CXCL11-dependent induction of FOXP3-negative regulatory T cells suppresses autoimmune encephalomyelitis
Yaniv Zohar, Gizi Wildbaum, Rostislav Novak, Andrew L. Salzman, Marcus Thelen, Ronen Alon, Yiftah Barsheshet, Christopher L. Karp, Nathan Karin
Yaniv Zohar, Gizi Wildbaum, Rostislav Novak, Andrew L. Salzman, Marcus Thelen, Ronen Alon, Yiftah Barsheshet, Christopher L. Karp, Nathan Karin
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Research Article

CXCL11-dependent induction of FOXP3-negative regulatory T cells suppresses autoimmune encephalomyelitis

Abstract

A single G protein–coupled receptor (GPCR) can activate multiple signaling cascades based on the binding of different ligands. The biological relevance of this feature in immune regulation has not been evaluated. The chemokine-binding GPCR CXCR3 is preferentially expressed on CD4+ T cells, and canonically binds 3 structurally related chemokines: CXCL9, CXCL10, and CXCL11. Here we have shown that CXCL10/CXCR3 interactions drive effector Th1 polarization via STAT1, STAT4, and STAT5 phosphorylation, while CXCL11/CXCR3 binding induces an immunotolerizing state that is characterized by IL-10hi (Tr1) and IL-4hi (Th2) cells, mediated via p70 kinase/mTOR in STAT3- and STAT6-dependent pathways. CXCL11 binds CXCR3 with a higher affinity than CXCL10, suggesting that CXCL11 has the potential to restrain inflammatory autoimmunity. We generated a CXCL11-Ig fusion molecule and evaluated its use in the EAE model of inflammatory autoimmune disease. Administration of CXCL11-Ig during the first episode of relapsing EAE in SJL/J mice not only led to rapid remission, but also prevented subsequent relapse. Using GFP-expressing effector CD4+ T cells, we observed that successful therapy was associated with reduced accumulation of these cells at the autoimmune site. Finally, we showed that very low doses of CXCL11 rapidly suppress signs of EAE in C57BL/6 mice lacking functional CXCL11.

Authors

Yaniv Zohar, Gizi Wildbaum, Rostislav Novak, Andrew L. Salzman, Marcus Thelen, Ronen Alon, Yiftah Barsheshet, Christopher L. Karp, Nathan Karin

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Figure 1

Distinct CXCR3 ligands differentially polarize CD4+ T cells in a CXCR3-dependent manner.

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Distinct CXCR3 ligands differentially polarize CD4+ T cells in a CXCR3-d...
(A) CXCR3 expression on CD4+CD3+ cells increased over the first 24 hours of anti-CD3/anti-CD28 activation, as detected by flow cytometry. (B) CD4+ T cells were anti-CD3/anti-CD28 activated in cultures supplemented with CXCL10 or CXCL11, and IL-4, IL-17, and IFN-γ were detected by flow cytometry. CXCL10 polarized IFN-γhiIL-4lo Th1 cells, whereas CXCL11 polarized IL-10hi Tr1 cells and IFN-γloIL-4hi Th2 cells. (C) After anti-CD3/anti-CD28 activation, CD4+ T cells were analyzed for CXCR3 and FOXP3 expression. A small portion of CXCR3+CD4+ T cells were FOXP3+ (3.3%). (D) 24 hours after anti-CD3/anti-CD28 activation of CD4+ T cells, cultures were supplemented with CXCL10 or CXCL11. 48 hours later, IL-10 was determined by flow cytometry. CXCL11 did not induce IL-10 production in FOXP3+CD4+ cells. (E) CD4+ T cells from SJL/J mice were anti-CD3/anti-CD28–activated; 24 hours later, cultures were supplemented with CXCL11 or CXCL10. 48 hours later, cytokine levels were determined by ELISA. CXCL11 induced IL-10 production, whereas CXCL10 induced IFN-γ production. (F) Freshly isolated human CD4+ T cells from healthy donors were activated with anti-CD3/anti-CD28. After 24 hours, cultures were supplemented with CXCL10 or CXCL11 together with anti-CXCR3 blocking mAb, anti-CXCR7 blocking mAb (R&D), PBS, or isotype-matched control IgG. 48 hours later, IL-10 and IL-4 levels were recorded. In all experiments, recombinant chemokines were added at 100 ng/ml (R&D systems). Data (mean ± SEM of triplicates) are from 1 of 3 independent experiments with similar results. *P < 0.001.