title: Induction of cytomegalovirus-specific CD4+ cytotoxic T lymphocytes from seropositive or negative healthy subjects or stem cell transplant recipients.
abstract: 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.
authors.affiliation: Department of Hematology, Tokai University School of Medicine, Kanagawa, Japan.
authors.forename: Kei
authors.initials: K
authors.lastname: Tazume
authors.forename: Masao
authors.initials: M
authors.lastname: Hagihara
authors.forename: Balgansuren
authors.initials: B
authors.lastname: Gansuvd
authors.forename: Ayako
authors.initials: A
authors.lastname: Higuchi
authors.forename: Yoko
authors.initials: Y
authors.lastname: Ueda
authors.forename: Kaoru
authors.initials: K
authors.lastname: Hirabayashi
authors.forename: Motoharu
authors.initials: M
authors.lastname: Hojo
authors.forename: Akiko
authors.initials: A
authors.lastname: Tanabe
authors.forename: Ayako
authors.initials: A
authors.lastname: Okamoto
authors.forename: Shunichi
authors.initials: S
authors.lastname: Kato
authors.forename: Tomomitsu
authors.initials: T
authors.lastname: Hotta
mesh_terms: 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*
chemical_list: D000956:Antigens, Viral; D006684:HLA-DR Antigens