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An IL-9 fate reporter demonstrates the induction of an innate IL-9 response in lung inflammation.

Wilhelm C, Hirota K, Stieglitz B, Van Snick J, Tolaini M, Lahl K, Sparwasser T, Helmby H, Stockinger B - Nat. Immunol. (2011)

Bottom Line: We found that during papain-induced lung inflammation, IL-9 production was largely restricted to innate lymphoid cells (ILCs).IL-9 production by ILCs depended on IL-2 from adaptive immune cells and was rapidly lost in favor of other cytokines, such as IL-13 and IL-5.Blockade of IL-9 production via neutralizing antibodies resulted in much lower expression of IL-13 and IL-5, which suggested that ILCs provide the missing link between the well-established functions of IL-9 in the regulation of type 2 helper T cell cytokines and responses.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Immunology, Medical Research Council National Institute for Medical Research, Mill Hill, UK.

ABSTRACT
Interleukin 9 (IL-9) is a cytokine linked to lung inflammation, but its cellular origin and function remain unclear. Here we describe a reporter mouse strain designed to map the fate of cells that have activated IL-9. We found that during papain-induced lung inflammation, IL-9 production was largely restricted to innate lymphoid cells (ILCs). IL-9 production by ILCs depended on IL-2 from adaptive immune cells and was rapidly lost in favor of other cytokines, such as IL-13 and IL-5. Blockade of IL-9 production via neutralizing antibodies resulted in much lower expression of IL-13 and IL-5, which suggested that ILCs provide the missing link between the well-established functions of IL-9 in the regulation of type 2 helper T cell cytokines and responses.

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Intranasal challenge with IL-33 induces ILC poised for IL-9 productiona) Flow cytometry of lung cells isolated from B6 mice challenged intranasally with PBS, IL-25 or IL-33, stained for surface lineage markers CD4, CD8α, TCRβ, TCRγδ, CD19, Nkp46, Gr-1, CD11c, Ter-119, CD11b (lineage) and Thy-1.2. (upper panel) and lung cells restimulated with PdBU and ionomycin for 2.5 h, stained for intracellular IL-9 and IL-13 and gated on ILC (lower panel). b) Absolute number of ILC (Lin− Thy1.2+ cells) in the lung and c) frequencies of IL-13+ ILC analyzed in the lung after the indicated treatments. d) Cytokine concentration in the supernatant of MACS sorted ILC isolated from PBS, IL-25 or IL-33 treated mice stimulated in vitro with IL-2 overnight. e) Flow cytometry of lung cells isolated from IL9CreR26ReYFP mice treated as in a), stained for lineage markers and Thy1.2 (upper panel) and assessed for eYFP and CD45 expression in ILC (lower panel). Numbers in gates or quadrants (a,e) indicate percent cells in each. Data represents at least two independent experiments with 3 mice in each experimental group (mean ±SEM in b,c,d,e).
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Figure 4: Intranasal challenge with IL-33 induces ILC poised for IL-9 productiona) Flow cytometry of lung cells isolated from B6 mice challenged intranasally with PBS, IL-25 or IL-33, stained for surface lineage markers CD4, CD8α, TCRβ, TCRγδ, CD19, Nkp46, Gr-1, CD11c, Ter-119, CD11b (lineage) and Thy-1.2. (upper panel) and lung cells restimulated with PdBU and ionomycin for 2.5 h, stained for intracellular IL-9 and IL-13 and gated on ILC (lower panel). b) Absolute number of ILC (Lin− Thy1.2+ cells) in the lung and c) frequencies of IL-13+ ILC analyzed in the lung after the indicated treatments. d) Cytokine concentration in the supernatant of MACS sorted ILC isolated from PBS, IL-25 or IL-33 treated mice stimulated in vitro with IL-2 overnight. e) Flow cytometry of lung cells isolated from IL9CreR26ReYFP mice treated as in a), stained for lineage markers and Thy1.2 (upper panel) and assessed for eYFP and CD45 expression in ILC (lower panel). Numbers in gates or quadrants (a,e) indicate percent cells in each. Data represents at least two independent experiments with 3 mice in each experimental group (mean ±SEM in b,c,d,e).

