HIV propagation. Acutely infected donor cells were mixed with carboxyfluorescein diacetate succinimidyl ester-labeled lymphocytes as targets, and Viral production was followed by measuring HIV Gag expression at different time points by flow cytometry. We report that cellular contacts drastically enhance productive Viral Transfer compared to what is seen with infection with free virus. Productive cell-to-cell Viral transmission required fusogenic Viral envelope glycoproteins on donor cells and adequate receptors on targets. Only a few syncytia were observed in this coculture system. Virus release from donor cells was unaffected when cultures were gently shaken, whereas virus Transfer to recipient cells was severely impaired. Altogether, these results indicate that cell-to-cell Transfer is the predominant mode of HIV spread and help to explain why this virus replicates so efficiently in lymphoid organs.

  • Dendritic cells and HIV-specific CD4+ T cells: HIV antigen presentation, T-cell activation, and Viral Transfer.
    Blood, 2006
    Co-Authors: Arnaud Moris, Fabien Blanchet, Anthony Pajot, Florence Guivel-benhassine, Margarita Salcedo, Olivier Schwartz
    Abstract:

    Human immunodeficiency virus (HIV)-specific CD4+ lymphocytes are preferentially infected in HIV-positive individuals. To study this preferential infection, we have derived several HIV-specific (HS) CD4+ clones. We show that in dendritic cells (DCs), HIV virion capture led to major histocompatibility complex class-II (MHC-II)-restricted Viral antigen presentation and to activation of HS cells. In contrast, neither cell-free virions nor infected lymphocytes activated HS cells. In DCs, the dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN/CD209), which internalizes virions, promoted MHC-II presentation of HIV antigens. Activation of HS cells by HIV-exposed DCs triggered an efficient Viral spread in lymphocytes. CD4+ clones with irrelevant antigenic specificities were not activated by HIV-exposed DCs and poorly supported Viral replication under this setting. Our results unravel the mechanisms of MHC-II-restricted HIV antigen presentation by DCs and describe how HIV gains access to the very cells designed by the immune system to counteract this pathogen.

Benjamin K Chen - One of the best experts on this subject based on the ideXlab platform.

  • quantitative 3d video microscopy of hiv Transfer across t cell virological synapses
    Science, 2009
    Co-Authors: Wolfgang Hubner, Gregory P Mcnerney, Ping Chen, Benjamin M Dale, Ronald E Gordon, Frank Y S Chuang, David M Asmuth, Thomas R Huser, Benjamin K Chen
    Abstract:

    The spread of HIV between immune cells is greatly enhanced by cell-cell adhesions called virological synapses, although the underlying mechanisms have been unclear. With use of an infectious, fluorescent clone of HIV, we tracked the movement of Gag in live CD4 T cells and captured the direct translocation of HIV across the virological synapse. Quantitative, high-speed three-dimensional (3D) video microscopy revealed the rapid formation of micrometer-sized "buttons" containing oligomerized Viral Gag protein. Electron microscopy showed that these buttons were packed with budding Viral crescents. Viral Transfer events were observed to form virus-laden internal compartments within target cells. Continuous time-lapse monitoring showed preferential infection through synapses. Thus, HIV dissemination may be enhanced by virological synapse-mediated cell adhesion coupled to Viral endocytosis.

  • predominant mode of human immunodeficiency virus Transfer between t cells is mediated by sustained env dependent neutralization resistant virological synapses
    Journal of Virology, 2007
    Co-Authors: Ping Chen, Wolfgang Hubner, Matthew A Spinelli, Benjamin K Chen
    Abstract:

