Human T-lymphoblastoid cells selected for growth in serum-free medium provide new tools for study of HIV replication and cytopathogenicity

Yahi, Nouara; Fantini, Jacques; Baghdiguian, Stephen; Chermann, Jean‐Claude

doi:10.1016/0166-0934(91)90099-l

Cited by 10 publications

(4 citation statements)

References 18 publications

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“…In agreement with previous observations [31], the lack of serum proteins in the assay significantly decreased the infectivity of HIV-1 resulting in a reduction in virus entry from 30 to 70% depending on the HIV-1 strain used (data not shown). Therefore, to increase virus uptake by target cells we used the spinoculation method, which was reported to significantly improve the efficiency of infection [32].…”

Section: Resultssupporting

confidence: 92%

Inhibition of HIV-1 Infection by Human α-Defensin-5, a Natural Antimicrobial Peptide Expressed in the Genital and Intestinal Mucosae

Furci¹,

Tolazzi²,

Sironi³

et al. 2012

PLoS ONE

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Backgroundα-defensin-5 (HD5) is a key effector of the innate immune system with broad anti-bacterial and anti-viral activities. Specialized epithelial cells secrete HD5 in the genital and gastrointestinal mucosae, two anatomical sites that are critically involved in HIV-1 transmission and pathogenesis. We previously found that human neutrophil defensins (HNP)-1 and -2 inhibit HIV-1 entry by specific bilateral interaction both with the viral envelope and with its primary cellular receptor, CD4. Despite low amino acid identity, human defensin-5 (HD5) shares with HNPs a high degree of structural homology.Methodology/Principal FindingsHere, we demonstrate that HD5 inhibits HIV-1 infection of primary CD4+ T lymphocytes at low micromolar concentration under serum-free and low-ionic-strength conditions similar to those occurring in mucosal fluids. Blockade of HIV-1 infection was observed with both primary and laboratory-adapted strains and was independent of the viral coreceptor-usage phenotype. Similar to HNPs, HD5 inhibits HIV-1 entry into the target cell by interfering with the reciprocal interaction between the external envelope glycoprotein, gp120, and CD4. At high concentrations, HD5 was also found to downmodulate expression of the CXCR4 coreceptor, but not of CCR5. Consistent with its broad spectrum of activity, antibody competition studies showed that HD5 binds to a region overlapping with the CD4- and coreceptor-binding sites of gp120, but not to the V3 loop region, which contains the major determinants of coreceptor-usage specificity.Conclusion/SignificanceThese findings provide new insights into the first line of immune defense against HIV-1 at the mucosal level and open new perspectives for the development of preventive and therapeutic strategies.

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Section: Resultssupporting

confidence: 92%

Inhibition of HIV-1 Infection by Human α-Defensin-5, a Natural Antimicrobial Peptide Expressed in the Genital and Intestinal Mucosae

Furci¹,

Tolazzi²,

Sironi³

et al. 2012

PLoS ONE

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“…Human T lymphoblastoid CEM-CL5 cells were cloned in our laboratory [17].These cells were used for co-cultivation experiments because of their high sensitivity to HIV infection, due to a constitutive overexpression of CD4. Recombinant IFN-y was purchased from Sigma (St. Louis, MO) .…”

Section: Cell Culturementioning

confidence: 99%

Inhibition of human immunodeficiency virus infection in human colon epithelial cells by recombinant interferon‐γ

et al. 1992

Self Cite

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Pretreatment of human colon epithelial cells HT29 by recombinant gamma interferon (IFN)-gamma was found to protect the cells from infection with various isolates of human immunodeficiency virus (HIV)-1 and HIV-2, as assessed by co-cultivation with human T lymphoblastoid cells and gene amplification by polymerase chain reaction technique. Additionally, IFN-gamma induced a dose-dependent inhibition of HIV-1 and HIV-2 production in chronically infected HT29 cells. In situ hybridization studies demonstrated that IFN-treated cells were still able to synthesize viral messenger ribonucleic acid. However, the expression of the p24 product of the gag gene was markedly decreased after IFN treatment as demonstrated by radio-immunoprecipitation assay. Taken together, these data suggested that the cytokine acted at the post-translational level by inhibiting the processing of structural viral proteins. It is concluded from this study that IFN-gamma has a potent anti-HIV effect on epithelial gastrointestinal cells.

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“…When indicated, HT29-D4 cells were grown on Nuclepore filters (pore size, 3 ,um) in two-compartment Transwell cell culture chambers (Costar) as previously described (8). Human T-lymphoblastoid CEM-CL5 cells (27) were grown in RPMI 1640 supplemented with 10% FCS. Human peripheral blood lymphocytes (PBL) obtained from normal seronegative donors were separated by Ficoll-Hypaque gradient centrifugation, cultured in RPMI 1640 supplemented with 10% FCS, and stimulated with phytohemagglutinin 3 days before infection.…”

Section: Materuils and Methodsmentioning

confidence: 99%

Galactosyl ceramide (or a closely related molecule) is the receptor for human immunodeficiency virus type 1 on human colon epithelial HT29 cells

Yahi

Baghdiguian

Moreau

et al. 1992

J Virol

229

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The gastrointestinal tract is considered to be a major route of infection for human immunodeficiency virus (HIV). Infection of human colon epithelial cells by HIV is not blocked by anti-CD4 antibodies known to block infection of lymphoid cells (J. Fantini, N. Yahi, and J. C. Chermann, Proc. Natl. Acad. Sci. USA 88:9297-9301, 1991), suggesting the presence of an alternate receptor for HIV on these cells. In this report, we show that (i) a monoclonal antibody specifically directed against galactosyl ceramide inhibited the infection of HT29 cells by two markedly different strains of HIV-1, as assessed by polymerase chain reaction amplification and reverse transcriptase assay; (ii) this antibody strongly labeled the surface of HT29 cells by immunofluorescence and electron microscopic immunolocalization; (iii) the labeling was preferentially but not totally restricted to the basolateral membrane domain of differentiated colonic cells, in agreement with the ability of HIV to infect both the apical and basolateral surfaces of these epithelial cells; and (iv) in thin-layer chromatography-immunostaining experiments with neutral glycolipids prepared from HT29 cells, the antibody specifically reacted with a ceramide monoglycoside fraction corresponding to galactosyl ceramide. We did not detect this glycolipid in lymphoid cells, and anti-galactosyl ceramide antibodies consistently failed to inhibit HIV infection of these cells. These data suggest that galactosyl ceramide (or a derivative) is an essential component of the receptor for HIV on the surface of HT29 cells.

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Human T-lymphoblastoid cells selected for growth in serum-free medium provide new tools for study of HIV replication and cytopathogenicity

Cited by 10 publications

References 18 publications

Inhibition of HIV-1 Infection by Human α-Defensin-5, a Natural Antimicrobial Peptide Expressed in the Genital and Intestinal Mucosae

Inhibition of HIV-1 Infection by Human α-Defensin-5, a Natural Antimicrobial Peptide Expressed in the Genital and Intestinal Mucosae

Inhibition of human immunodeficiency virus infection in human colon epithelial cells by recombinant interferon‐γ

Galactosyl ceramide (or a closely related molecule) is the receptor for human immunodeficiency virus type 1 on human colon epithelial HT29 cells

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