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CXCL12 (SDF-1alpha) suppresses ongoing experimental autoimmune encephalomyelitis by selecting antigen-specific regulatory T cells.

Meiron M, Zohar Y, Anunu R, Wildbaum G, Karin N - J. Exp. Med. (2008)

Bottom Line: The beneficial effect included selection of antigen-specific T cells that were CD4(+)CD25(-)Foxp3(-)IL-10(high), which could adoptively transfer disease resistance, and suppression of Th17 selection.However, in vitro functional analysis of these cells suggested that, even though CXCL12-Ig-induced tolerance is IL-10 dependent, IL-10-independent mechanisms may also contribute to their regulatory function.Collectively, our results not only demonstrate, for the first time, that a chemokine functions as a regulatory mediator, but also suggest a novel way for treating multiple sclerosis and possibly other inflammatory autoimmune diseases.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Bruce Rappaport Faculty of Medicine, Technion, Haifa 31096, Israel.

ABSTRACT
Experimental autoimmune encephalomyelitis (EAE) is a T cell-mediated autoimmune disease of the central nervous system induced by antigen-specific effector Th17 and Th1 cells. We show that a key chemokine, CXCL12 (stromal cell-derived factor 1alpha), redirects the polarization of effector Th1 cells into CD4(+)CD25(-)Foxp3(-)interleukin (IL) 10(high) antigen-specific regulatory T cells in a CXCR4-dependent manner, and by doing so acts as a regulatory mediator restraining the autoimmune inflammatory process. In an attempt to explore the therapeutic implication of these findings, we have generated a CXCL12-immunoglobulin (Ig) fusion protein that, when administered during ongoing EAE, rapidly suppresses the disease in wild-type but not IL-10-deficient mice. Anti-IL-10 neutralizing antibodies could reverse this suppression. The beneficial effect included selection of antigen-specific T cells that were CD4(+)CD25(-)Foxp3(-)IL-10(high), which could adoptively transfer disease resistance, and suppression of Th17 selection. However, in vitro functional analysis of these cells suggested that, even though CXCL12-Ig-induced tolerance is IL-10 dependent, IL-10-independent mechanisms may also contribute to their regulatory function. Collectively, our results not only demonstrate, for the first time, that a chemokine functions as a regulatory mediator, but also suggest a novel way for treating multiple sclerosis and possibly other inflammatory autoimmune diseases.

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CXCL12-Ig redirects the polarization of antigen-specific effector (Th1) cells into IL-10–producing regulatory T cells that suppress EAE. (A) The MOGp35-55 CD4+ T cell line was selected during two subsequent stimulation cycles in the presence of the target antigen and the combination of recombinant mouse IL-12 and anti–IL-4–neutralizing antibodies, and were activated in the third cycle in the presence or absence of 50 μg/ml CXCL12-Ig. Cells were subjected to intracellular cytokine staining (percentages are shown). (B) Cytokine levels in the culture media were also recorded using a standard ELISA method. (C) 3 × 106 T cells per mouse from the CXCL12-Ig–supplemented MOGp35-55 line (closed squares), the MOGp35-55 line (open squares), or PBS (open circles, no cells) were administered to EAE mice on day 12. Mice were monitored daily for the progression of the disease. Results of one out of two independent experiments with similar data (n = 6 mice per each group) are shown as the mean EAE score ± SE. The arrow indicates the day of cell therapy.
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fig6: CXCL12-Ig redirects the polarization of antigen-specific effector (Th1) cells into IL-10–producing regulatory T cells that suppress EAE. (A) The MOGp35-55 CD4+ T cell line was selected during two subsequent stimulation cycles in the presence of the target antigen and the combination of recombinant mouse IL-12 and anti–IL-4–neutralizing antibodies, and were activated in the third cycle in the presence or absence of 50 μg/ml CXCL12-Ig. Cells were subjected to intracellular cytokine staining (percentages are shown). (B) Cytokine levels in the culture media were also recorded using a standard ELISA method. (C) 3 × 106 T cells per mouse from the CXCL12-Ig–supplemented MOGp35-55 line (closed squares), the MOGp35-55 line (open squares), or PBS (open circles, no cells) were administered to EAE mice on day 12. Mice were monitored daily for the progression of the disease. Results of one out of two independent experiments with similar data (n = 6 mice per each group) are shown as the mean EAE score ± SE. The arrow indicates the day of cell therapy.

