Characterization of ebv‐carrying b‐cell populations in healthy seropositive individuals with regard to density, release of transforming virus and spontaneous outgrowth

Abstract

Peripheral or tonsil lymphocyte populations of EBV-seropositive donors give rise to EBV-carrying LCLs upon in vitro explantation. Such lines can arise either by a 2-step mechanism, namely release of virus from some of the explanted cells followed by infection of previously uninfected B cells, or by direct outgrowth of virus-harboring B cells (Rickinson et al., 1974; Dalens et al., 1975; Hinuma and Katsuki 1978; Katsuki et al., 1979). We observed that cells responsible for both the 2-step mechanism and for direct outgrowth are found in the purified B-cell compartment. Virus release was more frequent than direct outgrowth. The majority of virus-releasing cells were found in the low-density fraction that contains large, activated B blasts. Cells that were capable of spontaneous outgrowth in the presence of the viral inhibitor PFA and of virus-neutralizing antibody gave rise to cell lines that carried the sex chromosome marker of the original donor, rather than that of admixed cord blood lymphocyte of the opposite sex. Such cells were found in both the low- and the high-density fractions. The majority of the EBV-carrying B cells in vivo are thus low-density blasts. Rare small B cells of high density harboring EBV were capable of spontaneous outgrowth. This may be indicative of a host control mechanism that is removed upon cultivation in vitro.

Non-correlation of in vivo and in vitro parameters of Epstein-Barr virus persistence suggests heterogeneity of B cell infection.

“…Early studies using spontaneous in vitro outgrowth from T cell-depleted PBMC populations from normal donors show that this is predominantly a two-step process where virus-carrying cells undergo lytic infection early in the culture period and the virus released infects and immortalizes co-cultured normal B cells (Rickinson et al, 1977). This process is inhibited by the drug acyclovir, which blocks lytic reactivation (Collins, 1983), and by neutralizing antibody, which prevents secondary infection ; only very rarely have B cells capable of direct outgrowth in culture been detected (Lewin et al, 1987). Outgrowth of LCL is also prevented by co-cultured, autologous T cells which contain an EBV-specific cytotoxic population (Moss et al, 1978) and these cells are therefore removed or inactivated prior to culture.…”

“…This technique estimates the level of EBVcarrying B cells in the circulation at 1 : 10& to 1:10' (Lam et al, 1991). In this in vitro assay system, outgrowth of LCL can be suppressed by the viral DNA polymerase inhibitor acyclovir I. Johannessen and others I. Johannessen and others (Collins, 1983), or by serum containing neutralizing antibody, indicating that they arise indirectly by release of virus from the in vivo EBV-carrying B cells and infection of bystanding normal B cells (Rickinson et al, 1977 ;Lewin et al, 1987). Thus, only cells with the capacity to enter a productive cycle in culture are measured in this assay.…”

The Role of Methylation in the Phenotype-dependent Modulation of Epstein-Barr Nuclear Antigen 2 and Latent Membrane Protein Genes in Cells Latently Infected with Epstein-Barr Virus

“…The normal precursor of the BL cell is a likely candidate for this role. Lewin et al (1987) showed I. ERNBERG AND OTHERS recently that small, high density B cells from EBV-positive peripheral blood are capable for direct spontaneous outgrowth of LCLs carrying EBV, although this occurs infrequently. These small, presumably long-lived resting B cells in GO may be the cells with a high intrinsic methylation activity, carrying latent viral genomes in vivo in healthy EBV seropositive individuals.…”

Epstein-Barr Virus LMP2A-Induced B-Cell Survival in Two Unique Classes of EμLMP2A Transgenic Mice

“…Of the nine latent gene products detected in vitro, only EBNA1 and LMP2A transcripts are reproducibly detected in latently infected human B cells (6,34,40,46). Although latent virus has been isolated from peripheral B cells in otherwise healthy individuals (1,6,8,15,34,35,40,46), latently infected B cells may reside in bone marrow or other lymphatic sites producing latently infected progeny cells which account for the stable number of infected peripheral B cells in otherwise healthy individuals (21,23,35,50,51).…”