T regulatory cell chemokine production mediates pathogenic T cell attraction and suppression
Scott J. Patterson, Anne M. Pesenacker, Adele Y. Wang, Jana Gillies, Majid Mojibian, Kim Morishita, Rusung Tan, Timothy J. Kieffer, C. Bruce Verchere, Constadina Panagiotopoulos, Megan K. Levings
Scott J. Patterson, Anne M. Pesenacker, Adele Y. Wang, Jana Gillies, Majid Mojibian, Kim Morishita, Rusung Tan, Timothy J. Kieffer, C. Bruce Verchere, Constadina Panagiotopoulos, Megan K. Levings
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Research Article Immunology

T regulatory cell chemokine production mediates pathogenic T cell attraction and suppression

Abstract

T regulatory cells (Tregs) control immune homeostasis by preventing inappropriate responses to self and nonharmful foreign antigens. Tregs use multiple mechanisms to control immune responses, all of which require these cells to be near their targets of suppression; however, it is not known how Treg-to-target proximity is controlled. Here, we found that Tregs attract CD4+ and CD8+ T cells by producing chemokines. Specifically, Tregs produced both CCL3 and CCL4 in response to stimulation, and production of these chemokines was critical for migration of target T cells, as Tregs from Ccl3–/– mice, which are also deficient for CCL4 production, did not promote migration. Moreover, CCR5 expression by target T cells was required for migration of these cells to supernatants conditioned by Tregs. Tregs deficient for expression of CCL3 and CCL4 were impaired in their ability to suppress experimental autoimmune encephalomyelitis or islet allograft rejection in murine models. Moreover, Tregs from subjects with established type 1 diabetes were impaired in their ability to produce CCL3 and CCL4. Together, these results demonstrate a previously unappreciated facet of Treg function and suggest that chemokine secretion by Tregs is a fundamental aspect of their therapeutic effect in autoimmunity and transplantation.

Authors

Scott J. Patterson, Anne M. Pesenacker, Adele Y. Wang, Jana Gillies, Majid Mojibian, Kim Morishita, Rusung Tan, Timothy J. Kieffer, C. Bruce Verchere, Constadina Panagiotopoulos, Megan K. Levings

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Figure 4

Tregs attract CD4+ and CD8+ T cells in vitro via a CCL3/4-CCR5–dependent mechanism.

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Tregs attract CD4+ and CD8+ T cells in vitro via a CCL3/4-CCR5–dependent...
Tregs and Tconv cells were sorted from WT or Ccl3–/– mice, and CD4+ or CD8+ T cells were sorted from WT or Ccr5–/– mice. (A–C) Supernatants from TCR-stimulated Tregs or Tconv cells were placed in the bottom of a Transwell chamber, and CD4+ or CD8+ T cells were placed on top. The number of migrating cells is expressed as a fold increase compared with medium alone. Data are the average ± SEM; statistical analysis by t test (A, n = 4 B, n = 5) or 1-way ANOVA with Bonferroni’s post-hoc test (C, n = 9). (D) 200,000 ex vivo Thy1.1+CD4+ or Thy1.1+CD8+ T cells were placed in the upper chamber of a Transwell plate in the absence or presence of 200,000 stimulated Thy1.2+ CD4+ Tconv cells or Tregs from WT or Ccl3–/– mice or media alone. The bottom chambers contained supernatants from the indicated stimulated or unstimulated cells. After 3 hours, the numbers of Thy1.1+ cells in the bottom chamber were counted by flow cytometry. Shown are averaged data from 3 independent experiments, represented as means with SEM, 2-way ANOVA with Bonferroni’s post-hoc test. (E) 300,000 ex vivo CD8+ or CD4+ cells were placed in the upper chamber of a Transwell plate, and the bottom chambers contained supernatants from the stimulated Tregs or media in the absence or presence of neutralizing antibodies (1 mg/ml) against CCL3 and/or CCL4 as indicated. After 3 hours, the numbers of migrating cells in the bottom chamber were counted by flow cytometry. The absolute number of migrating cells is shown. Data are the average ± SEM of technical duplicates, 2-way ANOVA with Bonferroni’s post-hoc test. *P < 0.05; **P < 0.01; ***P < 0.005; ****P < 0.0001.