Intragraft memory-like CD127hiCD4+Foxp3+ Tregs maintain transplant tolerance
Yuanfei Zhao, Leigh Nicholson, Hannah Wang, Yi Wen Qian, Wayne J. Hawthorne, Elvira Jimenez-Vera, Brian S. Gloss, Joey Lai, Adwin Thomas, Yi Vee Chew, Heather Burns, Geoff Y. Zhang, Yuan Min Wang, Natasha M. Rogers, Guoping Zheng, Shounan Yi, Stephen I. Alexander, Philip J. O’Connell, Min Hu
Yuanfei Zhao, Leigh Nicholson, Hannah Wang, Yi Wen Qian, Wayne J. Hawthorne, Elvira Jimenez-Vera, Brian S. Gloss, Joey Lai, Adwin Thomas, Yi Vee Chew, Heather Burns, Geoff Y. Zhang, Yuan Min Wang, Natasha M. Rogers, Guoping Zheng, Shounan Yi, Stephen I. Alexander, Philip J. O’Connell, Min Hu
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Research Article Transplantation

Intragraft memory-like CD127hiCD4+Foxp3+ Tregs maintain transplant tolerance

Abstract

CD4+Foxp3+ regulatory T cells (Tregs) play an essential role in suppressing transplant rejection, but their role within the graft and heterogeneity in tolerance are poorly understood. Here, we compared phenotypic and transcriptomic characteristics of Treg populations within lymphoid organs and grafts in an islet xenotransplant model of tolerance. We showed Tregs were essential for tolerance induction and maintenance. Tregs demonstrated heterogeneity within the graft and lymphoid organs of tolerant mice. A subpopulation of CD127hi Tregs with memory features were found in lymphoid organs, presented in high proportions within long-surviving islet grafts, and had a transcriptomic and phenotypic profile similar to tissue Tregs. Importantly, these memory-like CD127hi Tregs were better able to prevent rejection by effector T cells, after adoptive transfer into secondary Rag–/– hosts, than naive Tregs or unselected Tregs from tolerant mice. Administration of IL-7 to the CD127hi Treg subset was associated with a strong activation of phosphorylation of STAT5. We proposed that memory-like CD127hi Tregs developed within the draining lymph node and underwent further genetic reprogramming within the graft toward a phenotype that had shared characteristics with other tissue or tumor Tregs. These findings suggested that engineering Tregs with these characteristics either in vivo or for adoptive transfer could enhance transplant tolerance.

Authors

Yuanfei Zhao, Leigh Nicholson, Hannah Wang, Yi Wen Qian, Wayne J. Hawthorne, Elvira Jimenez-Vera, Brian S. Gloss, Joey Lai, Adwin Thomas, Yi Vee Chew, Heather Burns, Geoff Y. Zhang, Yuan Min Wang, Natasha M. Rogers, Guoping Zheng, Shounan Yi, Stephen I. Alexander, Philip J. O’Connell, Min Hu

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Figure 1

Depletion of Tregs in induction phase abolishes transplant tolerance induced by short-term treatment with CTLA4-Fc/MR1 in DEREG mouse recipients of porcine NICC grafts.

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Depletion of Tregs in induction phase abolishes transplant tolerance ind...
(A) Schematic illustration of CTLA4-Fc/MR1 treatment (tolerant group) and Treg depletion (depletion group) timelines for DEREG recipients receiving porcine NICC transplants. DEREG recipients assigned to the rejection group did not receive any treatments. Samples were collected from all groups on days 8, 20, and ≥100 after transplantation. (B) Functional assessments of porcine NICC transplants in DEREG recipients day 100. Micrographs of H&E (left) and immunohistochemistry (IHC) insulin-positive staining (brown) (right) of porcine NICC grafts in the tolerant group are shown. Data for serum porcine C-peptide in tolerant group (n = 69), rejection group (n = 12), depletion group (n = 5), and control DEREG mice, which were neither transplanted nor treated with CTLA4-Fc/MR1 (naive group/mice) (n = 6). (C) Confirmation of Treg depletion in CTLA4-Fc/MR1–treated DEREG recipients with the DT injection regimen. Representative pseudocolor plots of CD4 versus GFP (gated on CD4+ T cells) showing CD4+GFP+ Tregs in peripheral blood mononuclear cells (PBMCs) from deletion group recipients before transplant and days 0, 3, and 10 after transplant. The proportion of CD4+GFP+ Tregs in CD4+ T cells of PBMCs in the depletion group (n = 5) and tolerant group (n = 3) before treatment and on day 0, day 3, day 10, and day 15. (D) Micrographs of H&E-stained porcine NICC grafts at day 8 and day 20 and immunofluorescence insulin-stained porcine NICC grafts day 20 in the rejection, tolerant, and depletion groups. The positive insulin staining was shown in red. H&E and IHC images were visualized using Aperio ImageScope (v.12.4.0.7018) software (Leica Biosystems). The immunofluorescence images were imaged on Olympus FV ≥1000 confocal microscope with FV10-ASW 4.2 software. Scale bar: 200 μm (B and H&E, D); 50 μm (immunofluorescence, D) Kruskal-Wallis test was used in B, and Mann-Whitney test was used in C. Data were from 9 independent experiments and shown as mean ± SEM. Label of statistical significance: *P < 0.05, ***P < 0.001, ****P < 0.0001.