ABSTRACT
BACKGROUND: CpG oligodeoxynucleotides (ODNs), resembling bacterial DNA, are currently tested in clinical trials as vaccine adjuvants. They have the nuclease-resistant phosphorothioate bond; the immune responses elicited differ according to the CpG ODN sequence and vaccination method. To develop a CpG ODN that can induce plasmacytoid dendritic cell (pDC)-mediated T(H)1 immunity through the mucosa, we constructed phosphodiester G9.1 comprising one palindromic CpG motif with unique polyguanosine-runs that allows degradation similar to naturally occurring bacterial DNA. METHODS: T(H)1 and T(H)2 immunity activation was evaluated by cytokine production pattern and T-bet/GATA-3 ratio in human peripheral blood mononuclear cells and mouse bone marrow cells. Adjuvanticity was evaluated in mice administered G9.1 with diphtheria toxoid (DT) through nasal vaccination. RESULTS: G9.1 exhibited stronger IFN-α-inducing activity than A-class CpG ODN2216 and increased T-bet/GATA-3 ratio by enhancing T-bet expression. Nasally administered G9.1 plus DT induced DT-specific mucosal IgA and serum IgG, but not IgE, responses with antitoxin activity in C57BL/6 and BALB/c mice, possibly due to IFN/BAFF production. Induction of T(H)1, but not T(H)2-type Abs depended completely on pDCs, the first in vivo demonstration by CpG ODNs. CONCLUSIONS: G9.1 is a promising mucosal adjuvant for induction of pDC-mediated T(H)1 immunity.
Subject(s)
Adjuvants, Immunologic/pharmacology , Adjuvants, Pharmaceutic/pharmacology , Dendritic Cells/immunology , Mucous Membrane/immunology , Oligodeoxyribonucleotides/immunology , Th1 Cells/immunology , Animals , Bone Marrow Cells/drug effects , Bone Marrow Cells/immunology , DNA, Bacterial/immunology , Dendritic Cells/drug effects , Diphtheria Toxoid/immunology , Female , Humans , Interferon-alpha/immunology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mucous Membrane/drug effects , Th1 Cells/drug effectsABSTRACT
Human plasmacytoid or CD4(+)CD11c(-) type 2 dendritic cell precursors (PDC) were identified as natural type I interferon (IFN)-producing cells in response to viral and bacterial infection. They represent effector cells of innate immunity and link it to the distinct adaptive immunity by differentiating into mature DC. It has been reported that oligodeoxyribonucleotides containing unmethylated CpG motifs (CpG DNA) stimulate PDC to produce IFN-alpha, but the molecular mechanisms involved remain unknown. We found that CpG-DNA-induced IFN-alpha production in PDC was completely impaired by the inhibitor of the p38 mitogen-activated protein kinase (MAPK) pathway. Expression of IFN regulatory factor (IRF)-7 was enhanced by CpG-DNA treatment, which was preceded by the phosphorylation of signal transducer and activator of transcription (STAT)1 on Tyr-701, as well as its enhanced phosphorylation on Ser-727. All of these events were also suppressed by the p38 MAPK inhibitor. STAT1, STAT2, and IRF-9, components of IFN-stimulated gene factor 3 (ISGF3), were recognized in the nuclear fraction of CpG-DNA-treated cells. Neither anti-IFN-alpha/beta antibodies (Ab) nor anti-IFNAR Ab suppressed STAT1 phosphorylation, enhancement of IRF-7 expression, or IFN-alpha production in the early phase of the culture. These results suggest that CpG DNA induces p38 MAPK-dependent phosphorylation of STAT1 in a manner independent of IFN-alpha/beta, which may cause ISGF3 formation to increase the transcription of the IRF-7 gene, thereby leading to IFN-alpha production in human PDC.
