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Characterization of the interferon-producing cell in mice infected with Listeria monocytogenes.

Stockinger S, Kastner R, Kernbauer E, Pilz A, Westermayer S, Reutterer B, Soulat D, Stengl G, Vogl C, Frenz T, Waibler Z, Taniguchi T, Rülicke T, Kalinke U, Müller M, Decker T - PLoS Pathog. (2009)

Bottom Line: Synthesis of serum IFN-I did not require TLR9.In contrast, in vitro-differentiated pDC infected with L. monocytogenes needed TLR9 to transcribe IFN-I mRNA.Based on these data, we propose that the engagement of pDC, the mode of IFN-I mobilization, as well as the shaping of the antimicrobial innate immune response by IFN-I differ between intracellular pathogens.

View Article: PubMed Central - PubMed

Affiliation: Max F. Perutz Laboratories, Department of Microbiology and Immunobiology, University of Vienna, Vienna, Austria.

ABSTRACT
Production of type I interferons (IFN-I, mainly IFNalpha and IFNbeta) is a hallmark of innate immune responses to all classes of pathogens. When viral infection spreads to lymphoid organs, the majority of systemic IFN-I is produced by a specialized "interferon-producing cell" (IPC) that has been shown to belong to the lineage of plasmacytoid dendritic cells (pDC). It is unclear whether production of systemic IFN-I is generally attributable to pDC irrespective of the nature of the infecting pathogen. We have addressed this question by studying infections of mice with the intracellular bacterium Listeria monocytogenes. Protective innate immunity against this pathogen is weakened by IFN-I activity. In mice infected with L. monocytogenes, systemic IFN-I was amplified via IFN-beta, the IFN-I receptor (IFNAR), and transcription factor interferon regulatory factor 7 (IRF7), a molecular circuitry usually characteristic of non-pDC producers. Synthesis of serum IFN-I did not require TLR9. In contrast, in vitro-differentiated pDC infected with L. monocytogenes needed TLR9 to transcribe IFN-I mRNA. Consistent with the assumption that pDC are not the producers of systemic IFN-I, conditional ablation of the IFN-I receptor in mice showed that most systemic IFN-I is produced by myeloid cells. Furthermore, results obtained with FACS-purified splenic cell populations from infected mice confirmed the assumption that a cell type with surface antigens characteristic of macrophages and not of pDC is responsible for bulk IFN-I synthesis. The amount of IFN-I produced in the investigated mouse lines was inversely correlated to the resistance to lethal infection. Based on these data, we propose that the engagement of pDC, the mode of IFN-I mobilization, as well as the shaping of the antimicrobial innate immune response by IFN-I differ between intracellular pathogens.

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Serum levels of IFN-I in mice deficient in components of the IFN-I signaling pathway.Mice of the indicated genotypes were injected intraperitoneally (i.p.) with 5×106 L. monocytogenes (C57BL/6 n = 25, IFNβ−/− n = 25, IFNAR1−/− n = 24, IRF3−/− n = 25, IRF7−/− n = 20) or PBS as a control (data not shown). After 24 h, serum was collected and ELISAs for IFNβ (A) and IFNα (B) were performed. The data presented here are a summary of several individual experiments with groups of four to five infected mice per genotype. Data representing IFNβ (A) and IFNα (B) concentrations were log-transformed (after adding one) to achieve approximate normality. Linear models with genotype and experiment as fixed effects were fitted using SPSS. Means plus/minus standard errors of wt and mutant genotypes are plotted (after back-transformation). Significant values are indicated by: n.s. not significant p>0.05, * p≤0.05, ** p≤0.01, *** p≤0.001.
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ppat-1000355-g001: Serum levels of IFN-I in mice deficient in components of the IFN-I signaling pathway.Mice of the indicated genotypes were injected intraperitoneally (i.p.) with 5×106 L. monocytogenes (C57BL/6 n = 25, IFNβ−/− n = 25, IFNAR1−/− n = 24, IRF3−/− n = 25, IRF7−/− n = 20) or PBS as a control (data not shown). After 24 h, serum was collected and ELISAs for IFNβ (A) and IFNα (B) were performed. The data presented here are a summary of several individual experiments with groups of four to five infected mice per genotype. Data representing IFNβ (A) and IFNα (B) concentrations were log-transformed (after adding one) to achieve approximate normality. Linear models with genotype and experiment as fixed effects were fitted using SPSS. Means plus/minus standard errors of wt and mutant genotypes are plotted (after back-transformation). Significant values are indicated by: n.s. not significant p>0.05, * p≤0.05, ** p≤0.01, *** p≤0.001.

