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Targeting of a Fixed Bacterial Immunogen to Fc Receptors Reverses the Anti-Inflammatory Properties of the Gram-Negative Bacterium, Francisella tularensis, during the Early Stages of Infection.

Babadjanova Z, Wiedinger K, Gosselin EJ, Bitsaktsis C - PLoS ONE (2015)

Bottom Line: It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity.The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date.These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Seton Hall University, South Orange, New Jersey, United States of America.

ABSTRACT
Production of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

No MeSH data available.


Related in: MedlinePlus

Administration of mAb-iFt immune complexes reverses the anti-inflammatory properties of LVS in the lungs of immunized mice.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. Lung tissue homogenates were obtained from immunized mice 24, 48 and 96 hours post-infection as indicated above and spun down at 15,000g for 30 minutes at room temperature to remove tissue debris. Cytokine levels were detected by using the IL-6, IL-10, TNF-α and IFN-γ ELISA kits and following vendor instructions (Biolegend). Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.
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pone.0129981.g005: Administration of mAb-iFt immune complexes reverses the anti-inflammatory properties of LVS in the lungs of immunized mice.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. Lung tissue homogenates were obtained from immunized mice 24, 48 and 96 hours post-infection as indicated above and spun down at 15,000g for 30 minutes at room temperature to remove tissue debris. Cytokine levels were detected by using the IL-6, IL-10, TNF-α and IFN-γ ELISA kits and following vendor instructions (Biolegend). Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.

Mentions: The balance and kinetics of pro- and anti-inflammatory cytokine secretion during F. tularensis challenge are key players in controlling the outcome of infection [18, 32, 33]. Consequently, having shown that immunization of mice with mAb-iFt favors a pro-inflammatory cytokine profile secreted by PECs in vitro, we attempted to determine the effect of our FcγR targeting approach on the levels of inflammatory cytokines in the lungs of immunized mice during the early stages of LVS infection. To accomplish this, C57BL/6 mice were immunized with PBS, or iFt, or mAb-iFt, boosted on day 21 and infected with a lethal dose of LVS on day 35 post-immunization. The lungs of euthanized mice were harvested after 24, 48 and 96 hours post-challenge, homogenized, and the IL-6, IL-10, TNF-α and IFN-γ cytokine levels were measured by direct sandwich ELISAs. Our results indicate a faster pro-inflammatory response in the lungs of mAb-iFt immunized mice compared to mice immunized with iFt alone, as assessed by the IL-6 and TNF-α production kinetics. Levels of IL-6 and TNF-α peaked at 48 hours post-infection, while they were significantly decreased by day 4 post-challenge (Fig 5). This observation was accompanied by a drop of the bacterial load (data not shown). On the other hand, the pro-inflammatory cytokine levels tested continued to increase at 96 hours post infection in both the PBS and iFt immunized mice accompanied by an increase in the bacterial burden in the lungs (Fig 5 and data not shown). Similar results among the three groups of mice were noted with IFN-γ (Fig 5). On the contrary, the levels of IL-10 were significantly higher in mice immunized with either iFt or PBS versus mAb-iFt within the first 24 of infection, indicating the early anti-inflammatory properties of F. tularensis LVS. Importantly, the decrease of IL-10 in the mAb-iFt immunized mice observed at 96 hours post-infection was consistent with our previous observation, indicating that FcγR targeting shifts towards a pro-inflammatory cytokine profile at the early stages of F. tularensis infection (Figs 2 and 5).


Targeting of a Fixed Bacterial Immunogen to Fc Receptors Reverses the Anti-Inflammatory Properties of the Gram-Negative Bacterium, Francisella tularensis, during the Early Stages of Infection.

Babadjanova Z, Wiedinger K, Gosselin EJ, Bitsaktsis C - PLoS ONE (2015)

Administration of mAb-iFt immune complexes reverses the anti-inflammatory properties of LVS in the lungs of immunized mice.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. Lung tissue homogenates were obtained from immunized mice 24, 48 and 96 hours post-infection as indicated above and spun down at 15,000g for 30 minutes at room temperature to remove tissue debris. Cytokine levels were detected by using the IL-6, IL-10, TNF-α and IFN-γ ELISA kits and following vendor instructions (Biolegend). Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.
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Related In: Results  -  Collection

