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Dendritic cells induce Th2-mediated airway inflammatory responses to house dust mite via DNA-dependent protein kinase.

Mishra A, Brown AL, Yao X, Yang S, Park SJ, Liu C, Dagur PK, McCoy JP, Keeran KJ, Nugent GZ, Jeffries KR, Qu X, Yu ZX, Levine SJ, Chung JH - Nat Commun (2015)

Bottom Line: Here we show that house dust mite induces DNA-PK phosphorylation, which is a marker of DNA-PK activation, in DCs via the generation of intracellular reactive oxygen species.We also demonstrate that pharmacological inhibition of DNA-PK, as well as the specific deletion of DNA-PK in DCs, attenuates the induction of allergic sensitization and Th2 immunity via a mechanism that involves the impaired presentation of mite antigens.Furthermore, pharmacological inhibition of DNA-PK following antigen priming similarly reduces the manifestations of mite-induced airway disease.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Asthma and Lung Inflammation, Division of Intramural Research, NHLBI, NIH, Bethesda, Maryland 20892, USA.

ABSTRACT
DNA-dependent protein kinase (DNA-PK) mediates double-stranded DNA break repair, V(D)J recombination and immunoglobulin class switch recombination, as well as innate immune and pro-inflammatory responses. However, there is limited information regarding the role of DNA-PK in adaptive immunity mediated by dendritic cells (DCs), which are the primary antigen-presenting cells in allergic asthma. Here we show that house dust mite induces DNA-PK phosphorylation, which is a marker of DNA-PK activation, in DCs via the generation of intracellular reactive oxygen species. We also demonstrate that pharmacological inhibition of DNA-PK, as well as the specific deletion of DNA-PK in DCs, attenuates the induction of allergic sensitization and Th2 immunity via a mechanism that involves the impaired presentation of mite antigens. Furthermore, pharmacological inhibition of DNA-PK following antigen priming similarly reduces the manifestations of mite-induced airway disease. Collectively, these findings suggest that DNA-PK may be a potential target for treatment of allergic asthma.

No MeSH data available.


Related in: MedlinePlus

The adoptive transfer of CD11b+ DCs fromCD11c-specificDNA-PKcsKnockout Mice have an Impaired Ability to InduceHDM-mediated Airway InflammationA. DNA-PKcsfl/fl andDNA-PKcsfl/fl; CD11c-Cre donor mice received a singleintranasal dose of 100 µg of HDM extract or saline and after 4 days, mediastinallymph nodes were removed andCD11c+/CD11b+/SiglecF−/MHCII+ DCswere isolated by flow cytometry. 2.5 × 104CD11c+/CD11b+/SiglecF−/MHCII+ DCswere adoptively transferred to wild type C57BL6 recipient mice by intranasaladministration on day 4 and daily intranasal HDM challenges (25 µg) wereadministered on days 13 through 18 to all recipient mice. Mice were sacrificed forend-point analysis on day 19. B. The number of total BALF inflammatory cellsand inflammatory cell subtypes in recipients of adoptively transferred CD11b+DCs (n = 6 – 10, * P < 0.05, one way ANOVA with Bonferroni multiplecomparison test). C. Representative histologic lung sections stained withhematoxylin and eosin (H&E) and periodic acid-Schiff (PAS). Scale bars denote 100µm for the x200 images and 20 µm for the x1000 images. D.Quantification of mucous cell metaplasia. (n = 10, * P = 0.0005, unpairedt test). 48.7 + 3.3 airways were analyzed per mouse. E.Serum HDM-specific IgE (n = 6 – 10 mice, * P < 0.05, one way ANOVA withBonferroni multiple comparison test). F. Cytokine secretion by exvivo cultures of mediastinal lymph node cells that had been re-stimulated withHDM (100 µg/ml) (n = 6 – 8 mice, * P < 0.01, one way ANOVA withBonferroni multiple comparison test). Results are pooled data from two independentexperiments.
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Figure 7: The adoptive transfer of CD11b+ DCs fromCD11c-specificDNA-PKcsKnockout Mice have an Impaired Ability to InduceHDM-mediated Airway InflammationA. DNA-PKcsfl/fl andDNA-PKcsfl/fl; CD11c-Cre donor mice received a singleintranasal dose of 100 µg of HDM extract or saline and after 4 days, mediastinallymph nodes were removed andCD11c+/CD11b+/SiglecF−/MHCII+ DCswere isolated by flow cytometry. 2.5 × 104CD11c+/CD11b+/SiglecF−/MHCII+ DCswere adoptively transferred to wild type C57BL6 recipient mice by intranasaladministration on day 4 and daily intranasal HDM challenges (25 µg) wereadministered on days 13 through 18 to all recipient mice. Mice were sacrificed forend-point analysis on day 19. B. The number of total BALF inflammatory cellsand inflammatory cell subtypes in recipients of adoptively transferred CD11b+DCs (n = 6 – 10, * P < 0.05, one way ANOVA with Bonferroni multiplecomparison test). C. Representative histologic lung sections stained withhematoxylin and eosin (H&E) and periodic acid-Schiff (PAS). Scale bars denote 100µm for the x200 images and 20 µm for the x1000 images. D.Quantification of mucous cell metaplasia. (n = 10, * P = 0.0005, unpairedt test). 48.7 + 3.3 airways were analyzed per mouse. E.Serum HDM-specific IgE (n = 6 – 10 mice, * P < 0.05, one way ANOVA withBonferroni multiple comparison test). F. Cytokine secretion by exvivo cultures of mediastinal lymph node cells that had been re-stimulated withHDM (100 µg/ml) (n = 6 – 8 mice, * P < 0.01, one way ANOVA withBonferroni multiple comparison test). Results are pooled data from two independentexperiments.

