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Blockage of Eosinopoiesis by IL-17A Is Prevented by Cytokine and Lipid Mediators of Allergic Inflammation.

Xavier-Elsas P, de Luca B, Queto T, Vieira BM, Masid-de-Brito D, Dahab EC, Alves Filho JC, Cunha FQ, Gaspar-Elsas MI - Mediators Inflamm. (2015)

Bottom Line: While IL-17A (0.1-10 ng/mL) had no IL-5-independent effect on eosinopoiesis, it dose-dependently suppressed IL-5-induced eosinophil differentiation, by acting during the initial 24 hours.Its effectiveness was abolished by caspase inhibitor, zVAD-fmk.By contrast, a higher IL-17A concentration (10 ng/mL) retained significant suppressive effect in both conditions, unmasking a high-end iNOS-independent mechanism.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Paulo de Góes Institute for Microbiology, Federal University of Rio de Janeiro (UFRJ), 21941-590 Rio de Janeiro, RJ, Brazil.

ABSTRACT
Interleukin- (IL-) 17A, a pleiotropic mediator of inflammation and autoimmunity, potently stimulates bone-marrow neutrophil production. To explore IL-17A effects on eosinopoiesis, we cultured bone-marrow from wild-type mice, or mutants lacking inducible nitric oxide synthase (iNOS-/-), CD95 (lpr), IL-17RA, or IL-4, with IL-5, alone or associated with IL-17A. Synergisms between IL-17A-activated, NO-dependent, and NO-independent mechanisms and antagonisms between IL-17A and proallergic factors were further examined. While IL-17A (0.1-10 ng/mL) had no IL-5-independent effect on eosinopoiesis, it dose-dependently suppressed IL-5-induced eosinophil differentiation, by acting during the initial 24 hours. Its effectiveness was abolished by caspase inhibitor, zVAD-fmk. The effect of IL-17A (0.1-1 ng/mL) was sensitive to the iNOS-selective inhibitor aminoguanidine and undetectable in iNOS-/- bone-marrow. By contrast, a higher IL-17A concentration (10 ng/mL) retained significant suppressive effect in both conditions, unmasking a high-end iNOS-independent mechanism. Lower IL-17A concentrations synergized with NO donor nitroprusside. Eosinopoiesis suppression by IL-17A was (a) undetectable in bone-marrow lacking IL-17RA or CD95 and (b) actively prevented by LTD4, LTC4, IL-13, and eotaxin. Sensitivity to IL-17A was increased in bone-marrow lacking IL-4; adding IL-4 to the cultures restored IL-5 responses to control levels. Therefore, effects of both IL-17A and proallergic factors are transduced by the iNOS-CD95 pathway in isolated bone-marrow.

No MeSH data available.


Related in: MedlinePlus

IL-17, effect on progenitor responses and requirement for IL-17RA. (a, b) Semisolid cultures were established for 7 days, from 2 × 105 bone-marrow cells of naive BALB/c mice, in the presence of GM-CSF (2 ng/mL), alone or in association with IL-17A at the indicated concentrations. Total (a) and differential (b) colony counts were carried out under the inverted microscope, directly (a) or after staining dried agar layers for EPO and counterstaining by Harris' Hematoxylin (b). Colony types in (b) are [36] pure granulocyte-macrophage (GM), mixed granulocyte-macrophage-eosinophil (GMEos), pure eosinophil (Eos), pure granulocyte (G), and pure macrophage (M). GM-CSF alone, black bar. GM-CSF in association with IL-17A (0.1 to 1 ng/mL), white bars. (c) Liquid cultures were established as described by Gaspar-Elsas et al. [38], in a two-phase culture with progenitor expansion in Flt3L and SCF, followed by eosinophil differentiation for up to 10 days with GM-CSF, IL-3, and IL-5, in the absence (white bar) or presence (black bar) of IL-17A (1 ng/mL). (d) Liquid cultures were established as in Figure 1(a), with bone-marrow from BALB/c mutants lacking IL-17RA. Data are mean ± SEM. ∗P < 0.05. Significant differences relative to GM controls (a, b) or IL-5 controls (c, d). All groups, n = 3.
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fig3: IL-17, effect on progenitor responses and requirement for IL-17RA. (a, b) Semisolid cultures were established for 7 days, from 2 × 105 bone-marrow cells of naive BALB/c mice, in the presence of GM-CSF (2 ng/mL), alone or in association with IL-17A at the indicated concentrations. Total (a) and differential (b) colony counts were carried out under the inverted microscope, directly (a) or after staining dried agar layers for EPO and counterstaining by Harris' Hematoxylin (b). Colony types in (b) are [36] pure granulocyte-macrophage (GM), mixed granulocyte-macrophage-eosinophil (GMEos), pure eosinophil (Eos), pure granulocyte (G), and pure macrophage (M). GM-CSF alone, black bar. GM-CSF in association with IL-17A (0.1 to 1 ng/mL), white bars. (c) Liquid cultures were established as described by Gaspar-Elsas et al. [38], in a two-phase culture with progenitor expansion in Flt3L and SCF, followed by eosinophil differentiation for up to 10 days with GM-CSF, IL-3, and IL-5, in the absence (white bar) or presence (black bar) of IL-17A (1 ng/mL). (d) Liquid cultures were established as in Figure 1(a), with bone-marrow from BALB/c mutants lacking IL-17RA. Data are mean ± SEM. ∗P < 0.05. Significant differences relative to GM controls (a, b) or IL-5 controls (c, d). All groups, n = 3.