Mentions: Primary T cells from EAE donors were subjected to two subsequent stimulation cycles in the presence of recombinant mouse IL-12 and anti–IL-4 neutralizing antibodies, and then to a subsequent stimulation in cultures that were or were not supplemented with 50 μg/ml CXCL12-Ig. Intracellular flow cytometry analysis showed that in the absence of CXCL12-Ig, the vast majority of the polarized CD4+ T cells were IFN-γhighIL-4low Th1 cells, whereas the addition of CXCL12-Ig to the culture medium only during the third stimulation cycle redirected the polarization of a significant portion of these cells into IL-10highIL-4low cells (from 1.6 to 23%; Fig. 6 A), resulting in a >10-fold increase in the level of secreted IL-10, as determined by ELISA (from 40 ± 5 to 580 ± 25 pg/ml; P < 0.0001; Fig. 6 B). Notably, the relative number of IL-4highIL-10low CD4+ T cells was also significantly increased (from 0.1 to 9%), and so did the level of IL-4, as determined by ELISA (from 64 ±6 to 215 ± 20 pg/ml; P < 0.001), together with a significant reduction in the production of IFN-γ (from 5,850 ± 430 to 1,930 ± 210 pg/ml; P < 0.001) and TNF-α (from 440 ± 55 to 180 ± 24 pg/ml; P < 0.001). No changes were observed in the level of TGF-β (Fig. 6 B). Intracellular analysis of IL-10 and IFN-γ in these cells clearly showed a highly significant increase in IL-10highIFN-γlow CD4+ T cells (from 1.8 to 22.5%), accompanied by a reciprocal decrease in the number of IL-10lowIFN-γhigh (from 37 to 7.8%) CD4+ T cells after reselection in the presence of CXCL12-Ig (Fig. 6 A). These data further demonstrate the apparent shift from Th1- to IL-10–producing regulatory T cells in the presence of CXCL12.


CXCL12 (SDF-1alpha) suppresses ongoing experimental autoimmune encephalomyelitis by selecting antigen-specific regulatory T cells.

Meiron M, Zohar Y, Anunu R, Wildbaum G, Karin N - J. Exp. Med. (2008)

CXCL12-Ig redirects the polarization of antigen-specific effector (Th1) cells into IL-10–producing regulatory T cells that suppress EAE. (A) The MOGp35-55 CD4+ T cell line was selected during two subsequent stimulation cycles in the presence of the target antigen and the combination of recombinant mouse IL-12 and anti–IL-4–neutralizing antibodies, and were activated in the third cycle in the presence or absence of 50 μg/ml CXCL12-Ig. Cells were subjected to intracellular cytokine staining (percentages are shown). (B) Cytokine levels in the culture media were also recorded using a standard ELISA method. (C) 3 × 106 T cells per mouse from the CXCL12-Ig–supplemented MOGp35-55 line (closed squares), the MOGp35-55 line (open squares), or PBS (open circles, no cells) were administered to EAE mice on day 12. Mice were monitored daily for the progression of the disease. Results of one out of two independent experiments with similar data (n = 6 mice per each group) are shown as the mean EAE score ± SE. The arrow indicates the day of cell therapy.
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Related In: Results  -  Collection

