Identification of the epithelial cell as the cell permissive for EBV replication in the oropharynx has reordered our perception of EBV biology. We will reexamine the phenomena of EBV-induced growth transformation, persistance and reactivation in the context of epithelial cell-lymphocyte interactions. To this end, our specific aims are: (1) to determine the relationship between EBV activation in epithelium and cellular expression of differentiation antigens; (2) to examine the role of latent EBV infection in the induction of epithelial hyperproliferation; (3) to analyses lymphocyte-epithelial cell interactions in the epidermis with regard to EBV reactivation and intercellular exchange of virus. Laboratory and wild-type EBV will be used to infect epithelial primary explant cultures capable of in vitro differentiation. To correlate stage of cell maturation with EBV antigen expression, we will use monoclonal antibodies to cell differentiation antigens (keratin, involucrin). Functional interrelationships between viral proteins and elements of epithelial cytoskeleton transiently expressed during cell differentiation will be examined by immunoelectron microscopy and microinjection. Expression of the EBV latent membrane protein, important in cell transformation, will be examined in epithelium.
Under aim #2, EBV's ability to induce proliferation of basal epithelial cells in a manner analogous to B cells immortalization will be examined in epithelial grafts in nude mice and in hyperplastic epithelial lesions of the uterine cervix. Evidence for an etiologic role of EBV in epithelial proliferation will be obtained by hybridization analysis for EBV gene products known to induce cell transformation; analysis of cell clonality using the structure of EBV termini; and determination of EBV DNA organization in proliferating cells (integrated, episomal, linear).
In aim #3, the ability of EBV to reactivate in latently infected B cells in the epidermis will be analysed by cocultivation of the two cell types and by cytohybridization to inflammatory infiltrates of the cervix. The issue of enogenous spread of EBV by B cells will be further addressed by analysis of EBV isolates, shed from separate anatomical sites, for restriction fragment length polymorphisms. Unlike in lymphocytes, the entire viral cycle - from latency to replication - is manifest in epithelial cells. These studies will provide new information on EBV pathobiology and cell regulation of the viral gene expression.
