Allograft rejection is restrained by short-lived TIM-3+PD-1+Foxp3+ Tregs
Shipra Gupta, … , Vijay K. Kuchroo, Terry B. Strom
Shipra Gupta, … , Vijay K. Kuchroo, Terry B. Strom
Published June 11, 2012
Citation Information: J Clin Invest. 2012;122(7):2395-2404. https://doi.org/10.1172/JCI45138.
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Research Article Immunology

Allograft rejection is restrained by short-lived TIM-3+PD-1+Foxp3+ Tregs

Abstract

Tregs play a pivotal role in inducing and maintaining donor-specific transplant tolerance. The T cell immunoglobulin and mucin domain-3 protein (TIM-3) is expressed on many fully activated effector T cells. Along with program death 1 (PD-1), TIM-3 is used as a marker for exhausted effector T cells, and interaction with its ligand, galectin-9, leads to selective death of TIM-3+ cells. We report herein the presence of a galectin-9–sensitive CD4+FoxP3+TIM-3+ population of T cells, which arose from CD4+FoxP3+TIM-3– proliferating T cells in vitro and in vivo and were often PD-1+. These cells became very prominent among graft-infiltrating Tregs during allograft response. The frequency and number of TIM-3+ Tregs peaked at the time of graft rejection and declined thereafter. Moreover, these cells also arise in a tolerance-promoting donor-specific transfusion model, representing a pool of proliferating, donor-specific Tregs. Compared with TIM-3– Tregs, TIM-3+ Tregs, which are often PD-1+ as well, exhibited higher in vitro effector function and more robust expression of CD25, CD39, CD73, CTLA-4, IL-10, and TGF-β but not galectin-9. However, these TIM-3+ Tregs did not flourish when passively transferred to newly transplanted hosts. These data suggest that a heretofore unrecognized graft-infiltrating, short-lived subset of Tregs can restrain rejection.

Authors

Shipra Gupta, Thomas B. Thornley, Wenda Gao, Rafael Larocca, Laurence A. Turka, Vijay K. Kuchroo, Terry B. Strom

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Figure 5

CD4+TIM-3+ Tregs are potent suppressors of Teff proliferation in an in vitro MLR culture and robustly express Treg markers.

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CD4+TIM-3+ Tregs are potent suppressors of Teff proliferation in an in v...
(A) In an MLR culture, naive CD4+GFP– Teffs from spleens of BALB/c-KI mice were stimulated with irradiated CD90– splenocytes from BALB/c-KI mice for 3 days in the presence or absence of Tregs. Sorted CD4+GFP+TIM-3+/TIM-3– Tregs were obtained from spleens of BALB/c-KI mice injected with DBA/2 DST 5 days prior to harvesting. T cell proliferation in these cultures was analyzed by the mean values of incorporated thymidine of duplicate/triplicate wells and compared with that of MLR cultures with Teff alone (normalized as 100%). Data are represented as mean ± SEM; n = 3 independent experiments done in duplicate, triplicates, or quadruplicates leading to a total of ≥ 8 independent values for each condition. **P = 0.002; ***P < 0.001. (B) TIM-3+GFP+FoxP3+ and TIM-3–GFP+FoxP3+ cells were sorted from harvested dLNs of 30 to 35 C57BL/6-KI mice grafted with BALB/c skin on day 7. RNA was isolated from sorted TIM-3+GFP+FoxP3+ and TIM-3–GFP+FoxP3+ populations. Gene expression of various markers was assessed by quantitative real-time PCR. Data are representative of 3 independent experiments. (C) CD4+TIM-3+ Tregs and CD4+TIM-3– Tregs harvested from the dLNs on day 7 were stained and analyzed by flow cytometry for various Treg functional markers as shown. Arrows in the histograms depict TIM-3– Tregs (a) and TIM-3+ Tregs (b). Numbers represent the percentage of TIM-3+ and TIM-3– Tregs, respectively. n ≥ 5 mice.