Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2013 Aug 1;191(3):1110-7.
doi: 10.4049/jimmunol.1203089. Epub 2013 Jun 24.

Distinct chemokine receptor axes regulate Th9 cell trafficking to allergic and autoimmune inflammatory sites

Ervin E Kara  1 , Iain Comerford, Cameron R Bastow, Kevin A Fenix, Wendel Litchfield, Tracy M Handel, Shaun R McColl
Affiliations

Distinct chemokine receptor axes regulate Th9 cell trafficking to allergic and autoimmune inflammatory sites

Ervin E Kara et al. J Immunol. .

Abstract

Migration of Th cells to peripheral sites of inflammation is essential for execution of their effector function. The recently described Th9 subset characteristically produces IL-9 and has been implicated in both allergy and autoimmunity. Despite this, the migratory properties of Th9 cells remain enigmatic. In this study, we examined chemokine receptor usage by Th9 cells and demonstrate, in models of allergy and autoimmunity, that these cells express functional CCR3, CCR6, and CXCR3, chemokine receptors commonly associated with other, functionally opposed effector Th subsets. Most Th9 cells that express CCR3 also express CXCR3 and CCR6, and expression of these receptors appears to account for the recruitment of Th9 cells to disparate inflammatory sites. During allergic inflammation, Th9 cells use CCR3 and CCR6, but not CXCR3, to home to the peritoneal cavity, whereas Th9 homing to the CNS during experimental autoimmune encephalomyelitis involves CXCR3 and CCR6 but not CCR3. To our knowledge, these data provide the first insights into regulation of Th9 cell trafficking in allergy and autoimmunity.

PubMed Disclaimer

Conflict of interest statement

CONFLICT OF INTEREST

The authors disclose no competing financial interests.