Mentions: Because PGE2 has a strong inhibitory effect on myeloid colony formation [42], while IL-17 is considered a strong stimulant of granulopoiesis in vivo [21, 22, 24], we examined in detail the effects of IL-17A on myeloid colony-forming cells (progenitors). We evaluated whether IL-17A significantly affected colony formation in the presence of GM-CSF (Figures 3(a) and 3(b)), taking care to distinguish between effects on total colony counts (Figure 3(a)) and on different colony types (Figure 3(b)). In the 0.1–1 ng/mL concentration range, IL-17A had no significant effect on colony counts, total or differential, in comparison with the GM-CSF control. By contrast, IL-17A had a significant enhancing effect on total colony counts at 10 ng/mL (Figure 3(a)). This effect was accounted for by a significant increase in granulocyte-macrophage (GM) colony counts (Figure 3(b)), but not in eosinophil-containing GM colonies (GMEos), which were scored as a separate category [36], nor in pure eosinophil colonies (Eos). All myeloid colony types other than pure GM colonies, including both classes of eosinophil-containing colonies, were unaffected by IL-A17 over the 0.1–10 ng/mL concentration range. Therefore, no suppression of eosinophil progenitors was observed in semisolid culture, suggesting that the inhibition of eosinopoiesis by IL-17A is due to an effect on lineage-committed precursors, which are but no longer able to form colonies.


Blockage of Eosinopoiesis by IL-17A Is Prevented by Cytokine and Lipid Mediators of Allergic Inflammation.

Xavier-Elsas P, de Luca B, Queto T, Vieira BM, Masid-de-Brito D, Dahab EC, Alves Filho JC, Cunha FQ, Gaspar-Elsas MI - Mediators Inflamm. (2015)

