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Blood, 15 February 2001, Vol. 97, No. 4, pp. 994-1000
IMMUNOBIOLOGY
Induction of cytomegalovirus (CMV)-specific T-cell responses
using dendritic cells pulsed with CMV antigen: a novel culture system
free of live CMV virions
Karl Peggs,
Stephanie Verfuerth, and
Stephen Mackinnon
From the Department of Haematology, University College
London, London, United Kingdom.
Recipients of allogeneic transplants are at risk of cytomegalovirus
(CMV) infection and disease during the period of immune compromise
after transplantation. The limitations of current antiviral pharmacotherapy have led to attempts to develop alternative strategies for preventing or treating CMV infection, such as adoptive transfer of
donor-derived virus-specific T cells. Methods for generating CMV-specific T cells either use live CMV to infect autologous antigen-presenting cells (APCs) or require some knowledge of the immunodominant peptides involved in the immune response. A novel culture system was developed that does not use live virions and in
which the APCs are monocyte-derived dendritic cells (DCs). APCs were
pulsed with CMV antigen and cocultured with autologous peripheral blood
lymphocytes from donors seropositive for CMV. The culture-output cells
consisted of both CD4- and CD8-expressing T cells. Proliferation, as
determined by a tritium-thymidine-incorporation assay, showed
significant CMV-antigen specificity in cultures from 15 of 15 donors
seropositive for CMV. In cytotoxicity assays, cytotoxic T lymphocytes
from 10 of 12 cocultures specifically lysed autologous CMV-infected
fibroblasts or DCs but not HLA-mismatched or uninfected target cells,
and this activity was shown to be blocked by HLA class 1 blocking
antibodies. T-cell-receptor spectratyping of cells from the cultures
typically showed complex size-distribution patterns, with all size
classes of a normal preculture distribution present. However, a few
size-class peaks were expanded compared with the preculture patterns
and these may have represented expansions of CMV-specific T-cell
clones. Advantages of this culture system are that it requires no live
virions and no detailed knowledge of the antigenic peptides involved
and it is applicable to CMV-seropositive donors of any HLA type.

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