Mentions: Knock-out mice were used to study pathways contributing to increase of IFNβ and IFNα in the serum of mice infected with L. monocytogenes. We established 24 h post intraperitoneal (i.p.) injection of bacteria as the point of maximum IFN-I induction in preliminary experiments (data not shown); therefore, IFNβ and IFNα were measured 24 h after infection with L. monocytogenes in subsequent experiments. Serum levels of IFNβ were generally at the detection limit of the ELISA and differences between wt and the IRF3−/− or IRF7−/− genotypes could not be reliably determined. By contrast, statistically significant differences were observed comparing wt and IFNAR1−/− animals (Figure 1A). This finding most likely reflects the importance of the IFNAR for IFNβ clearance. Increased accumulation of the cytokine in the blood of IFNAR−/− mice is therefore not in contradiction to subsequent findings showing that a fraction of IFNβ production occurs via feed-forward amplification through the IFNAR (e.g. Figures 2 and 3). Serum of infected wt mice contained higher levels of IFNα than IFNβ. IFNα species were reduced in absence of IFNβ, IRF3 or IRF7 (Figure 1B). Serum IFNα was virtually absent in mice lacking IFNAR1. Collectively the data show that the levels of serum IFNα critically depend on ‘early’ IFNβ and on IFN-I signaling, hence feed-forward amplification. Bacterial loads in liver and spleen after 24 h of infection were virtually identical between wt and IFNAR−/− mice (Figure S1). Therefore, differences in IFN-I production did not result from lower numbers of infecting bacteria.


Characterization of the interferon-producing cell in mice infected with Listeria monocytogenes.

Stockinger S, Kastner R, Kernbauer E, Pilz A, Westermayer S, Reutterer B, Soulat D, Stengl G, Vogl C, Frenz T, Waibler Z, Taniguchi T, Rülicke T, Kalinke U, Müller M, Decker T - PLoS Pathog. (2009)

Serum levels of IFN-I in mice deficient in components of the IFN-I signaling pathway.Mice of the indicated genotypes were injected intraperitoneally (i.p.) with 5×106 L. monocytogenes (C57BL/6 n = 25, IFNβ−/− n = 25, IFNAR1−/− n = 24, IRF3−/− n = 25, IRF7−/− n = 20) or PBS as a control (data not shown). After 24 h, serum was collected and ELISAs for IFNβ (A) and IFNα (B) were performed. The data presented here are a summary of several individual experiments with groups of four to five infected mice per genotype. Data representing IFNβ (A) and IFNα (B) concentrations were log-transformed (after adding one) to achieve approximate normality. Linear models with genotype and experiment as fixed effects were fitted using SPSS. Means plus/minus standard errors of wt and mutant genotypes are plotted (after back-transformation). Significant values are indicated by: n.s. not significant p>0.05, * p≤0.05, ** p≤0.01, *** p≤0.001.
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Related In: Results  -  Collection