    Cell-free human immunodeficiency virus type 1 (HIV-1) can initiate infections, but contact between infected and uninfected T cells can enhance Viral spread through intercellular structures called virological synapses (VS). The relative contribution of VS to cell-free Viral Transfer has not been carefully measured. Using an ultrasensitive, fluorescent virus Transfer assay, we estimate that when VS between HIV-expressing Jurkat T cells and primary CD4 + T cells are formed, cell-associated Transfer of virus is 18,000-fold more efficient than uptake of cell-free virus. Furthermore, in contrast to cell-free virus uptake, the VS deposits virus rapidly into focal, trypsin-resistant compartments in target T cells. This massive virus internalization requires Env-CD4 receptor interactions but is resistant to inhibition by patient-derived neutralizing antisera that inhibit homologous cell-free virus. Deleting the Env cytoplasmic tail does not abrogate VS-mediated Transfer, but it renders the VS sensitive to neutralizing antibodies, suggesting that the tail limits exposure of VS-neutralizing epitopes on the surface of infected cells. Dynamic live imaging of the VS reveals that HIV-expressing cells are polarized and make sustained, Env-dependent contacts with target cells through uropod-like structures. The polarized T-cell morphology, Env-CD4 coordinated adhesion, and Viral Transfer from HIV-infected to uninfected cells suggest that VS allows HIV-1 to evade antibody neutralization and to disseminate efficiently. Future studies will discern to what extent this massive Viral Transfer contributes to productive infection or Viral dissemination through the migration of virus-carrying T cells.

Howard Clark - One of the best experts on this subject based on the ideXlab platform.

  • surfactant protein a binds to hiv and inhibits direct infection of cd4 cells but enhances dendritic cell mediated Viral Transfer
    Journal of Immunology, 2008
    Co-Authors: Gaurav D Gaiha, Tao Dong, Nades Palaniyar, Daniel A Mitchell, Kenneth B M Reid, Howard Clark
    Abstract:

    The identification of surfactant protein A (SP-A) as an important innate immune factor of the lungs, amniotic fluid, and the vaginal tract suggests that it could play an important role during various stages of HIV disease progression and transmission. Therefore, we examined whether SP-A could bind to HIV and also had any effect on Viral infectivity. Our data demonstrate that SP-A binds to HIV in a calcium-dependent manner that is inhibitable by mannose and EDTA. Affinity capture of the HIV Viral lysate reveals that SP-A targets the envelope glycoprotein of HIV (gp120), which was confirmed by ELISA using recombinant gp120. Digestion of gp120 with endoglycosidase H abrogates the binding of SP-A, indicating that the high mannose structures on gp120 are the target of the collectin. Infectivity studies reveal that SP-A inhibits the infection of CD4+ T cells by two strains of HIV (BaL, IIIB) by >80%. Competition assays with CD4 and mAbs F105 and b12 suggest that SP-A inhibits infectivity by occlusion of the CD4-binding site. Studies with dendritic cells (DCs) demonstrate that SP-A enhances the binding of gp120 to DCs, the uptake of Viral particles, and the Transfer of virus from DCs to CD4+ T cells by >5-fold at a pH representative of the vaginal tract. Collectively, these results suggest that SP-A acts as a dual modulator of HIV infection by protecting CD4+ T cells from direct infection but enhancing the Transfer of infection to CD4+ T cells mediated by DCs.

Liyen Huang - One of the best experts on this subject based on the ideXlab platform.

  • adeno associated Viral Transfer of opioid receptor gene to primary sensory neurons a strategy to increase opioid antinociception
    Proceedings of the National Academy of Sciences of the United States of America, 2003
    Co-Authors: Liyen Huang
    Abstract:

    To develop a genetic approach for the treatment of pain, we introduced a recombinant adeno-associated Viral (rAAV) vector containing the cDNA for the μ-opioid receptor (μOR) into primary afferent neurons in dorsal root ganglia (DRGs) of rats, which resulted in a long-lasting (>6 months) increase in μOR expression in DRG neurons. The increase greatly potentiated the antinociceptive effects of morphine in rAAV-μOR-infected rats with and without inflammation. Perforated patch recordings indicated that the efficacy and potency of opioid inhibition of voltage-dependent Ca2+ channels were enhanced in infected neurons, which may underlie the increase in opiate efficacy. These data suggest that Transfer of opioid receptor genes into DRG cells with rAAV vectors may offer a new therapeutic strategy for pain management.