Mentions: Although neither IL-25 nor IL-33 were able to induce IL-9 production from ILC, previous analysis did not test the capacity of these cytokines to induce recruitment of ILC and the potential for subsequent IL-9 induction following exposure to IL-2. To test whether IL-25 and IL-33-induced ILC were able to produce IL-9 upon IL-2 stimulation we intranasally treated wild-type mice with IL-25 or IL-33. Intranasal challenge with IL-33 was much more potent than IL-25 treatment in inducing the accumulation of ILC in the lung (Fig. 4a, b). Although IL-25 challenge resulted in the induction or recruitment of ILC producing IL-13 (Fig 4a and c), this was again higher after IL-33 application. However, while no IL-9-positive cells were detected by intracellular cytokine staining, IL-33 induced high IL-9 production from ILC upon IL-2 stimulation (Fig. 4d). Additionally, IL-33-primed ILC were also more potent producers of IL-5 and IL-13 than IL-25-primed ILC (Fig. 4d). IL-33 was also more potent to induce eYFP+ ILC upon intranasal administration (Fig. 4e) and such ILC expressed IL-13 and displayed similar surface markers as the eYFP+ ILC generated during papain-induced lung inflammation (Supplementary Fig. 6). In order to compare IL-33 and papain-induced ILC with other ILC2 cell types, we investigated the induction of eYFP expression in nuocytes in the mesenteric lymph nodes after intraperitoneal injection of IL-25 and IL-33, NHCs in the FALT and Ih2 cells in the liver. IL-33, but not IL-25 administration induced a small percentage of eYFP+ nuocytes, while no substantial NHCs or Ih2 cells were detected (Supplementary Fig. 7). Thus, the induction of ILC competent to produce IL-9 is mainly driven by IL-33.


An IL-9 fate reporter demonstrates the induction of an innate IL-9 response in lung inflammation.

Wilhelm C, Hirota K, Stieglitz B, Van Snick J, Tolaini M, Lahl K, Sparwasser T, Helmby H, Stockinger B - Nat. Immunol. (2011)