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ppat-1000355-g001: Serum levels of IFN-I in mice deficient in components of the IFN-I signaling pathway.Mice of the indicated genotypes were injected intraperitoneally (i.p.) with 5×106 L. monocytogenes (C57BL/6 n = 25, IFNβ−/− n = 25, IFNAR1−/− n = 24, IRF3−/− n = 25, IRF7−/− n = 20) or PBS as a control (data not shown). After 24 h, serum was collected and ELISAs for IFNβ (A) and IFNα (B) were performed. The data presented here are a summary of several individual experiments with groups of four to five infected mice per genotype. Data representing IFNβ (A) and IFNα (B) concentrations were log-transformed (after adding one) to achieve approximate normality. Linear models with genotype and experiment as fixed effects were fitted using SPSS. Means plus/minus standard errors of wt and mutant genotypes are plotted (after back-transformation). Significant values are indicated by: n.s. not significant p>0.05, * p≤0.05, ** p≤0.01, *** p≤0.001.
Mentions: Knock-out mice were used to study pathways contributing to increase of IFNβ and IFNα in the serum of mice infected with L. monocytogenes. We established 24 h post intraperitoneal (i.p.) injection of bacteria as the point of maximum IFN-I induction in preliminary experiments (data not shown); therefore, IFNβ and IFNα were measured 24 h after infection with L. monocytogenes in subsequent experiments. Serum levels of IFNβ were generally at the detection limit of the ELISA and differences between wt and the IRF3−/− or IRF7−/− genotypes could not be reliably determined. By contrast, statistically significant differences were observed comparing wt and IFNAR1−/− animals (Figure 1A). This finding most likely reflects the importance of the IFNAR for IFNβ clearance. Increased accumulation of the cytokine in the blood of IFNAR−/− mice is therefore not in contradiction to subsequent findings showing that a fraction of IFNβ production occurs via feed-forward amplification through the IFNAR (e.g. Figures 2 and 3). Serum of infected wt mice contained higher levels of IFNα than IFNβ. IFNα species were reduced in absence of IFNβ, IRF3 or IRF7 (Figure 1B). Serum IFNα was virtually absent in mice lacking IFNAR1. Collectively the data show that the levels of serum IFNα critically depend on ‘early’ IFNβ and on IFN-I signaling, hence feed-forward amplification. Bacterial loads in liver and spleen after 24 h of infection were virtually identical between wt and IFNAR−/− mice (Figure S1). Therefore, differences in IFN-I production did not result from lower numbers of infecting bacteria.

Bottom Line: Synthesis of serum IFN-I did not require TLR9.In contrast, in vitro-differentiated pDC infected with L. monocytogenes needed TLR9 to transcribe IFN-I mRNA.Based on these data, we propose that the engagement of pDC, the mode of IFN-I mobilization, as well as the shaping of the antimicrobial innate immune response by IFN-I differ between intracellular pathogens.

View Article: PubMed Central - PubMed

Affiliation: Max F. Perutz Laboratories, Department of Microbiology and Immunobiology, University of Vienna, Vienna, Austria.

ABSTRACT
Production of type I interferons (IFN-I, mainly IFNalpha and IFNbeta) is a hallmark of innate immune responses to all classes of pathogens. When viral infection spreads to lymphoid organs, the majority of systemic IFN-I is produced by a specialized "interferon-producing cell" (IPC) that has been shown to belong to the lineage of plasmacytoid dendritic cells (pDC). It is unclear whether production of systemic IFN-I is generally attributable to pDC irrespective of the nature of the infecting pathogen. We have addressed this question by studying infections of mice with the intracellular bacterium Listeria monocytogenes. Protective innate immunity against this pathogen is weakened by IFN-I activity. In mice infected with L. monocytogenes, systemic IFN-I was amplified via IFN-beta, the IFN-I receptor (IFNAR), and transcription factor interferon regulatory factor 7 (IRF7), a molecular circuitry usually characteristic of non-pDC producers. Synthesis of serum IFN-I did not require TLR9. In contrast, in vitro-differentiated pDC infected with L. monocytogenes needed TLR9 to transcribe IFN-I mRNA. Consistent with the assumption that pDC are not the producers of systemic IFN-I, conditional ablation of the IFN-I receptor in mice showed that most systemic IFN-I is produced by myeloid cells. Furthermore, results obtained with FACS-purified splenic cell populations from infected mice confirmed the assumption that a cell type with surface antigens characteristic of macrophages and not of pDC is responsible for bulk IFN-I synthesis. The amount of IFN-I produced in the investigated mouse lines was inversely correlated to the resistance to lethal infection. Based on these data, we propose that the engagement of pDC, the mode of IFN-I mobilization, as well as the shaping of the antimicrobial innate immune response by IFN-I differ between intracellular pathogens.

Show MeSH
Related in: MedlinePlus