Mentions: Since uptake of HDM antigen in the lung and trafficking to MLNs byCD11b+ DCs from DNA-PKcsfl/fl; Cd11c-Cre micewere not reduced, we next assessed whether CD11b+ DCs fromDNA-PKcsfl/fl; Cd11c-Cre mice have an impairment inantigen presentation that limits their ability to induce allergic sensitization andTh2-mediated airway inflammation. CD11b+ DCs were isolated from MLNs ofDNA-PKcsfl/fl and DNA-PKcsfl/fl;Cd11c-Cre mice that had been challenged with HDM or saline and adoptivelytransferred to WT mice that subsequently received intranasal HDM challenges to induceairway inflammation (Figure 7A). As shown in Figure 7B, the number alveolar macrophages, eosinophils,neutrophils and lymphocytes were significantly reduced in BALF from recipients ofCD11b+ DCs from DNA-PKcsfl/fl; Cd11c-Cre donormice as compared to DNA-PKcsfl/fl donor mice. Similarly, lunghistology showed a reduction in peri-bronchial inflammatory cell infiltrates and mucouscell metaplasia in recipients of CD11b+ DCs fromDNA-PKcsfl/fl; Cd11c-Cre donor mice as compared toDNA-PKcsfl/fl donor mice (Figures 7C and 7D). Serum levels of HDM-specific IgE were significantly reducedin recipients of CD11b+ DCs from DNA-PKcsfl/fl;Cd11c-Cre donor mice, while ex vivo cultures of MLNs that hadbeen re-stimulated with HDM showed an impaired ability to produce IL-5 and IL-13 (Figures 7E and 7F). Collectively, these experimentsdemonstrate that CD11b+ DCs from DNA-PKcsfl/fl;Cd11c-Cre mice have an impairment in antigen presentation and the subsequentinduction of allergic sensitization and Th2 immunity to HDM.


Dendritic cells induce Th2-mediated airway inflammatory responses to house dust mite via DNA-dependent protein kinase.

Mishra A, Brown AL, Yao X, Yang S, Park SJ, Liu C, Dagur PK, McCoy JP, Keeran KJ, Nugent GZ, Jeffries KR, Qu X, Yu ZX, Levine SJ, Chung JH - Nat Commun (2015)

The adoptive transfer of CD11b+ DCs fromCD11c-specificDNA-PKcsKnockout Mice have an Impaired Ability to InduceHDM-mediated Airway InflammationA. DNA-PKcsfl/fl andDNA-PKcsfl/fl; CD11c-Cre donor mice received a singleintranasal dose of 100 µg of HDM extract or saline and after 4 days, mediastinallymph nodes were removed andCD11c+/CD11b+/SiglecF−/MHCII+ DCswere isolated by flow cytometry. 2.5 × 104CD11c+/CD11b+/SiglecF−/MHCII+ DCswere adoptively transferred to wild type C57BL6 recipient mice by intranasaladministration on day 4 and daily intranasal HDM challenges (25 µg) wereadministered on days 13 through 18 to all recipient mice. Mice were sacrificed forend-point analysis on day 19. B. The number of total BALF inflammatory cellsand inflammatory cell subtypes in recipients of adoptively transferred CD11b+DCs (n = 6 – 10, * P < 0.05, one way ANOVA with Bonferroni multiplecomparison test). C. Representative histologic lung sections stained withhematoxylin and eosin (H&E) and periodic acid-Schiff (PAS). Scale bars denote 100µm for the x200 images and 20 µm for the x1000 images. D.Quantification of mucous cell metaplasia. (n = 10, * P = 0.0005, unpairedt test). 48.7 + 3.3 airways were analyzed per mouse. E.Serum HDM-specific IgE (n = 6 – 10 mice, * P < 0.05, one way ANOVA withBonferroni multiple comparison test). F. Cytokine secretion by exvivo cultures of mediastinal lymph node cells that had been re-stimulated withHDM (100 µg/ml) (n = 6 – 8 mice, * P < 0.01, one way ANOVA withBonferroni multiple comparison test). Results are pooled data from two independentexperiments.
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Related In: Results  -  Collection

