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Visualizing regulatory T cell control of autoimmune responses in nonobese diabetic mice

Nature Immunology volume 7, pages 83–92 (2006)Cite this article

Abstract

The in vivo mechanism of regulatory T cell (Treg cell) function in controlling autoimmunity remains controversial. Here we have used two-photon laser-scanning microscopy to analyze lymph node priming of diabetogenic T cells and to delineate the mechanisms of Treg cell control of autoimmunity in vivo. Islet antigen–specific CD4+CD25− T helper cells (TH cells) and Treg cells swarmed and arrested in the presence of autoantigens. These TH cell activities were progressively inhibited in the presence of increasing numbers of Treg cells. There were no detectable stable associations between Treg and TH cells during active suppression. In contrast, Treg cells directly interacted with dendritic cells bearing islet antigen. Such persistent Treg cell–dendritic cell contacts preceded the inhibition of TH cell activation by dendritic cells, supporting the idea that dendritic cells are central to Treg cell function in vivo.

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Figure 1: Treg cells suppress the priming of islet antigen–specific CD4+CD25− TH cells in the pancreatic lymph node.
Figure 2: Movement dynamics of BDC2.5 CD4+CD25− TH cells in explanted lymph nodes.
Figure 3: Quantitative characterization of movement dynamics of BDC2.5 CD4+CD25− TH cells.
Figure 4: Treg cells alter the movement dynamics of BDC2.5 CD4+CD25− TH cells in explanted lymph nodes.
Figure 5: Autoreactive CD4+CD25− TH cells and Treg cells from BDC2.5 mice home to the T cell zone and preferentially accumulate at the T cell–B cell boundary in the presence of autoantigen.
Figure 6: In vivo suppression by Treg cells is not associated with stable Treg cell–TH cell interactions.
Figure 7: Islet antigen–bearing DCs form stable interaction with BDC2.5 CD4+CD25− TH cells.
Figure 8: BDC2.5 Treg cells stably interact with islet antigen–bearing DCs.

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Acknowledgements

We thank S. Jiang, C. McArthur, C. Bennett, R.A. Hwang, F. Siedenberg, L. Braun, J. Belgum, S. Hayden, M. Sanderson and M. Bengttson for technical assistance, and members of the Bluestone laboratory for support. Supported by the Juvenile Diabetes Research Foundation (4-1999-841 and 32004232), National Insitutes of Health (R37 AI46643, P30 DK063720, R21 AI066097 and AI30663), Sandler–Howard Hughes Medical Institute Biomedical Research Support Program (5300246), Sandler New Technologies and the Canadian Institutes of Health Research and Alberta Heritage Foundation for Medical Research.

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Author notes

  1. Matthew F Krummel and Jeffrey A Bluestone: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Medicine, University of California, San Francisco, Diabetes Center, University of California, San Francisco, 94143, California, USA

    Qizhi Tang, Jason Y Adams, Mingying Bi, Brian T Fife & Jeffrey A Bluestone

  2. Department of Pathology, University of California, San Francisco, 94143, California, USA

    Qizhi Tang, Aaron J Tooley, Matthew F Krummel & Jeffrey A Bluestone

  3. Howard Hughes Medical Institute and Department of Medicine, University of California, San Francisco, 94143, California, USA

    Richard M Locksley

  4. Julia McFarlane Diabetes Research Centre and Department of Microbiology & Infectious Diseases, University of Calgary, Calgary, T2N 4N1, Alberta, Canada

    Pau Serra & Pere Santamaria

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Supplementary information

Supplementary Fig. 1

TPLSM survey of pancreatic LN for the presence and distribution of transferred cells in the pancreatic LN. (PDF 372 kb)

Supplementary Video 1

BDC2.5 TH cells in inguinal LN of NOD recipients. The trajectories for some randomly selected cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 15 minute in real-time, which is compressed to 5 sec in this video. (MOV 409 kb)

Supplementary Video 2

BDC2.5 TH cells in pancreatic LN of NOD recipients. The trajectories for some free moving (top half) and swarming (lower middle) cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 15 minute in real-time, which is compressed to 5 sec in this video. (MOV 359 kb)

Supplementary Video 3

BDC2.5 TH cells in pancreatic LN of a NOD.Cd28−/− recipient. The trajectories for some clustering and arrested cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 30 minute in real-time, which is compressed to 10 sec in this video. (MOV 727 kb)

Supplementary Video 4

BDC2.5 TH cells in pancreatic LN of a NOD.CD80−/−CD86−/− recipient. The trajectories for some clustering and arrested cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 30 minute in real-time, which is compressed to 10 sec in this video. (MOV 1155 kb)

Supplementary Video 5

BDC2.5 TH cells in pancreatic LN of a NOD.Cd28−/− recipient reconstituted with NOD Treg cells. The trajectories for some swarming cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 30 minute in real-time, which is compressed to 10 seconds in this video. (MOV 997 kb)

Supplementary Video 6

BDC2.5 TH cells in pancreatic LN of a NOD.Cd28−/− recipient reconstituted with BDC2.5 Treg cells. The trajectories for some randomly selected cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 30 minute in real-time, which is compressed to 10 sec in this video. (MOV 1415 kb)

Supplementary Video 7

Lack of stable interaction between Treg cells and TH cells during in vivo suppression. BDC2.5 TH cells (greenish yellow) and BDC2.5 Treg cells (red) in pancreatic LN of a NOD.Cd28−/− recipient were imaged together showing lack of stable association of the two cell types. This duration of the imaging is 6 minute in real-time, which is compressed to 4 sec in this video. (MOV 414 kb)

Supplementary Video 8

BDC2.5 TH cells (red) interacting with GFP+ DCs in the pancreatic LN of a NOD.MIP.GFP recipient. This video is generated by projecting 10 images spanning 20 μm in the z direction onto a single plane. The duration of this imaging is 15 minute in real-time, which is compressed to 5 sec in this video. (MOV 180 kb)

Supplementary Video 9

BDC2.5 Treg cells in pancreatic LN of NOD recipients. The trajectories for some swarming cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 30 minute in real-time, which is compressed to 10 sec in this video. (MOV 1107 kb)

Supplementary Video 10

BDC2.5 Treg cells in pancreatic LN of a NOD.Cd28−/− recipient. The trajectories for some clustering cells are shown. The tracks are color coded to indicate time progression from the beginning (blue) to the end of imaging (yellow). The duration of the imaging is 15 minute in real-time, which is compressed to 4.5 sec in this video. (MOV 736 kb)

Supplementary Video 11

BDC2.5 Treg cells (red) interacting with a GFP+ DC in the pancreatic LN of a NOD.MIP.GFP recipient. This video is generated by projecting 10 images spanning 24 μm in the z direction onto a single plane. The duration of the imaging is 10 minute in real-time, which is compressed to 3 sec in this video. (MOV 195 kb)

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Tang, Q., Adams, J., Tooley, A. et al. Visualizing regulatory T cell control of autoimmune responses in nonobese diabetic mice. Nat Immunol 7, 83–92 (2006). https://doi.org/10.1038/ni1289

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