A study of Epstein-Barr virus (EBV)-transformed cells in vitro

Benjamin, Pearline A. (1981) A study of Epstein-Barr virus (EBV)-transformed cells in vitro. PhD thesis, University of Glasgow.

Abstract

The aim of my studies for this thesis was to establish and characterize pairs of EBV-transformed lymphoblastoid cell lines (LCLs) which resulted from the transformation of human umbilical cord blood lymphocyte (HUCL) populations, by different strains of EBV. EBV-transformed human LCLs have been shown to represent bone marrow (B)-derived cell populations. Moreover, the immunoglobulin (ig) synthesizing cells vitro among LCLs are thought to represent populations of lymphocytes which had been synthesizing Ig in vivo (Steel et. al., (1977)). LCLs are also thought to contain in their populations, other B-cell types. Finally, different strains of EBV are thought to have different biological characteristics in vitro. Two main premises to be tested were :- (1) strains of EBV have different characteristics in vitro and (11) EBV transforms both Ig as well as non-Ig synthesizing cells, that is, individual LCLs should represent different populations of Ig and non-Ig synthesizing cells vivo. Each pair of cell lines was obtained from one common pool of HUCL population. HUCL populations have been shown to be devoid of natural killer cells and EBV-specific cytotoxic thymus (T)-derived cells. This feature of HUCL populations, renders them excellent tools for the, study of EBV-induced transformation vitro. I established and characterized 16 pairs of LCLs. One of two pools from each of 16 samples of HUCL populations was either EBV (B958) or EBV (QIMR-WIL) transformed. Transformed cells were characterized after the establishment of the transformed state as judged by acidity in the growth medium, coupled with the appearance of transformed foci of cells. The time lapse between infection of HUCLs and the establishment of the transformed state was shorter among EBV (B958) than among EBV (QIMR-WIL) transformed cells. The difference in the time lapse among the two groups of infected cells was attributable to differences in the transforming efficiences (TE). While EBV B958 had TE of 103.4 transforming dose50/ml (TD50/ml), TE for EBV QIMR-WIL was found to be 101.0 TD50/ml. The higher TE for EBV B958 was attributed to the characteristic features of the virus and the B958 cells. It was found that more than one EBV DNA core could become encapsidated in the EBV B95S capsid. This feature could contribute to the high transforming efficiency for EBV B95S,, B958 cells were found to survive for up to ten days in culture without medium replenishment, and a high viability among the cell populations retained. On electron micrographs, encapsidated EBV B958 particles were seen attached to the surfaces of undisrupted B958 cells. A cyclical phenomenon was proposed whereby, EBV B958 particles once released from disrupted cells, reinfect viable cells still in culture. The reinfected cells subsequently lysed and released viral particles, thus contributing to the maintenance of a high TE for EBV B958. All 32 cell lines were found to synthesize the EBV- associated nuclear antigen, EBNA. There was no significant difference in the percentages of EBNA synthesizing cells between EBV (B958) and EBV (QIMR-WIL) transformed cells. However, EBV (B958) transformed cell lines were found to represent a higher number of producer cell lines than EBV (QIMR-WIL) transformed cell lines. EBV (B958) transformed cell populations had higher percentages of Ig synthesizing cells than EBV (QIMR-WIL) transformed cell populations. EBV (B958) transformed cells had percentages of IgM and IgG synthesizing cells which were comparable to the percentages of IgM and IgG synthesizing cells among uninfected, untransformed HUCL populations. On the other hand, EBV (QIMR-WIL) transformed cell populations had fewer Ig synthesizing cells than uninfected, untransformed HUCL populations. Among long-term cultures of both EBV (B958) and EBV (QIMR-WIL) transformed cultures, populations of cells were selected for continual proliferation, which had higher percentages of EBV-associated early antigen positive (EA)+ cells, than newly established cultures. Among these (EA)+ cell populations, there was also increased percentages of cytoplasmic Ig positive cells. It was therefore postulated that EBV-transformed cells in culture could proliferate and synthesize Ig as a direct response to EBV-associated antigenic stimulation. One factor was very obvious among long-term cultures which were analyzed by SDS-polyacrylamide gel-electrophoresis. Irrespective of the EBV strain which was used to induce transformation, the cells which survived in culture, were IgM synthesizing.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Additional Information: Adviser: A R Williamson
• Keywords: Virology.
• Colleges/Schools: College of Medical Veterinary and Life Sciences
• Supervisor's Name: Supervisor, not known
• Date of Award: 1981
• Depositing User: Enlighten Team
• Unique ID: glathesis:1981-72036
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 17 May 2019 13:17
• Last Modified: 25 Aug 2022 10:17
• URI: https://theses.gla.ac.uk/id/eprint/72036