Figures

Figure 1
Figure 1. In vitro-generated TH9 cells express chemokine receptors associated with other effector TH cell subsets
(A) Representative histograms of chemokine receptor expression on in vitro-generated TH cell subsets. Mean ± SEM over 3 independent experiments indicated within respective histograms. Grey histogram: isotype control; open histogram: anti-CKR staining. (B) Flow cytometric analyses of chemokine receptor mean fluorescence intensity (MFI) on in vitro-generated TH9 cells compared with TH0, TH1, TH17, iTreg and TH2 cells. Chemokine receptor MFI was calculated by subtracting respective isotype-matched negative control staining from anti-CKR staining MFI. Data are presented as mean ± SEM (n=3 independent experiments). Comparisons of MFI on TH9 cells compared with other TH subsets were performed using a one-way ANOVA with a Bonferroni’s multiple comparison post-test. * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001. (C) Quantitative PCR analysis of Ccr3, Ccr6 and Cxcr3 mRNA expression by TH0 and TH9 cells generated from FACS-purified naïve CD4+ T cell precursors, presented relative to the house-keeping gene Rplp0. Data are presented as mean ± SD and are representative of one of two independent experiments performed with similar results. * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001.
Figure 2
Figure 2. In vitro-generated TH9 cells co-express CCR3, CCR6 and CXCR3
(A) Representative FACS contour plots of dual chemokine receptor expression on TH0 and TH9 cells generated in vitro. (B) CCR6 expression on CCR3−CXCR3−, CCR3+CXCR3−, CCR3−CXCR3+ and CCR3−CXCR3+ TH9 cell populations. Quantitation is shown. Data are presented as mean ± SEM (n=3 independent experiments). * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001.
Figure 3
Figure 3. Functional expression of CCR3, CCR6 and CXCR3 on TH9 cells
(A) Transwell chemotaxis of TH9 cells relative to naïve, TH1 and TH17 cells. Migration index was calculated by dividing the number of positive events in test wells by the number of positive events in which no chemokine was added to the bottom chamber. Cells were enumerated as follows: naïve CD4+: CD4+CD8−CD62L+CD44lo; TH1: CD4+CD8−IFN-γ+IL-17A−; TH17: CD4+CD8−IFN-γ−IL-17A+; TH9: CD4+CD8−IFN-γ−IL-9+. Data presented as mean ± SEM (n=3 independent experiments). (B) Inhibition of TH9 cell migration to CCL11, CCL20 and CXCL11 using chemokine receptor antagonists. Migration assays were performed as in (A) with the inclusion of the indicated chemokine receptor antagonist in the lower chamber. Data presented as mean ± SEM (n=2 independent experiments). * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001.
Figure 4
Figure 4. TH9 cells generated in vivo during allergic inflammation express CCR3, CCR6 and CXCR3
(A) Representative FACS contour plots of TH9 cells generated in a type-2 immunisation strategy. BALB/c mice were sensitised with OVA/AlOH gel on days 0, 3 and 7, inguinal lymph nodes collected 7 days later and TH9 cells identified in flow cytometry as live CD3+CD4+, IL-4/IL-17A/FoxP3− (bottom gate in left plot), IL-9+ (gated in right plot). (B) Representative FACS histograms of CCR3, CCR6 and CXCR3 expression on TH9 cells gated in (A) with % of chemokine receptor positive TH9 cells indicated. Grey histogram: isotype control; open histogram: anti-CKR staining. Quantitation is shown (n=5 mice).
Figure 5
Figure 5. TH9 cells generated in vivo during PLP-induced EAE express CCR3, CCR6 and CXCR3
(A) Representative FACS contour plots of TH9 cells generated in a type-1/-17 immunisation strategy. SJL/j mice were immunised with PLP139–151/CFA, inguinal lymph nodes collected 10 days later and TH9 cells identified in flow cytometry as live CD3+CD4+, IL-17A/FoxP3− (bottom gate in left plot), IL-9+ (gated in right plot). (D) Representative histograms of CCR3, CCR6 and CXCR3 expression on TH9 cells gated in (B) with % of chemokine receptor positive TH9 cells indicated. Grey histogram: isotype control; open histogram: anti-CKR staining. Quantitation is shown (n=5 mice).
Figure 6
Figure 6. CCR3 and CCR6 mediate recruitment of TH9 cells to an allergic microenvironment
(A) Schematic of experimental strategy. (B) Peritoneal cavity (PC)-infiltrated TH9 cells 6hrs post-OVA challenge. Representative FACS contour plots of peritoneal cavity-infiltrated TH9 cells (CD4+IL-9+) 6hrs post-OVA challenge. Plots are pre-gated on live IL-4/IL-17A/FoxP3 (Dump)− lymphocytes. Number and proportion of TH9 cells in the PC 6hrs post-OVA challenge is shown. (C) Number and proportion of TH9 cells 6hrs post-OVA challenge in lymph node and peripheral blood (live IL-4/IL-17A/FoxP3 (Dump)− lymphocytes that are CD4+IL-9+). Data presented as mean ± SEM (n=4 mice/group). Two tailed unpaired student t-tests were performed where appropriate. * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001.
Figure 7
Figure 7. CCR6 and CXCR3 mediate recruitment of TH9 cells to an autoimmune microenvironment
(A) Schematic of experimental strategy. (B) CNS-infiltrated TH9 cells 15 days post-EAE induction. Representative FACS contour plots of CNS-infiltrated TH9 cells (CD4+IL-9+) on d15-post EAE induction. Plots are pre-gated on live IL-17A/FoxP3 (Dump)−. Number and proportion of TH9 cells in CNS d15 post-EAE induction is shown. (C) Number and proportion of TH9 cells in lymph node and peripheral blood (live IL-4/IL-17A/FoxP3 (Dump)− lymphocytes that are CD4+IL-9+). Data presented as mean ± SEM (n=3–4 mice/group). Two tailed unpaired student t-tests were performed where appropriate. * denotes p<0.05; ** denotes p<0.01; *** denotes p<0.001.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Veldhoen M, Uyttenhove C, van Snick J, Helmby H, Westendorf A, Buer J, Martin B, Wilhelm C, Stockinger B. Transforming growth factor-beta 'reprograms' the differentiation of T helper 2 cells and promotes an interleukin 9-producing subset. Nat Immunol. 2008;9:1341–1346. - PubMed
    1. Dardalhon V, Awasthi A, Kwon H, Galileos G, Gao W, Sobel RA, Mitsdoerffer M, Strom TB, Elyaman W, Ho IC, Khoury S, Oukka M, Kuchroo VK. IL-4 inhibits TGF-beta-induced Foxp3+ T cells and, together with TGF-beta, generates IL-9+ IL-10+ Foxp3(-) effector T cells. Nat Immunol. 2008;9:1347–1355. - PMC - PubMed
    1. Jabeen R, Kaplan MH. The symphony of the ninth: the development and function of Th9 cells. Curr Opin Immunol. 2012;24:303–307. - PMC - PubMed
    1. McMillan SJ, Bishop B, Townsend MJ, McKenzie AN, Lloyd CM. The absence of interleukin 9 does not affect the development of allergen-induced pulmonary inflammation nor airway hyperreactivity. J Exp Med. 2002;195:51–57. - PMC - PubMed
    1. Cheng G, Arima M, Honda K, Hirata H, Eda F, Yoshida N, Fukushima F, Ishii Y, Fukuda T. Anti-interleukin-9 antibody treatment inhibits airway inflammation and hyperreactivity in mouse asthma model. Am J Respir Criti Care Med. 2002;166:409–416. - PubMed

Publication types

MeSH terms

LinkOut - more resources