Infection of Normal Human Epithelial Cells by Epstein-Barr Virus

Shapiro, I.M.; Volsky, David J.

doi:10.1126/science.6298935

Cited by 48 publications

(20 citation statements)

References 12 publications

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“…The close correlation observed between the percentages of cells binding EBV and HB-5 in epithelial cell suspensions, the similar localization of virus and HB-5 at sites on the epithelial cell surface, and the evidence both from cultured cell suspensions and from tissue sections indicating a selective involvement of less differentiated cells all strongly suggest that the C3d/EBV receptor structure is expressed on epithelia in a differentiation-linked manner much as it is on B lymphocytes. Past difficulty in demonstrating the virus receptor on cultured epithelial cells (Bayliss & Wolf, 1981 ;Glaser et al, 1980;Shapiro & Volsky, 1983) might indeed be explained by the tendency of such cells to differentiate rapidly in vitro (Banks-Schlegel & Green, 1981), a process reflected in our study by detection of an EBV receptor on just 5 ~ of cells. Enrichment for undifferentiated cells may prove crucial in future studies to characterize further the HB-5/anti-B2-defined molecule on the epithelial cell membrane and to examine its putative role as an EBV receptor.…”

Section: Virus Bindingmentioning

confidence: 82%

Human Epithelial Cell Expression of an Epstein-barr Virus Receptor

Sixbey

Davis

Young

et al. 1987

Journal of General Virology

129

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SUMMARYCultured human epithelium bound and internalized radiolabelled Epstein-Barr virus (EBV) within 1 h of exposure. A similar percentage of cultured cells also were reactive with monoclonal antibodies to the EBV/C3d receptor of B lymphocytes. In cross-sections of fresh frozen, stratified epithelium, receptor expression seemed limited to the less differentiated subpopulation of cells. These findings support the notion of direct infection of epithelial cells by EBV and suggest a viral life cycle in epithelium dependent on the stage of cell differentiation. INTRODUCTIONThe concept of strict B lymphocyte tropism has been central to discussions of the biology of the Epstein-Barr virus (EBV) since the discovery of this herpesvirus in 1964. EBV's restricted host cell range is based on an apparent lack of membrane receptors for the virus on human cells other than B lymphocytes, and virus-associated diseases such as Burkitt's lymphoma, infectious mononucleosis, and diffuse lymphoproliferative disorders of the immunocompromised host all involve infections of the B cell lineage. Yet, important aspects of virus-host interaction remain unclear when viewed in terms of an absolute tropism by EBV for B lymphocytes (Miller, 1984;Rickinson, 1984). First, EBV is linked to an epithelial cell malignancy, undifferentiated nasopharyngeal carcinoma, with viral DNA present in 1009/o of the tumours worldwide, an association more striking than in Burkitt's lymphoma. Second, despite clear evidence that EBV replicates and is shed at sites in the oropharynx, lymphocytes permissive for viral replication can not be demonstrated in vivo, and certainly experimental infection of B cells in vitro results not in viral replication, but in viral latency and cell immortalization. Recent reports suggest that viral replication in epithelial cells may be a regular feature both of acute EBV infection in man and of the chronic viral carrier state (Lemon et al., 1977;Wolf et al., 1984;Greenspan et al., 1985), but the means whereby the virus gains entry into such cells has been the subject of considerable debate (Bayliss & Wolf 1980Sixbey et al., 1983;Takimoto et al., 1983). We present here evidence for viral recognition of a cell surface receptor in vitro on an undifferentiated subset of human epithelial cells, suggesting direct infection of these cells by EBV.

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Section: Virus Bindingmentioning

confidence: 82%

Human Epithelial Cell Expression of an Epstein-barr Virus Receptor

Sixbey

Davis

Young

et al. 1987

Journal of General Virology

129

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“…12,[14][15][16] This might in part result from the requirement of primary epithelial cells to be cultured under conditions favouring differentiation. In addition, these studies might have exclusively analysed primary samples with a low sensitivity to EBV infection.…”

Section: Discussionmentioning

confidence: 99%

“…12,[14][15][16] Transfection of CD21, one of the cellular ligand of EBV gp350 surface glycoprotein, in epithelial cell lines markedly increases efficiency of infection suggesting that virus binding to target cells is a limiting factor in most epithelial cell lines. 17 In the same vein, Tugizov et al reported increased infection efficiency in epithelial cell subclones selected for their high virus binding levels.…”

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confidence: 99%

Epstein‐Barr virus B95.8 produced in 293 cells shows marked tropism for differentiated primary epithelial cells and reveals interindividual variation in susceptibility to viral infection