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Figure 7: The adoptive transfer of CD11b+ DCs fromCD11c-specificDNA-PKcsKnockout Mice have an Impaired Ability to InduceHDM-mediated Airway InflammationA. DNA-PKcsfl/fl andDNA-PKcsfl/fl; CD11c-Cre donor mice received a singleintranasal dose of 100 µg of HDM extract or saline and after 4 days, mediastinallymph nodes were removed andCD11c+/CD11b+/SiglecF−/MHCII+ DCswere isolated by flow cytometry. 2.5 × 104CD11c+/CD11b+/SiglecF−/MHCII+ DCswere adoptively transferred to wild type C57BL6 recipient mice by intranasaladministration on day 4 and daily intranasal HDM challenges (25 µg) wereadministered on days 13 through 18 to all recipient mice. Mice were sacrificed forend-point analysis on day 19. B. The number of total BALF inflammatory cellsand inflammatory cell subtypes in recipients of adoptively transferred CD11b+DCs (n = 6 – 10, * P < 0.05, one way ANOVA with Bonferroni multiplecomparison test). C. Representative histologic lung sections stained withhematoxylin and eosin (H&E) and periodic acid-Schiff (PAS). Scale bars denote 100µm for the x200 images and 20 µm for the x1000 images. D.Quantification of mucous cell metaplasia. (n = 10, * P = 0.0005, unpairedt test). 48.7 + 3.3 airways were analyzed per mouse. E.Serum HDM-specific IgE (n = 6 – 10 mice, * P < 0.05, one way ANOVA withBonferroni multiple comparison test). F. Cytokine secretion by exvivo cultures of mediastinal lymph node cells that had been re-stimulated withHDM (100 µg/ml) (n = 6 – 8 mice, * P < 0.01, one way ANOVA withBonferroni multiple comparison test). Results are pooled data from two independentexperiments.
Mentions: Since uptake of HDM antigen in the lung and trafficking to MLNs byCD11b+ DCs from DNA-PKcsfl/fl; Cd11c-Cre micewere not reduced, we next assessed whether CD11b+ DCs fromDNA-PKcsfl/fl; Cd11c-Cre mice have an impairment inantigen presentation that limits their ability to induce allergic sensitization andTh2-mediated airway inflammation. CD11b+ DCs were isolated from MLNs ofDNA-PKcsfl/fl and DNA-PKcsfl/fl;Cd11c-Cre mice that had been challenged with HDM or saline and adoptivelytransferred to WT mice that subsequently received intranasal HDM challenges to induceairway inflammation (Figure 7A). As shown in Figure 7B, the number alveolar macrophages, eosinophils,neutrophils and lymphocytes were significantly reduced in BALF from recipients ofCD11b+ DCs from DNA-PKcsfl/fl; Cd11c-Cre donormice as compared to DNA-PKcsfl/fl donor mice. Similarly, lunghistology showed a reduction in peri-bronchial inflammatory cell infiltrates and mucouscell metaplasia in recipients of CD11b+ DCs fromDNA-PKcsfl/fl; Cd11c-Cre donor mice as compared toDNA-PKcsfl/fl donor mice (Figures 7C and 7D). Serum levels of HDM-specific IgE were significantly reducedin recipients of CD11b+ DCs from DNA-PKcsfl/fl;Cd11c-Cre donor mice, while ex vivo cultures of MLNs that hadbeen re-stimulated with HDM showed an impaired ability to produce IL-5 and IL-13 (Figures 7E and 7F). Collectively, these experimentsdemonstrate that CD11b+ DCs from DNA-PKcsfl/fl;Cd11c-Cre mice have an impairment in antigen presentation and the subsequentinduction of allergic sensitization and Th2 immunity to HDM.

Bottom Line: Here we show that house dust mite induces DNA-PK phosphorylation, which is a marker of DNA-PK activation, in DCs via the generation of intracellular reactive oxygen species.We also demonstrate that pharmacological inhibition of DNA-PK, as well as the specific deletion of DNA-PK in DCs, attenuates the induction of allergic sensitization and Th2 immunity via a mechanism that involves the impaired presentation of mite antigens.Furthermore, pharmacological inhibition of DNA-PK following antigen priming similarly reduces the manifestations of mite-induced airway disease.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Asthma and Lung Inflammation, Division of Intramural Research, NHLBI, NIH, Bethesda, Maryland 20892, USA.

ABSTRACT
DNA-dependent protein kinase (DNA-PK) mediates double-stranded DNA break repair, V(D)J recombination and immunoglobulin class switch recombination, as well as innate immune and pro-inflammatory responses. However, there is limited information regarding the role of DNA-PK in adaptive immunity mediated by dendritic cells (DCs), which are the primary antigen-presenting cells in allergic asthma. Here we show that house dust mite induces DNA-PK phosphorylation, which is a marker of DNA-PK activation, in DCs via the generation of intracellular reactive oxygen species. We also demonstrate that pharmacological inhibition of DNA-PK, as well as the specific deletion of DNA-PK in DCs, attenuates the induction of allergic sensitization and Th2 immunity via a mechanism that involves the impaired presentation of mite antigens. Furthermore, pharmacological inhibition of DNA-PK following antigen priming similarly reduces the manifestations of mite-induced airway disease. Collectively, these findings suggest that DNA-PK may be a potential target for treatment of allergic asthma.

No MeSH data available.


Related in: MedlinePlus