Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells
Due to the intracellular expression of Foxp3 it is impossible to purify viable Foxp3+ cells on the basis of Foxp3 staining. Consequently CD4+Foxp3+ regulatory T cells (Tregs) in mice have mostly been characterized using CD4+CD25+ T cells or GFP-Foxp3 reporter T cells. However, these two populations...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2024-01-01
|
| Series: | Frontiers in Immunology |
| Subjects: | |
| Online Access: | https://www.frontiersin.org/articles/10.3389/fimmu.2024.1292158/full |
| _version_ | 1797346877891411968 |
|---|---|
| author | Zilei Liu Zilei Liu Zilei Liu Katherine J. Baines Katherine J. Baines Natalie M. Niessen Munish K. Heer David Clark David Clark G. Alexander Bishop Paul R. Trevillian Paul R. Trevillian |
| author_facet | Zilei Liu Zilei Liu Zilei Liu Katherine J. Baines Katherine J. Baines Natalie M. Niessen Munish K. Heer David Clark David Clark G. Alexander Bishop Paul R. Trevillian Paul R. Trevillian |
| author_sort | Zilei Liu |
| collection | DOAJ |
| description | Due to the intracellular expression of Foxp3 it is impossible to purify viable Foxp3+ cells on the basis of Foxp3 staining. Consequently CD4+Foxp3+ regulatory T cells (Tregs) in mice have mostly been characterized using CD4+CD25+ T cells or GFP-Foxp3 reporter T cells. However, these two populations cannot faithfully represent Tregs as the expression of CD25 and Foxp3 does not completely overlap and GFP+Foxp3+ reporter T cells have been reported to be functionally altered. The aim of this study was to characterize normal Tregs without separating Foxp3+ and Foxp3- cells for the expression of the main functional molecules and proliferation behaviors by flow cytometry and to examine their gene expression characteristics through differential gene expression. Our data showed that the expressions of Foxp3, CD25, CTLA-4 (both intracellular and cell surface) and PD-1 was mostly confined to CD4+ T cells and the expression of Foxp3 did not completely overlap with the expression of CD25, CTLA-4 or PD-1. Despite higher levels of expression of the T cell inhibitory molecules CTLA-4 and PD-1, Tregs maintained higher levels of Ki-67 expression in the homeostatic state and had greater proliferation in vivo after allo-activation than Tconv. Differential gene expression analysis revealed that resting Tregs exhibited immune activation markers characteristic of activated Tconv. This is consistent with the flow data that the T cell activation markers CD25, CTLA-4, PD-1, and Ki-67 were much more strongly expressed by Tregs than Tconv in the homeostatic state. |
| first_indexed | 2024-03-08T11:39:34Z |
| format | Article |
| id | doaj.art-943c498dcdd649c7a2c1a8e047584bee |
| institution | Directory Open Access Journal |
| issn | 1664-3224 |
| language | English |
| last_indexed | 2024-03-08T11:39:34Z |
| publishDate | 2024-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Immunology |
| spelling | doaj.art-943c498dcdd649c7a2c1a8e047584bee2024-01-25T08:01:33ZengFrontiers Media S.A.Frontiers in Immunology1664-32242024-01-011510.3389/fimmu.2024.12921581292158Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cellsZilei Liu0Zilei Liu1Zilei Liu2Katherine J. Baines3Katherine J. Baines4Natalie M. Niessen5Munish K. Heer6David Clark7David Clark8G. Alexander Bishop9Paul R. Trevillian10Paul R. Trevillian11Transplant Unit, John Hunter Hospital, Newcastle, NSW, AustraliaTransplant and Surgical Immunology Theme, Immune Health Research Program, Hunter Medical Research Institute, The University of Newcastle, Newcastle, NSW, AustraliaSchool of Medicine and Public Health, College of Medicine, Health and Wellbeing, The University of Newcastle, Newcastle, NSW, AustraliaTransplant and Surgical Immunology Theme, Immune Health Research Program, Hunter Medical Research Institute, The University of Newcastle, Newcastle, NSW, AustraliaSchool of Biomedical Sciences and Pharmacy, College of Medicine, Health and Wellbeing, The University of Newcastle, Newcastle, NSW, AustraliaTransplant and Surgical Immunology Theme, Immune Health Research Program, Hunter Medical Research Institute, The University of Newcastle, Newcastle, NSW, AustraliaTransplant Unit, John Hunter Hospital, Newcastle, NSW, AustraliaTransplant Unit, John Hunter Hospital, Newcastle, NSW, AustraliaSchool of Medicine and Public Health, College of