Hrishikesh Pandit - One of the best experts on this subject based on the ideXlab platform.

  • Image_3_Surfactant Protein D Reverses the Gene Signature of Transepithelial HIV-1 Passage and Restricts the Viral Transfer Across the Vaginal Barrier.TIF
    2019
    Co-Authors: Hrishikesh Pandit, Kavita Kale, Hidemi Yamamoto, Gargi Thakur, Sushama Rokade, Payal Chakraborty, Madavan Vasudevan, Uday Kishore, Taruna Madan, Raina Nakova Fichorova
    Abstract:

    Effective prophylactic strategy against the current epidemic of sexually transmitted HIV-1 infection requires understanding of the innate gatekeeping mechanisms at the genital mucosa. Surfactant protein D (SP-D), a member of the collectin family of proteins naturally present in the vaginal tract, is a potential HIV-1 entry inhibitor at the cellular level. Human EpiVaginal tissues compartmentalized in culture inserts were apically exposed to HIV-1 and/or a recombinant fragment of human SP-D (rfhSP-D) and Viral passage was assessed in the basal chamber containing mononuclear leukocytes. To map the gene signature facilitating or resisting the transepithelial Viral Transfer, microarray analysis of the HIV-1 challenged EpiVaginal tissues was performed in the absence or presence of rfhSP-D. Mucosal biocompatibility of rfhSP-D was assessed ex vivo and in the standard rabbit vaginal irritation model. The passage of virus through the EpiVaginal tissues toward the underlying target cells was associated with a global epithelial gene signature including differential regulation of genes primarily involved in inflammation, tight junctions and cytoskeletal framework. RfhSP-D significantly inhibited HIV-1 Transfer across the vaginal tissues and was associated with a significant reversal of virus induced epithelial gene signature. Pro-inflammatory NF-κB and mTOR transcripts were significantly downregulated, while expression of the tight junctions and cytoskeletal genes was upheld. In the absence of virus, rfhSP-D directly interacted with the EpiVaginal tissues and upregulated expression of genes related to structural stability of the cell and epithelial integrity. There was no increment in the Viral acquisition by the PBMCs present in basal chambers wherein, the EpiVaginal tissues in apical chambers were treated with rfhSP-D. The effective concentrations of rfhSP-D had no effect on lactobacilli, epithelial barrier integrity and were safe on repeated applications onto the rabbit vaginal mucosa. This pre-clinical safety data, coupled with its efficacy of restricting Viral passage via reversal of virus-induced gene expression of the vaginal barrier, make a strong argument for clinical trials of rfhSP-D as a topical anti-HIV microbicide.

  • Table_1_Surfactant Protein D Reverses the Gene Signature of Transepithelial HIV-1 Passage and Restricts the Viral Transfer Across the Vaginal Barrier.docx
    2019
    Co-Authors: Hrishikesh Pandit, Kavita Kale, Hidemi Yamamoto, Gargi Thakur, Sushama Rokade, Payal Chakraborty, Madavan Vasudevan, Uday Kishore, Taruna Madan, Raina Nakova Fichorova
    Abstract:

    Effective prophylactic strategy against the current epidemic of sexually transmitted HIV-1 infection requires understanding of the innate gatekeeping mechanisms at the genital mucosa. Surfactant protein D (SP-D), a member of the collectin family of proteins naturally present in the vaginal tract, is a potential HIV-1 entry inhibitor at the cellular level. Human EpiVaginal tissues compartmentalized in culture inserts were apically exposed to HIV-1 and/or a recombinant fragment of human SP-D (rfhSP-D) and Viral passage was assessed in the basal chamber containing mononuclear leukocytes. To map the gene signature facilitating or resisting the transepithelial Viral Transfer, microarray analysis of the HIV-1 challenged EpiVaginal tissues was performed in the absence or presence of rfhSP-D. Mucosal biocompatibility of rfhSP-D was assessed ex vivo and in the standard rabbit vaginal irritation model. The passage of virus through the EpiVaginal tissues toward the underlying target cells was associated with a global epithelial gene signature including differential regulation of genes primarily involved in inflammation, tight junctions and cytoskeletal framework. RfhSP-D significantly inhibited HIV-1 Transfer across the vaginal tissues and was associated with a significant reversal of virus induced epithelial gene signature. Pro-inflammatory NF-κB and mTOR transcripts were significantly downregulated, while expression of the tight junctions and cytoskeletal genes was upheld. In the absence of virus, rfhSP-D directly interacted with the EpiVaginal tissues and upregulated expression of genes related to structural stability of the cell and epithelial integrity. There was no increment in the Viral acquisition by the PBMCs present in basal chambers wherein, the EpiVaginal tissues in apical chambers were treated with rfhSP-D. The effective concentrations of rfhSP-D had no effect on lactobacilli, epithelial barrier integrity and were safe on repeated applications onto the rabbit vaginal mucosa. This pre-clinical safety data, coupled with its efficacy of restricting Viral passage via reversal of virus-induced gene expression of the vaginal barrier, make a strong argument for clinical trials of rfhSP-D as a topical anti-HIV microbicide.

  • Surfactant Protein D Reverses the Gene Signature of Transepithelial HIV-1 Passage and Restricts the Viral Transfer Across the Vaginal Barrier
    Frontiers Media S.A., 2019
    Co-Authors: Hrishikesh Pandit, Kavita Kale, Hidemi Yamamoto, Gargi Thakur, Sushama Rokade, Payal Chakraborty, Madavan Vasudevan, Uday Kishore, Taruna Madan
    Abstract:

    Effective prophylactic strategy against the current epidemic of sexually transmitted HIV-1 infection requires understanding of the innate gatekeeping mechanisms at the genital mucosa. Surfactant protein D (SP-D), a member of the collectin family of proteins naturally present in the vaginal tract, is a potential HIV-1 entry inhibitor at the cellular level. Human EpiVaginal tissues compartmentalized in culture inserts were apically exposed to HIV-1 and/or a recombinant fragment of human SP-D (rfhSP-D) and Viral passage was assessed in the basal chamber containing mononuclear leukocytes. To map the gene signature facilitating or resisting the transepithelial Viral Transfer, microarray analysis of the HIV-1 challenged EpiVaginal tissues was performed in the absence or presence of rfhSP-D. Mucosal biocompatibility of rfhSP-D was assessed ex vivo and in the standard rabbit vaginal irritation model. The passage of virus through the EpiVaginal tissues toward the underlying target cells was associated with a global epithelial gene signature including differential regulation of genes primarily involved in inflammation, tight junctions and cytoskeletal framework. RfhSP-D significantly inhibited HIV-1 Transfer across the vaginal tissues and was associated with a significant reversal of virus induced epithelial gene signature. Pro-inflammatory NF-κB and mTOR transcripts were significantly downregulated, while expression of the tight junctions and cytoskeletal genes was upheld. In the absence of virus, rfhSP-D directly interacted with the EpiVaginal tissues and upregulated expression of genes related to structural stability of the cell and epithelial integrity. There was no increment in the Viral acquisition by the PBMCs present in basal chambers wherein, the EpiVaginal tissues in apical chambers were treated with rfhSP-D. The effective concentrations of rfhSP-D had no effect on lactobacilli, epithelial barrier integrity and were safe on repeated applications onto the rabbit vaginal mucosa. This pre-clinical safety data, coupled with its efficacy of restricting Viral passage via reversal of virus-induced gene expression of the vaginal barrier, make a strong argument for clinical trials of rfhSP-D as a topical anti-HIV microbicide