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Extracellular vesicles are key intercellular mediators in the development of immune dysfunction to allergens in the airways.

Shin TS, Kim JH, Kim YS, Jeon SG, Zhu Z, Gho YS, Kim YK - Allergy (2010)

Bottom Line: The inhalation of LPS enhanced EVs release into the BAL fluid, when compared to the application of PBS.Airway sensitization with allergens and LPS-induced EVs resulted in a mixed Th1 and Th17 cell responses, although that with allergens and PBS-induced EVs induced immune tolerance.Moreover, the immune responses induced by the LPS-induced EVs were blocked by denaturation of the EV-bearing proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, POSTECH Biotech Center, Pohang University of Science and Technology (POSTECH), Pohang, Korea. juinea@postech.ac.kr

ABSTRACT

Background: Previous evidence indicates that inhalation of lipopolysaccharide (LPS)-containing with allergens induced mixed Th1 and Th17 cell responses in the airways. Extracellular vesicles (EVs) are nanometer-sized spherical, lipid-bilayered structures and are recently in the public eye as an intercellular communicator in immune responses.

Objective: To evaluate the role of EVs secreted by LPS inhalation in the development of airway immune dysfunction in response to allergens.

Methods: Extracellular vesicles in bronchoalveolar lavage fluids of BALB/c mice were isolated and characterized 24 h after applications to the airway of 10 μg of LPS for 3 days. To evaluate the role of LPS-induced EVs on the development of airway immune dysfunction, in vivo and in vitro experiments were performed using the isolated LPS-induced EVs.

Results: The inhalation of LPS enhanced EVs release into the BAL fluid, when compared to the application of PBS. Airway sensitization with allergens and LPS-induced EVs resulted in a mixed Th1 and Th17 cell responses, although that with allergens and PBS-induced EVs induced immune tolerance. In addition, LPS-induced EVs enhanced the production of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) by lung dendritic cells. Moreover, the immune responses induced by the LPS-induced EVs were blocked by denaturation of the EV-bearing proteins.

Conclusion: These data suggest that EVs (especially, the protein components) secreted by LPS inhalation are a key intercellular communicator in the development of airway immune dysfunction to inhaled LPS-containing allergens.

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The production levels of pro-inflammatory and immunomodulating cytokines are up-regulated by airway application of LPS-induced EVs. (A) Production levels of TNF-α and IL-6 in the RAW264.7 macrophage cell line after stimulation with PBS, LPS (10 ng/ml), PBS-induced EVs (0.5 μg/ml) or LPS-induced EVs (0.5 μg/ml). (B) Protocol for in vivo experiments to evaluate innate immune responses (n = 5 each group). (C) BAL cellularity 24 h after airway application of PBS-induced or LPS-induced EVs. (D) In vivo production levels of TNF-α and IL-6 at 6 h after airway application of PBS-induced or LPS-induced EVs. (E) Ex vivo production of IL-12p70 and IL-6 from lung dendritic cells (DCs) 6 h after stimulation with LPS-induced EVs (0.5 μg/ml). In (C–E) *P < 0.05 compared to PBS group.
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fig03: The production levels of pro-inflammatory and immunomodulating cytokines are up-regulated by airway application of LPS-induced EVs. (A) Production levels of TNF-α and IL-6 in the RAW264.7 macrophage cell line after stimulation with PBS, LPS (10 ng/ml), PBS-induced EVs (0.5 μg/ml) or LPS-induced EVs (0.5 μg/ml). (B) Protocol for in vivo experiments to evaluate innate immune responses (n = 5 each group). (C) BAL cellularity 24 h after airway application of PBS-induced or LPS-induced EVs. (D) In vivo production levels of TNF-α and IL-6 at 6 h after airway application of PBS-induced or LPS-induced EVs. (E) Ex vivo production of IL-12p70 and IL-6 from lung dendritic cells (DCs) 6 h after stimulation with LPS-induced EVs (0.5 μg/ml). In (C–E) *P < 0.05 compared to PBS group.

Mentions: The in vitro production levels of TNF-α and IL-6 from mouse macrophages (RAW264.7 cells) were enhanced by stimulation with LPS (10 ng/ml) or LPS-induced EVs (0.5 μg/ml), when compared to stimulation with PBS or PBS-induced EVs (0.5 μg/ml) (Fig. 3A). Therefore, we evaluated the effects of LPS-induced EVs on the development of innate immune responses in vivo. To test this objective, 1 μg of LPS-induced EVs was applied intranasally to naïve BALB/c mice (Fig. 3B). Bronchoalveolar lavage cellularity 24 h postapplication was significantly increased in the mice that received LPS-induced EVs, when compared to those that received PBS-induced EVs (Fig. 3C). Moreover, the in vivo production levels of TNF-α and IL-6 at 6 h postapplication were higher in the mice that received LPS-induced EVs than in those that received PBS-induced EVs (Fig. 3D). Based on the previous data that airway application of LPS-containing allergens induced mixed Th1 and Th17 cell responses via the up-regulation of IL-12p70 and IL-6 production, respectively (7, 8), we evaluated the effects of LPS-induced EVs on the production of T-cell polarizing cytokines from antigen-presenting cells. CD11c+ cells (DCs) were isolated from naïve BALB/c mice, and then stimulated with LPS-induced EVs or PBS in vitro. The production levels of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) were significantly higher following stimulation with LPS-induced EVs (Fig. 3E). Taken together, these data indicate that EVs secreted in response to LPS cause infiltration of inflammatory cells via the up-regulation of pro-inflammatory mediators, such as TNF-α, and modulate adaptive immune responses via the up-regulation of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) in antigen-presenting cells.


Extracellular vesicles are key intercellular mediators in the development of immune dysfunction to allergens in the airways.

