Limits...
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.

Show MeSH

Related in: MedlinePlus

Protein components of the EVs secreted in response to LPS play key roles in the mediation of innate immune responses. (A) The effects of pretreatment with polymyxin B (PMB) (an antagonist of LPS) on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. (B) The effects of pretreatment with protease K or heating on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. For (A and B) *P < 0.05 compared to the EV_PBS group; **P < 0.05 compared to the other groups. (C) Confocal and differential interference contrast (DIC) microscopy imaging of the effects of pretreatment with protease K or heating on the endocytosis of LPS-induced EVs into the cytoplasm of RAW264.7 macrophages. DAPI and cell tracker denote nucleus and cytoplasmic staining, respectively; DiI indicates lipid labeling. In both confocal merged and DIC images show that endocytosis does not occur in LPS-induced EV after pretreatment with protease K or with heat, although it occurred in LPS-induced EVs without these treatments. (a: EV_PBS; b: EV_LPS; c: EV_LPS/protease; d: EV_LPS/heat). (D) Study protocol for in vivo experiments (left panel), and the effects of pretreatment of proteinase K or heating on the in vivo production of TNF-α and IL-6 in the airways (right panel). *P < 0.05 compared to the other groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3066408&req=5

fig04: Protein components of the EVs secreted in response to LPS play key roles in the mediation of innate immune responses. (A) The effects of pretreatment with polymyxin B (PMB) (an antagonist of LPS) on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. (B) The effects of pretreatment with protease K or heating on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. For (A and B) *P < 0.05 compared to the EV_PBS group; **P < 0.05 compared to the other groups. (C) Confocal and differential interference contrast (DIC) microscopy imaging of the effects of pretreatment with protease K or heating on the endocytosis of LPS-induced EVs into the cytoplasm of RAW264.7 macrophages. DAPI and cell tracker denote nucleus and cytoplasmic staining, respectively; DiI indicates lipid labeling. In both confocal merged and DIC images show that endocytosis does not occur in LPS-induced EV after pretreatment with protease K or with heat, although it occurred in LPS-induced EVs without these treatments. (a: EV_PBS; b: EV_LPS; c: EV_LPS/protease; d: EV_LPS/heat). (D) Study protocol for in vivo experiments (left panel), and the effects of pretreatment of proteinase K or heating on the in vivo production of TNF-α and IL-6 in the airways (right panel). *P < 0.05 compared to the other groups.

Mentions: To evaluate the role of the LPS in the LPS-induced EVs in the innate immune responses, the in vitro production levels of TNF-α and IL-6 in lung macrophages were evaluated after incubation with LPS-induced EVs, with or without pretreatment with PMB (an antagonist of LPS). The production levels of TNF-α and IL-6 were similar after stimulation with LPS-induced EVs, with or without pretreatment with PMB, although the LPS-enhanced production of these cytokines was abolished by pretreatment with PMB (Fig. 4A). Taken the absence of any LPS in the LPS-induced EVs into the consideration, this finding indicates that LPS itself in host cell-derived EVs plays no role in the development of the innate immune responses induced by airway LPS exposure.


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)

Protein components of the EVs secreted in response to LPS play key roles in the mediation of innate immune responses. (A) The effects of pretreatment with polymyxin B (PMB) (an antagonist of LPS) on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. (B) The effects of pretreatment with protease K or heating on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. For (A and B) *P < 0.05 compared to the EV_PBS group; **P < 0.05 compared to the other groups. (C) Confocal and differential interference contrast (DIC) microscopy imaging of the effects of pretreatment with protease K or heating on the endocytosis of LPS-induced EVs into the cytoplasm of RAW264.7 macrophages. DAPI and cell tracker denote nucleus and cytoplasmic staining, respectively; DiI indicates lipid labeling. In both confocal merged and DIC images show that endocytosis does not occur in LPS-induced EV after pretreatment with protease K or with heat, although it occurred in LPS-induced EVs without these treatments. (a: EV_PBS; b: EV_LPS; c: EV_LPS/protease; d: EV_LPS/heat). (D) Study protocol for in vivo experiments (left panel), and the effects of pretreatment of proteinase K or heating on the in vivo production of TNF-α and IL-6 in the airways (right panel). *P < 0.05 compared to the other groups.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3066408&req=5

fig04: Protein components of the EVs secreted in response to LPS play key roles in the mediation of innate immune responses. (A) The effects of pretreatment with polymyxin B (PMB) (an antagonist of LPS) on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. (B) The effects of pretreatment with protease K or heating on the in vitro production levels of TNF-α and IL-6 in RAW264.7 macrophages. For (A and B) *P < 0.05 compared to the EV_PBS group; **P < 0.05 compared to the other groups. (C) Confocal and differential interference contrast (DIC) microscopy imaging of the effects of pretreatment with protease K or heating on the endocytosis of LPS-induced EVs into the cytoplasm of RAW264.7 macrophages. DAPI and cell tracker denote nucleus and cytoplasmic staining, respectively; DiI indicates lipid labeling. In both confocal merged and DIC images show that endocytosis does not occur in LPS-induced EV after pretreatment with protease K or with heat, although it occurred in LPS-induced EVs without these treatments. (a: EV_PBS; b: EV_LPS; c: EV_LPS/protease; d: EV_LPS/heat). (D) Study protocol for in vivo experiments (left panel), and the effects of pretreatment of proteinase K or heating on the in vivo production of TNF-α and IL-6 in the airways (right panel). *P < 0.05 compared to the other groups.
Mentions: To evaluate the role of the LPS in the LPS-induced EVs in the innate immune responses, the in vitro production levels of TNF-α and IL-6 in lung macrophages were evaluated after incubation with LPS-induced EVs, with or without pretreatment with PMB (an antagonist of LPS). The production levels of TNF-α and IL-6 were similar after stimulation with LPS-induced EVs, with or without pretreatment with PMB, although the LPS-enhanced production of these cytokines was abolished by pretreatment with PMB (Fig. 4A). Taken the absence of any LPS in the LPS-induced EVs into the consideration, this finding indicates that LPS itself in host cell-derived EVs plays no role in the development of the innate immune responses induced by airway LPS exposure.

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