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fig6: CXCL12-Ig redirects the polarization of antigen-specific effector (Th1) cells into IL-10–producing regulatory T cells that suppress EAE. (A) The MOGp35-55 CD4+ T cell line was selected during two subsequent stimulation cycles in the presence of the target antigen and the combination of recombinant mouse IL-12 and anti–IL-4–neutralizing antibodies, and were activated in the third cycle in the presence or absence of 50 μg/ml CXCL12-Ig. Cells were subjected to intracellular cytokine staining (percentages are shown). (B) Cytokine levels in the culture media were also recorded using a standard ELISA method. (C) 3 × 106 T cells per mouse from the CXCL12-Ig–supplemented MOGp35-55 line (closed squares), the MOGp35-55 line (open squares), or PBS (open circles, no cells) were administered to EAE mice on day 12. Mice were monitored daily for the progression of the disease. Results of one out of two independent experiments with similar data (n = 6 mice per each group) are shown as the mean EAE score ± SE. The arrow indicates the day of cell therapy.
Mentions: Primary T cells from EAE donors were subjected to two subsequent stimulation cycles in the presence of recombinant mouse IL-12 and anti–IL-4 neutralizing antibodies, and then to a subsequent stimulation in cultures that were or were not supplemented with 50 μg/ml CXCL12-Ig. Intracellular flow cytometry analysis showed that in the absence of CXCL12-Ig, the vast majority of the polarized CD4+ T cells were IFN-γhighIL-4low Th1 cells, whereas the addition of CXCL12-Ig to the culture medium only during the third stimulation cycle redirected the polarization of a significant portion of these cells into IL-10highIL-4low cells (from 1.6 to 23%; Fig. 6 A), resulting in a >10-fold increase in the level of secreted IL-10, as determined by ELISA (from 40 ± 5 to 580 ± 25 pg/ml; P < 0.0001; Fig. 6 B). Notably, the relative number of IL-4highIL-10low CD4+ T cells was also significantly increased (from 0.1 to 9%), and so did the level of IL-4, as determined by ELISA (from 64 ±6 to 215 ± 20 pg/ml; P < 0.001), together with a significant reduction in the production of IFN-γ (from 5,850 ± 430 to 1,930 ± 210 pg/ml; P < 0.001) and TNF-α (from 440 ± 55 to 180 ± 24 pg/ml; P < 0.001). No changes were observed in the level of TGF-β (Fig. 6 B). Intracellular analysis of IL-10 and IFN-γ in these cells clearly showed a highly significant increase in IL-10highIFN-γlow CD4+ T cells (from 1.8 to 22.5%), accompanied by a reciprocal decrease in the number of IL-10lowIFN-γhigh (from 37 to 7.8%) CD4+ T cells after reselection in the presence of CXCL12-Ig (Fig. 6 A). These data further demonstrate the apparent shift from Th1- to IL-10–producing regulatory T cells in the presence of CXCL12.

Bottom Line: The beneficial effect included selection of antigen-specific T cells that were CD4(+)CD25(-)Foxp3(-)IL-10(high), which could adoptively transfer disease resistance, and suppression of Th17 selection.However, in vitro functional analysis of these cells suggested that, even though CXCL12-Ig-induced tolerance is IL-10 dependent, IL-10-independent mechanisms may also contribute to their regulatory function.Collectively, our results not only demonstrate, for the first time, that a chemokine functions as a regulatory mediator, but also suggest a novel way for treating multiple sclerosis and possibly other inflammatory autoimmune diseases.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Bruce Rappaport Faculty of Medicine, Technion, Haifa 31096, Israel.

ABSTRACT
Experimental autoimmune encephalomyelitis (EAE) is a T cell-mediated autoimmune disease of the central nervous system induced by antigen-specific effector Th17 and Th1 cells. We show that a key chemokine, CXCL12 (stromal cell-derived factor 1alpha), redirects the polarization of effector Th1 cells into CD4(+)CD25(-)Foxp3(-)interleukin (IL) 10(high) antigen-specific regulatory T cells in a CXCR4-dependent manner, and by doing so acts as a regulatory mediator restraining the autoimmune inflammatory process. In an attempt to explore the therapeutic implication of these findings, we have generated a CXCL12-immunoglobulin (Ig) fusion protein that, when administered during ongoing EAE, rapidly suppresses the disease in wild-type but not IL-10-deficient mice. Anti-IL-10 neutralizing antibodies could reverse this suppression. The beneficial effect included selection of antigen-specific T cells that were CD4(+)CD25(-)Foxp3(-)IL-10(high), which could adoptively transfer disease resistance, and suppression of Th17 selection. However, in vitro functional analysis of these cells suggested that, even though CXCL12-Ig-induced tolerance is IL-10 dependent, IL-10-independent mechanisms may also contribute to their regulatory function. Collectively, our results not only demonstrate, for the first time, that a chemokine functions as a regulatory mediator, but also suggest a novel way for treating multiple sclerosis and possibly other inflammatory autoimmune diseases.

Show MeSH
Related in: MedlinePlus