Intranasal challenge with IL-33 induces ILC poised for IL-9 productiona) Flow cytometry of lung cells isolated from B6 mice challenged intranasally with PBS, IL-25 or IL-33, stained for surface lineage markers CD4, CD8α, TCRβ, TCRγδ, CD19, Nkp46, Gr-1, CD11c, Ter-119, CD11b (lineage) and Thy-1.2. (upper panel) and lung cells restimulated with PdBU and ionomycin for 2.5 h, stained for intracellular IL-9 and IL-13 and gated on ILC (lower panel). b) Absolute number of ILC (Lin− Thy1.2+ cells) in the lung and c) frequencies of IL-13+ ILC analyzed in the lung after the indicated treatments. d) Cytokine concentration in the supernatant of MACS sorted ILC isolated from PBS, IL-25 or IL-33 treated mice stimulated in vitro with IL-2 overnight. e) Flow cytometry of lung cells isolated from IL9CreR26ReYFP mice treated as in a), stained for lineage markers and Thy1.2 (upper panel) and assessed for eYFP and CD45 expression in ILC (lower panel). Numbers in gates or quadrants (a,e) indicate percent cells in each. Data represents at least two independent experiments with 3 mice in each experimental group (mean ±SEM in b,c,d,e).
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Figure 4: Intranasal challenge with IL-33 induces ILC poised for IL-9 productiona) Flow cytometry of lung cells isolated from B6 mice challenged intranasally with PBS, IL-25 or IL-33, stained for surface lineage markers CD4, CD8α, TCRβ, TCRγδ, CD19, Nkp46, Gr-1, CD11c, Ter-119, CD11b (lineage) and Thy-1.2. (upper panel) and lung cells restimulated with PdBU and ionomycin for 2.5 h, stained for intracellular IL-9 and IL-13 and gated on ILC (lower panel). b) Absolute number of ILC (Lin− Thy1.2+ cells) in the lung and c) frequencies of IL-13+ ILC analyzed in the lung after the indicated treatments. d) Cytokine concentration in the supernatant of MACS sorted ILC isolated from PBS, IL-25 or IL-33 treated mice stimulated in vitro with IL-2 overnight. e) Flow cytometry of lung cells isolated from IL9CreR26ReYFP mice treated as in a), stained for lineage markers and Thy1.2 (upper panel) and assessed for eYFP and CD45 expression in ILC (lower panel). Numbers in gates or quadrants (a,e) indicate percent cells in each. Data represents at least two independent experiments with 3 mice in each experimental group (mean ±SEM in b,c,d,e).
Mentions: Although neither IL-25 nor IL-33 were able to induce IL-9 production from ILC, previous analysis did not test the capacity of these cytokines to induce recruitment of ILC and the potential for subsequent IL-9 induction following exposure to IL-2. To test whether IL-25 and IL-33-induced ILC were able to produce IL-9 upon IL-2 stimulation we intranasally treated wild-type mice with IL-25 or IL-33. Intranasal challenge with IL-33 was much more potent than IL-25 treatment in inducing the accumulation of ILC in the lung (Fig. 4a, b). Although IL-25 challenge resulted in the induction or recruitment of ILC producing IL-13 (Fig 4a and c), this was again higher after IL-33 application. However, while no IL-9-positive cells were detected by intracellular cytokine staining, IL-33 induced high IL-9 production from ILC upon IL-2 stimulation (Fig. 4d). Additionally, IL-33-primed ILC were also more potent producers of IL-5 and IL-13 than IL-25-primed ILC (Fig. 4d). IL-33 was also more potent to induce eYFP+ ILC upon intranasal administration (Fig. 4e) and such ILC expressed IL-13 and displayed similar surface markers as the eYFP+ ILC generated during papain-induced lung inflammation (Supplementary Fig. 6). In order to compare IL-33 and papain-induced ILC with other ILC2 cell types, we investigated the induction of eYFP expression in nuocytes in the mesenteric lymph nodes after intraperitoneal injection of IL-25 and IL-33, NHCs in the FALT and Ih2 cells in the liver. IL-33, but not IL-25 administration induced a small percentage of eYFP+ nuocytes, while no substantial NHCs or Ih2 cells were detected (Supplementary Fig. 7). Thus, the induction of ILC competent to produce IL-9 is mainly driven by IL-33.

Bottom Line: We found that during papain-induced lung inflammation, IL-9 production was largely restricted to innate lymphoid cells (ILCs).IL-9 production by ILCs depended on IL-2 from adaptive immune cells and was rapidly lost in favor of other cytokines, such as IL-13 and IL-5.Blockade of IL-9 production via neutralizing antibodies resulted in much lower expression of IL-13 and IL-5, which suggested that ILCs provide the missing link between the well-established functions of IL-9 in the regulation of type 2 helper T cell cytokines and responses.

View Article: PubMed Central - PubMed

Affiliation: Division of Molecular Immunology, Medical Research Council National Institute for Medical Research, Mill Hill, UK.

ABSTRACT
Interleukin 9 (IL-9) is a cytokine linked to lung inflammation, but its cellular origin and function remain unclear. Here we describe a reporter mouse strain designed to map the fate of cells that have activated IL-9. We found that during papain-induced lung inflammation, IL-9 production was largely restricted to innate lymphoid cells (ILCs). IL-9 production by ILCs depended on IL-2 from adaptive immune cells and was rapidly lost in favor of other cytokines, such as IL-13 and IL-5. Blockade of IL-9 production via neutralizing antibodies resulted in much lower expression of IL-13 and IL-5, which suggested that ILCs provide the missing link between the well-established functions of IL-9 in the regulation of type 2 helper T cell cytokines and responses.

Show MeSH
Related in: MedlinePlus