Feederle

Nęuhierl

Bannert

et al. 2007

Intl Journal of Cancer

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Epstein-Barr virus (EBV), a well-characterised B-lymphotropic agent is aetiologically linked to B cell lymphoproliferations, but the spectrum of diseases the virus causes also includes oral hairy leukoplakia, a benign epithelial lesion, as well as carcinomas of the nasopharynx and of the stomach. However, it is still unclear how EBV accesses and transforms primary epithelial cells. Sixteen samples consisting of primary epithelial cells from the sphenoidal sinus or from tonsils were infected with GFP-tagged recombinant B95.8 EBVs produced in the 293 cell line. The rate of infection was assessed by counting GFP-positive cells and cells expressing viral proteins. Primary epithelial cells from all samples were found to be sensitive to EBV infection but there was a marked interindividual variation among the tested samples (2-48% positive cells). This suggests heterogeneity in terms of sensitivity to EBV infection in vivo and therefore possibly to EBV-associated diseases of the epithelium. The virus showed a preferential tropism for differentiated epithelial cells (p63 negative, involucrin positive). In all cases, infected cells expressed EBV lytic proteins but also the LMP1 protein. The viral tropism for differentiated cells and the permissivity of these cells for virus replication reproduced in vitro cardinal features of oral hairy leukoplakia. We have identified a source of EBV that shows unusually strong epitheliotropism for primary epithelial cells that will allow detailed analysis of virus-cell interactions during virus infection, replication and virus-mediated transformation. ' 2007 Wiley-Liss, Inc.

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“…Conjugation of the virus with fluorescein-isothiocyanate (FITC, Sigma) was generally as described by Khelifa and Menezes (7), with some modifications (17). Briefly, 500 times concentrated EBV in phosphate-buffered saline, pH 7.5 (PBS) was incubated with FITC dissolved in a carbonate buffer, pH 9.2, for 1 h a t room temperature.…”

Section: Virusesmentioning

confidence: 99%

Application of flow cytometry to studies on the distribution, persistence, and regeneration rate of Epstein‐Barr virus receptors

McCune

Volsky

1984

Cytometry

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Flow cytometry has been applied to study the persistence and regeneration rate of Epstein-Barr virus (EBV) receptors and other membrane proteins in normal and tumor cells. EBV receptors were detected in a binding assay utilizing fluorescein isothiocyanate-conjugated virions (FITC-EBV). After stripping receptors from the cell surface with trypsin, binding of FITC-EBV was rapidly regenerated in the lymphoma cell line Loukes, reaching 75% of the control levels by 3 h. Fresh human lymphocytes regenerated receptors at a much slower rate, reaching 50% of control levels by 24 h. The regenerating receptors did not reappear simultaneously on all the cells. Subpopulation of new receptor-positive cells increased gradually during the incubation at 37°C. Reappearance of receptors was inhibited in the absence of protein synthesis. Conversely, receptors persisted in the nontrypsinized cells in the absence of protein synthesis. These results show that certain parameters of membrane receptor turnover, such as the change in receptor distribution within a cell population, receptor persistance, and regeneration rate, can be measured at the single-cell level by flow cytometry.Key terms: Flow cytometry, Epstein-Barr virus (EBV), surface receptors, membrane protein turnover, fluoresceinated virus Recent approaches to the study of membrane protein turnover have involved detecting specific proteins by the time-consuming techniques of sodium dodecyl sulfate (SDS)-gel electrophoresis (5), immunoprecipitation (l), purification of individual proteins (201, and the use of fluorescent probes and fluorescence microscopy (2). Only the latter technique permits following the fate of a n individul membrane protein on single cells, but it is qualitative and insensitive. Flow cytometry offers a rapid, sensitive, and quantitative method of detecting cell surface markers at the single cell level (10). Here we describe flow cytometry as a convenient and effective tool for the study of two aspects of membrane protein turnover: the persistence of membrane receptors in the presence and absence of protein synthesis inhibitor and regeneration of membrane receptors after stripping with trypsin. We have chosen to investigate membrane receptors for Epstein-Barr virus (EBV-R) because of our interest in the host cell range and biological activity of this human tumor agent. EBV-R have been demonstrated only on human and certain other primate B lymphocytes (6,181, strengthening the association between the virus and Burkitt's lymphoma (9). However, viral genome and nuclear antigen, EBNA, have also been regularly found in nasopharyngeal carcinoma cells (8,131. This indicates that under certain pathologic conditions EBV can bind to and penetrate into human epithelial cells. Moreover, while EBV-infected lymphocytes are normally recognized and eliminated by the immune system (41, the BL and NPC cells can successfully avert immune response (14), presumably as a result of some unknown malignancy-related cell surface changes. We have compared the persistence and regenerat...

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Infection of Normal Human Epithelial Cells by Epstein-Barr Virus

Cited by 48 publications

References 12 publications

Human Epithelial Cell Expression of an Epstein-barr Virus Receptor

Human Epithelial Cell Expression of an Epstein-barr Virus Receptor

Epstein‐Barr virus B95.8 produced in 293 cells shows marked tropism for differentiated primary epithelial cells and reveals interindividual variation in susceptibility to viral infection

Application of flow cytometry to studies on the distribution, persistence, and regeneration rate of Epstein‐Barr virus receptors

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