IL-17, effect on progenitor responses and requirement for IL-17RA. (a, b) Semisolid cultures were established for 7 days, from 2 × 105 bone-marrow cells of naive BALB/c mice, in the presence of GM-CSF (2 ng/mL), alone or in association with IL-17A at the indicated concentrations. Total (a) and differential (b) colony counts were carried out under the inverted microscope, directly (a) or after staining dried agar layers for EPO and counterstaining by Harris' Hematoxylin (b). Colony types in (b) are [36] pure granulocyte-macrophage (GM), mixed granulocyte-macrophage-eosinophil (GMEos), pure eosinophil (Eos), pure granulocyte (G), and pure macrophage (M). GM-CSF alone, black bar. GM-CSF in association with IL-17A (0.1 to 1 ng/mL), white bars. (c) Liquid cultures were established as described by Gaspar-Elsas et al. [38], in a two-phase culture with progenitor expansion in Flt3L and SCF, followed by eosinophil differentiation for up to 10 days with GM-CSF, IL-3, and IL-5, in the absence (white bar) or presence (black bar) of IL-17A (1 ng/mL). (d) Liquid cultures were established as in Figure 1(a), with bone-marrow from BALB/c mutants lacking IL-17RA. Data are mean ± SEM. ∗P < 0.05. Significant differences relative to GM controls (a, b) or IL-5 controls (c, d). All groups, n = 3.
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig3: IL-17, effect on progenitor responses and requirement for IL-17RA. (a, b) Semisolid cultures were established for 7 days, from 2 × 105 bone-marrow cells of naive BALB/c mice, in the presence of GM-CSF (2 ng/mL), alone or in association with IL-17A at the indicated concentrations. Total (a) and differential (b) colony counts were carried out under the inverted microscope, directly (a) or after staining dried agar layers for EPO and counterstaining by Harris' Hematoxylin (b). Colony types in (b) are [36] pure granulocyte-macrophage (GM), mixed granulocyte-macrophage-eosinophil (GMEos), pure eosinophil (Eos), pure granulocyte (G), and pure macrophage (M). GM-CSF alone, black bar. GM-CSF in association with IL-17A (0.1 to 1 ng/mL), white bars. (c) Liquid cultures were established as described by Gaspar-Elsas et al. [38], in a two-phase culture with progenitor expansion in Flt3L and SCF, followed by eosinophil differentiation for up to 10 days with GM-CSF, IL-3, and IL-5, in the absence (white bar) or presence (black bar) of IL-17A (1 ng/mL). (d) Liquid cultures were established as in Figure 1(a), with bone-marrow from BALB/c mutants lacking IL-17RA. Data are mean ± SEM. ∗P < 0.05. Significant differences relative to GM controls (a, b) or IL-5 controls (c, d). All groups, n = 3.
Mentions: Because PGE2 has a strong inhibitory effect on myeloid colony formation [42], while IL-17 is considered a strong stimulant of granulopoiesis in vivo [21, 22, 24], we examined in detail the effects of IL-17A on myeloid colony-forming cells (progenitors). We evaluated whether IL-17A significantly affected colony formation in the presence of GM-CSF (Figures 3(a) and 3(b)), taking care to distinguish between effects on total colony counts (Figure 3(a)) and on different colony types (Figure 3(b)). In the 0.1–1 ng/mL concentration range, IL-17A had no significant effect on colony counts, total or differential, in comparison with the GM-CSF control. By contrast, IL-17A had a significant enhancing effect on total colony counts at 10 ng/mL (Figure 3(a)). This effect was accounted for by a significant increase in granulocyte-macrophage (GM) colony counts (Figure 3(b)), but not in eosinophil-containing GM colonies (GMEos), which were scored as a separate category [36], nor in pure eosinophil colonies (Eos). All myeloid colony types other than pure GM colonies, including both classes of eosinophil-containing colonies, were unaffected by IL-A17 over the 0.1–10 ng/mL concentration range. Therefore, no suppression of eosinophil progenitors was observed in semisolid culture, suggesting that the inhibition of eosinopoiesis by IL-17A is due to an effect on lineage-committed precursors, which are but no longer able to form colonies.

Bottom Line: While IL-17A (0.1-10 ng/mL) had no IL-5-independent effect on eosinopoiesis, it dose-dependently suppressed IL-5-induced eosinophil differentiation, by acting during the initial 24 hours.Its effectiveness was abolished by caspase inhibitor, zVAD-fmk.By contrast, a higher IL-17A concentration (10 ng/mL) retained significant suppressive effect in both conditions, unmasking a high-end iNOS-independent mechanism.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Paulo de Góes Institute for Microbiology, Federal University of Rio de Janeiro (UFRJ), 21941-590 Rio de Janeiro, RJ, Brazil.

ABSTRACT
Interleukin- (IL-) 17A, a pleiotropic mediator of inflammation and autoimmunity, potently stimulates bone-marrow neutrophil production. To explore IL-17A effects on eosinopoiesis, we cultured bone-marrow from wild-type mice, or mutants lacking inducible nitric oxide synthase (iNOS-/-), CD95 (lpr), IL-17RA, or IL-4, with IL-5, alone or associated with IL-17A. Synergisms between IL-17A-activated, NO-dependent, and NO-independent mechanisms and antagonisms between IL-17A and proallergic factors were further examined. While IL-17A (0.1-10 ng/mL) had no IL-5-independent effect on eosinopoiesis, it dose-dependently suppressed IL-5-induced eosinophil differentiation, by acting during the initial 24 hours. Its effectiveness was abolished by caspase inhibitor, zVAD-fmk. The effect of IL-17A (0.1-1 ng/mL) was sensitive to the iNOS-selective inhibitor aminoguanidine and undetectable in iNOS-/- bone-marrow. By contrast, a higher IL-17A concentration (10 ng/mL) retained significant suppressive effect in both conditions, unmasking a high-end iNOS-independent mechanism. Lower IL-17A concentrations synergized with NO donor nitroprusside. Eosinopoiesis suppression by IL-17A was (a) undetectable in bone-marrow lacking IL-17RA or CD95 and (b) actively prevented by LTD4, LTC4, IL-13, and eotaxin. Sensitivity to IL-17A was increased in bone-marrow lacking IL-4; adding IL-4 to the cultures restored IL-5 responses to control levels. Therefore, effects of both IL-17A and proallergic factors are transduced by the iNOS-CD95 pathway in isolated bone-marrow.

No MeSH data available.


Related in: MedlinePlus