Shin TS, Kim JH, Kim YS, Jeon SG, Zhu Z, Gho YS, Kim YK - Allergy (2010)

The production levels of pro-inflammatory and immunomodulating cytokines are up-regulated by airway application of LPS-induced EVs. (A) Production levels of TNF-α and IL-6 in the RAW264.7 macrophage cell line after stimulation with PBS, LPS (10 ng/ml), PBS-induced EVs (0.5 μg/ml) or LPS-induced EVs (0.5 μg/ml). (B) Protocol for in vivo experiments to evaluate innate immune responses (n = 5 each group). (C) BAL cellularity 24 h after airway application of PBS-induced or LPS-induced EVs. (D) In vivo production levels of TNF-α and IL-6 at 6 h after airway application of PBS-induced or LPS-induced EVs. (E) Ex vivo production of IL-12p70 and IL-6 from lung dendritic cells (DCs) 6 h after stimulation with LPS-induced EVs (0.5 μg/ml). In (C–E) *P < 0.05 compared to PBS group.
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fig03: The production levels of pro-inflammatory and immunomodulating cytokines are up-regulated by airway application of LPS-induced EVs. (A) Production levels of TNF-α and IL-6 in the RAW264.7 macrophage cell line after stimulation with PBS, LPS (10 ng/ml), PBS-induced EVs (0.5 μg/ml) or LPS-induced EVs (0.5 μg/ml). (B) Protocol for in vivo experiments to evaluate innate immune responses (n = 5 each group). (C) BAL cellularity 24 h after airway application of PBS-induced or LPS-induced EVs. (D) In vivo production levels of TNF-α and IL-6 at 6 h after airway application of PBS-induced or LPS-induced EVs. (E) Ex vivo production of IL-12p70 and IL-6 from lung dendritic cells (DCs) 6 h after stimulation with LPS-induced EVs (0.5 μg/ml). In (C–E) *P < 0.05 compared to PBS group.
Mentions: The in vitro production levels of TNF-α and IL-6 from mouse macrophages (RAW264.7 cells) were enhanced by stimulation with LPS (10 ng/ml) or LPS-induced EVs (0.5 μg/ml), when compared to stimulation with PBS or PBS-induced EVs (0.5 μg/ml) (Fig. 3A). Therefore, we evaluated the effects of LPS-induced EVs on the development of innate immune responses in vivo. To test this objective, 1 μg of LPS-induced EVs was applied intranasally to naïve BALB/c mice (Fig. 3B). Bronchoalveolar lavage cellularity 24 h postapplication was significantly increased in the mice that received LPS-induced EVs, when compared to those that received PBS-induced EVs (Fig. 3C). Moreover, the in vivo production levels of TNF-α and IL-6 at 6 h postapplication were higher in the mice that received LPS-induced EVs than in those that received PBS-induced EVs (Fig. 3D). Based on the previous data that airway application of LPS-containing allergens induced mixed Th1 and Th17 cell responses via the up-regulation of IL-12p70 and IL-6 production, respectively (7, 8), we evaluated the effects of LPS-induced EVs on the production of T-cell polarizing cytokines from antigen-presenting cells. CD11c+ cells (DCs) were isolated from naïve BALB/c mice, and then stimulated with LPS-induced EVs or PBS in vitro. The production levels of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) were significantly higher following stimulation with LPS-induced EVs (Fig. 3E). Taken together, these data indicate that EVs secreted in response to LPS cause infiltration of inflammatory cells via the up-regulation of pro-inflammatory mediators, such as TNF-α, and modulate adaptive immune responses via the up-regulation of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) in antigen-presenting cells.

Bottom Line: The inhalation of LPS enhanced EVs release into the BAL fluid, when compared to the application of PBS.Airway sensitization with allergens and LPS-induced EVs resulted in a mixed Th1 and Th17 cell responses, although that with allergens and PBS-induced EVs induced immune tolerance.Moreover, the immune responses induced by the LPS-induced EVs were blocked by denaturation of the EV-bearing proteins.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, POSTECH Biotech Center, Pohang University of Science and Technology (POSTECH), Pohang, Korea. juinea@postech.ac.kr

ABSTRACT

Background: Previous evidence indicates that inhalation of lipopolysaccharide (LPS)-containing with allergens induced mixed Th1 and Th17 cell responses in the airways. Extracellular vesicles (EVs) are nanometer-sized spherical, lipid-bilayered structures and are recently in the public eye as an intercellular communicator in immune responses.

Objective: To evaluate the role of EVs secreted by LPS inhalation in the development of airway immune dysfunction in response to allergens.

Methods: Extracellular vesicles in bronchoalveolar lavage fluids of BALB/c mice were isolated and characterized 24 h after applications to the airway of 10 μg of LPS for 3 days. To evaluate the role of LPS-induced EVs on the development of airway immune dysfunction, in vivo and in vitro experiments were performed using the isolated LPS-induced EVs.

Results: The inhalation of LPS enhanced EVs release into the BAL fluid, when compared to the application of PBS. Airway sensitization with allergens and LPS-induced EVs resulted in a mixed Th1 and Th17 cell responses, although that with allergens and PBS-induced EVs induced immune tolerance. In addition, LPS-induced EVs enhanced the production of Th1- and Th17-polarizing cytokines (IL-12p70 and IL-6, respectively) by lung dendritic cells. Moreover, the immune responses induced by the LPS-induced EVs were blocked by denaturation of the EV-bearing proteins.

Conclusion: These data suggest that EVs (especially, the protein components) secreted by LPS inhalation are a key intercellular communicator in the development of airway immune dysfunction to inhaled LPS-containing allergens.

Show MeSH
Related in: MedlinePlus