title: A simple immunomonitoring procedure for mRNA-loaded dendritic cell therapy.
abstract: To develop a simple immunomonitoring method for dendritic cell therapy using messenger RNA (mRNA) as antigen, we evaluated whether Mycobacterium tuberculosis antigen 85A (Ag85A) mRNA-transfected peripheral blood mononuclear cells (PBMCs) could be used to stimulate the induction of interferon (IFN)-gamma-producing T cells. PBMCs from 10 healthy donors were cocultured with autologous PBMCs transfected with mRNA overnight, and the number of IFN-gamma-producing T cells was measured by flow cytometry. IFN-gamma-producing CD4+ and CD8+ T cells were detected in 4 and 5 donors, respectively. PBMCs from 3 donors with negative results were then cocultured with Ag85A mRNA-transfected autologous PBMCs for 1 week to achieve in vitro primary induction of Ag85A-specific T cells. After restimulation with freshly prepared stimulator cells, a small but significant number of IFN-gamma-producing CD8+ T cells was detected. The induction of IFN-gamma-producing CD8+ T cells by overnight coculture was completely abolished by anti-class II or anti-interleukin-12 antibodies, whereas it was partially inhibited by anti-class I antibody. These data suggest that Ag85A mRNA-transfected PBMCs induce specific IFN-gamma-producing T cells and might be applicable for immunomonitoring of mRNA-loaded dendritic cell therapy.
authors.affiliation: Department of Laboratory Medicine, Yamaguchi University Graduate School of Medicine, Ube, Japan.
authors.forename: Mikako
authors.initials: M
authors.lastname: Hiura
authors.forename: Koji
authors.initials: K
authors.lastname: Ueno
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Suehiro
authors.forename: Shoichi
authors.initials: S
authors.lastname: Hazama
authors.forename: Masaaki
authors.initials: M
authors.lastname: Oka
authors.forename: Kohzoh
authors.initials: K
authors.lastname: Imai
authors.forename: Yuji
authors.initials: Y
authors.lastname: Hinoda
mesh_terms: D000217:Acyltransferases / Q000235:genetics*; D000942:Antigens, Bacterial / Q000235:genetics*; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism*; D018414:CD8-Positive T-Lymphocytes / Q000378:metabolism*; D018920:Coculture Techniques; D003713:Dendritic Cells / Q000378:metabolism*; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000235:genetics; D015234:HLA-A Antigens / Q000378:metabolism; D015235:HLA-B Antigens / Q000378:metabolism; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000378:metabolism*; D008213:Lymphocyte Activation; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012882:Skin Tests; D014162:Transfection; D014374:Tuberculin Test
chemical_list: D000942:Antigens, Bacterial; D015234:HLA-A Antigens; D015235:HLA-B Antigens; D012333:RNA, Messenger; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D007371:Interferon-gamma; D000217:Acyltransferases; C112580:antigen 85A, Mycobacterium tuberculosis