Induction of cytomegalovirus-specific CD4+ cytotoxic T lymphocytes from seropositive or negative healthy subjects or stem cell transplant recipients.
OBJECTIVE
We generated cytomegalovirus (CMV)-specific cytotoxic T lymphocytes (CTL) in vitro using dendritic cells (DC) pulsed with crude CMV antigens (Ag).


METHODS
Mononuclear cells from healthy CMV-seropositive or seronegative volunteers and from stem cell transplant (SCT) recipients were cultured with CD14(+) monocyte-derived DC prepulsed with CMV Ag and then matured in vitro with lipopolysaccharide and tumor necrosis factor-alpha. After proliferation, cells were checked for phenotype (CD4/CD8), while killing activity was measured by 51Cr-release assay.


RESULTS
CD4(+) T cells, the main proliferating cells from both seropositive and seronegative individuals, killed autologous Ag-pulsed DC but not vehicle-pulsed autologous DC or CMV-pulsed allogeneic DC. Similar CTL induction was accomplished from SCT recipients. Significant killing of autologous CMV-infected fibroblasts required 16-hour incubation as opposed to the standard 4-hour incubation, which was prevented by either a perforin inhibitor or anti-Fas ligand monoclonal antibody. CTL enhanced surface HLA-DR expression of CMV-infected fibroblasts, and their activity was neutralized by anti-HLA-DR monoclonal antibody.


CONCLUSIONS
CMV-specific CD4(+) CTL were inducible with or without antiviral humoral immunity, even from immunosuppressed SCT recipients. These CTL showed perforin- and Fas/Fas ligand-mediated cytotoxicity after long-term (16-hour) contact with CMV-infected targets.
Tazume
Kei
K
Department of Hematology, Tokai University School of Medicine, Kanagawa, Japan.
Hagihara
Masao
M
Gansuvd
Balgansuren
B
Higuchi
Ayako
A
Ueda
Yoko
Y
Hirabayashi
Kaoru
K
Hojo
Motoharu
M
Tanabe
Akiko
A
Okamoto
Ayako
A
Kato
Shunichi
S
Hotta
Tomomitsu
T
D000956:Antigens, Viral / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D003587:Cytomegalovirus / Q000276:immunology*; D003586:Cytomegalovirus Infections / Q000276:immunology*; D003602:Cytotoxicity, Immunologic; D003713:Dendritic Cells / Q000502:physiology; D006684:HLA-DR Antigens / Q000032:analysis; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D000956:Antigens, Viral; D006684:HLA-DR Antigens