Medicine, Health and Wellbeing, The University of Newcastle, Newcastle, NSW, AustraliaTransplantation Immunobiology Group, University of Sydney Central Clinical School, Charles Perkins Centre, Faculty of Medicine and Health, Sydney, NSW, AustraliaTransplant Unit, John Hunter Hospital, Newcastle, NSW, AustraliaTransplant and Surgical Immunology Theme, Immune Health Research Program, Hunter Medical Research Institute, The University of Newcastle, Newcastle, NSW, AustraliaDue to the intracellular expression of Foxp3 it is impossible to purify viable Foxp3+ cells on the basis of Foxp3 staining. Consequently CD4+Foxp3+ regulatory T cells (Tregs) in mice have mostly been characterized using CD4+CD25+ T cells or GFP-Foxp3 reporter T cells. However, these two populations cannot faithfully represent Tregs as the expression of CD25 and Foxp3 does not completely overlap and GFP+Foxp3+ reporter T cells have been reported to be functionally altered. The aim of this study was to characterize normal Tregs without separating Foxp3+ and Foxp3- cells for the expression of the main functional molecules and proliferation behaviors by flow cytometry and to examine their gene expression characteristics through differential gene expression. Our data showed that the expressions of Foxp3, CD25, CTLA-4 (both intracellular and cell surface) and PD-1 was mostly confined to CD4+ T cells and the expression of Foxp3 did not completely overlap with the expression of CD25, CTLA-4 or PD-1. Despite higher levels of expression of the T cell inhibitory molecules CTLA-4 and PD-1, Tregs maintained higher levels of Ki-67 expression in the homeostatic state and had greater proliferation in vivo after allo-activation than Tconv. Differential gene expression analysis revealed that resting Tregs exhibited immune activation markers characteristic of activated Tconv. This is consistent with the flow data that the T cell activation markers CD25, CTLA-4, PD-1, and Ki-67 were much more strongly expressed by Tregs than Tconv in the homeostatic state.https://www.frontiersin.org/articles/10.3389/fimmu.2024.1292158/fullregulatory T cellsFoxp3CTLA-4PD-1T cell proliferationdifferential gene expression |
| spellingShingle | Zilei Liu Zilei Liu Zilei Liu Katherine J. Baines Katherine J. Baines Natalie M. Niessen Munish K. Heer David Clark David Clark G. Alexander Bishop Paul R. Trevillian Paul R. Trevillian Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells Frontiers in Immunology regulatory T cells Foxp3 CTLA-4 PD-1 T cell proliferation differential gene expression |
| title | Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells |
| title_full | Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells |
| title_fullStr | Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells |
| title_full_unstemmed | Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells |
| title_short | Characterizing Foxp3+ and Foxp3- T cells in the homeostatic state and after allo-activation: resting CD4+Foxp3+ Tregs have molecular characteristics of activated T cells |
| title_sort | characterizing foxp3 and foxp3 t cells in the homeostatic state and after allo activation resting cd4 foxp3 tregs have molecular characteristics of activated t cells |
| topic | regulatory T cells Foxp3 CTLA-4 PD-1 T cell proliferation differential gene expression |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2024.1292158/full |
| work_keys_str_mv | AT zileiliu characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT zileiliu characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT zileiliu characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT katherinejbaines characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT katherinejbaines characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT nataliemniessen characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT munishkheer characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT davidclark characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT davidclark characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT galexanderbishop characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT paulrtrevillian characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells AT paulrtrevillian characterizingfoxp3andfoxp3tcellsinthehomeostaticstateandafteralloactivationrestingcd4foxp3tregshavemolecularcharacteristicsofactivatedtcells |