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pone.0129981.g005: Administration of mAb-iFt immune complexes reverses the anti-inflammatory properties of LVS in the lungs of immunized mice.C57BL/6 mice were immunized i.n. with PBS, iFt (2x107 CFUs), or mAb-iFt, boosted on day 21 and challenged on day 35 with 10,000 CFUs of Ft LVS. Lung tissue homogenates were obtained from immunized mice 24, 48 and 96 hours post-infection as indicated above and spun down at 15,000g for 30 minutes at room temperature to remove tissue debris. Cytokine levels were detected by using the IL-6, IL-10, TNF-α and IFN-γ ELISA kits and following vendor instructions (Biolegend). Results are representative of three independent experiments. (*) P-value < 0.1; (**) P-value < 0.05; bars represent the SD.
Mentions: The balance and kinetics of pro- and anti-inflammatory cytokine secretion during F. tularensis challenge are key players in controlling the outcome of infection [18, 32, 33]. Consequently, having shown that immunization of mice with mAb-iFt favors a pro-inflammatory cytokine profile secreted by PECs in vitro, we attempted to determine the effect of our FcγR targeting approach on the levels of inflammatory cytokines in the lungs of immunized mice during the early stages of LVS infection. To accomplish this, C57BL/6 mice were immunized with PBS, or iFt, or mAb-iFt, boosted on day 21 and infected with a lethal dose of LVS on day 35 post-immunization. The lungs of euthanized mice were harvested after 24, 48 and 96 hours post-challenge, homogenized, and the IL-6, IL-10, TNF-α and IFN-γ cytokine levels were measured by direct sandwich ELISAs. Our results indicate a faster pro-inflammatory response in the lungs of mAb-iFt immunized mice compared to mice immunized with iFt alone, as assessed by the IL-6 and TNF-α production kinetics. Levels of IL-6 and TNF-α peaked at 48 hours post-infection, while they were significantly decreased by day 4 post-challenge (Fig 5). This observation was accompanied by a drop of the bacterial load (data not shown). On the other hand, the pro-inflammatory cytokine levels tested continued to increase at 96 hours post infection in both the PBS and iFt immunized mice accompanied by an increase in the bacterial burden in the lungs (Fig 5 and data not shown). Similar results among the three groups of mice were noted with IFN-γ (Fig 5). On the contrary, the levels of IL-10 were significantly higher in mice immunized with either iFt or PBS versus mAb-iFt within the first 24 of infection, indicating the early anti-inflammatory properties of F. tularensis LVS. Importantly, the decrease of IL-10 in the mAb-iFt immunized mice observed at 96 hours post-infection was consistent with our previous observation, indicating that FcγR targeting shifts towards a pro-inflammatory cytokine profile at the early stages of F. tularensis infection (Figs 2 and 5).

Bottom Line: It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity.The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date.These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Seton Hall University, South Orange, New Jersey, United States of America.

ABSTRACT
Production of pro-inflammatory cytokines by innate immune cells at the early stages of bacterial infection is important for host protection against the pathogen. Many intracellular bacteria, including Francisella tularensis, the agent of tularemia, utilize the anti-inflammatory cytokine IL-10, to evade the host immune response. It is well established that IL-10 has the ability to inhibit robust antigen presentation by dendritic cells and macrophages, thus suppressing the generation of protective immunity. The pathogenesis of F. tularensis is not fully understood, and research has failed to develop an effective vaccine to this date. In the current study, we hypothesized that F. tularensis polarizes antigen presenting cells during the early stages of infection towards an anti-inflammatory status characterized by increased synthesis of IL-10 and decreased production of IL-12p70 and TNF-α in an IFN-ɣ-dependent fashion. In addition, F. tularensis drives an alternative activation of alveolar macrophages within the first 48 hours post-infection, thus allowing the bacterium to avoid protective immunity. Furthermore, we demonstrate that targeting inactivated F. tularensis (iFt) to Fcγ receptors (FcɣRs) via intranasal immunization with mAb-iFt complexes, a proven vaccine strategy in our laboratories, reverses the anti-inflammatory effects of the bacterium on macrophages by down-regulating production of IL-10. More specifically, we observed that targeting of iFt to FcγRs enhances the classical activation of macrophages not only within the respiratory mucosa, but also systemically, at the early stages of infection. These results provide important insight for further understanding the protective immune mechanisms generated when targeting immunogens to Fc receptors.

No MeSH data available.


Related in: MedlinePlus