Distinct Responses of Lung and Spleen Dendritic Cells to the TLR9 Agonist CpG Oligodeoxynucleotide

Synthetic oligodeoxyribonucleotides (ODNs) containing unmethylated CpG recapitulate the activation of TLR9 by microbial DNA. ODNs are potent stimulators of the immune response in cells expressing TLR9. Despite extensive use of mice as experimental animals in basic and applied immunological research, the key sequence determinants that govern the activation of mouse TLR9 by ODNs have not been well defined. We performed a systematic investigation of the sequence motif of B class phosphodiester ODNs to identify the sequence properties that govern mouse TLR9 activation. In contrast to ODNs activating human TLR9, where the minimal sequence motif for the receptor activation comprises a pair of closely positioned CpGs we found that the mouse TLR9 requires a single CpG positioned 4–6 nt from the 5′-end. Activation is augmented by a 5′TCC sequence one to three nucleotides from the CG. The distance of the CG dinucleotide of four to six nucleotides from the 5′-end and the ODN’s length fine-tunes activation of mouse macrophages. Length of the ODN <23 and >29 nt decreases activation of dendritic cells. The ODNs with minimal sequence induce Th1-type cytokine synthesis in dendritic cells and confirm the expression of cell surface markers in B cells. Identification of the minimal sequence provides an insight into the sequence selectivity of mouse TLR9 and points to the differences in the receptor selectivity between species probably as a result of differences in the receptor binding sites.

“…3E-H). The activation of TLR9 in DC subtypes is in agreement with the expression profile of TLR9 in mice DC subtypes: pDC, cDCs, and myeloid DCs (31).…”

Section: Response Of Mouse Dcs To the Minm Odnsupporting

confidence: 62%

Species-Specific Minimal Sequence Motif for Oligodeoxyribonucleotides Activating Mouse TLR9

Pohar

Lainšček

Fukui

et al. 2015

“…In addition, it has recently been demonstrated that both myeloid DCs and pDCs in the lung show no detectable expression of TLR9 while both subsets in the spleen express TLR9 (48). Consequently, CpG oligonucleotides exerted differential effects on lung and spleen DCs when administered to mice (48). Similarly, in our study, the absence of TLR9 resulted in differential effects in the lung and spleen.…”

Section: Discussionsupporting

confidence: 52%

“…However, it has been shown by Northern blot analysis that TLR9 is much stronger expressed in the spleen than in the lung and a variety of other tissues (47). In addition, it has recently been demonstrated that both myeloid DCs and pDCs in the lung show no detectable expression of TLR9 while both subsets in the spleen express TLR9 (48). Consequently, CpG oligonucleotides exerted differential effects on lung and spleen DCs when administered to mice (48).…”

Section: Discussionmentioning

confidence: 99%

“…It has been hypothesized that the biological significance of the absence of TLR9 in lung DCs may be a protective measure that has evolved to protect the lung from the development of diseases that are associated with high cytokine (IL-6) production such as pulmonary fibrosis (48). Thus, it is tempting to speculate that the same applies to MHV-68 lung infection, namely that the "physiological" absence of TLR9 in lung DCs may prevent an overwhelming innate immune response, for example, inflammatory cytokine production, and thereby limits pulmonary disease while having only little effect on virus replication.…”

Section: Discussionmentioning

confidence: 99%

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TLR9 Contributes to Antiviral Immunity during Gammaherpesvirus Infection

Guggemoos

Hangel

Hamm

et al. 2008

The human gammaherpesviruses Kaposi’s sarcoma-associated herpesvirus and EBV cause important infections. As pathogenetic studies of the human infections are restricted, murine gammaherpesvirus 68 serves as a model to study gammaherpesvirus pathogenesis. TLRs are a conserved family of receptors detecting microbial molecular patterns. Among the TLRs, TLR9 recognizes unmethylated CpG DNA motifs present in bacterial and viral DNA. The aim of this study was to assess the role of TLR9 in gammaherpesvirus pathogenesis. Upon stimulation with murine gammaherpesvirus 68, Flt3L-cultured bone marrow cells (dendritic cells) from TLR9−/− mice secreted reduced levels of IL-12, IFN-α, and IL-6, when compared with dendritic cells from wild-type mice. Intranasal infection of TLR9−/− and wild-type mice did not reveal any differences during lytic and latent infection. In contrast, when infected i.p., TLR9−/− mice showed markedly higher viral loads both during lytic and latent infection. Thus, we show for the first time that TLR9 is involved in gammaherpesvirus pathogenesis and contributes to organ-specific immunity.

“…This results in impaired T cell stimulation of both CD4 ϩ and CD8 ϩ T cells and contributes to the reduced pulmonary inflammatory response to inhaled Ags. The expression of TLR9 and the response of lung DC to CpG have been previously demonstrated by others (34,35). In contrast to studies using splenic or peripheral lymph node DC after systemic CpG injection (26), CpG-induced maturation of lung DC did not down-regulate their constitutive MHC class II-mediated Ag presentation.…”

Section: Discussionmentioning

confidence: 60%

Impaired Lung Dendritic Cell Migration and T Cell Stimulation Induced by Immunostimulatory Oligonucleotides Contribute to Reduced Allergic Airway Inflammation

Constabel

Stankov

Hartwig

et al. 2009

CpG-containing oligonucleotides (CpG) have been shown to reduce key features of allergic airway inflammation in mouse models. Given the inhibitory effects of CpG treatment on Ag presentation of subsequently encountered Ags via MHC class I and II molecules by dendritic cells (DC), we hypothesized that intranasal CpG treatment would lead to reduced Ag-specific T cell stimulation in the lung-draining lymph nodes, thereby reducing the inflammatory response in sensitized mice. Intranasal CpG administration led to phenotypic maturation of lung and mediastinal lymph node DC as determined by expression of MHC class II, CD80, and CD86. This was accompanied by a significant reduction in the proliferation of adoptively transferred Ag-specific CD4+ and CD8+ T cells in mediastinal lymph nodes, when CpG was given before inhalative OVA challenges. DC obtained from mediastinal lymph nodes of CpG-treated mice before OVA inhalation led to reduced T cell stimulation via MHC class I and II molecules. In addition, CpG diminished airway eosinophilia and pulmonary infiltration after sensitization or following adoptive transfer of Ag-specific Th2 cells. These results were explained by reduced CCL21 expression and inhibition of lung DC migration following CpG administration, which could be restored by transfer of bone marrow-derived DC, because CpG had no major impact on the constitutive MHC class II Ag presentation of protein-derived Ag by lung tissue-derived DC. We conclude that CpG treatment can effectively impair the DC-mediated Ag transport from the lungs to the lymph nodes, resulting in reduced T cell activation and blunted airway inflammation.