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In vivo tracking of 'color-coded' effector, natural and induced regulatory T cells in the allograft response

Nature Medicine volume 16, pages 718–722 (2010)Cite this article

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Abstract

Here we present methods to longitudinally track islet allograft–infiltrating T cells in live mice by endoscopic confocal microscopy and to analyze circulating T cells by in vivo flow cytometry. We developed a new reporter mouse whose T cell subsets express distinct, 'color-coded' proteins enabling in vivo detection and identification of effector T cells (Teff cells) and discrimination between natural and induced regulatory T cells (nTreg and iTreg cells). Using these tools, we observed marked differences in the T cell response in recipients receiving tolerance-inducing therapy (CD154-specific monoclonal antibody plus rapamycin) compared to untreated controls. These results establish real-time cell tracking as a powerful means to probe the dynamic cellular interplay mediating immunologic rejection or transplant tolerance.

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Figure 1: In vivo imaging of color-coded T cells.
Figure 2: Analysis of infiltrating T cells within islet allografts.
Figure 3: Serial endomicroscopy of infiltrating T cells within islet allografts.
Figure 4: Detection of nTreg, Teff and iTreg cells by in vivo flow cytometry in the peripheral blood.

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Acknowledgements

This work was supported by grants from the US National Institutes of Health (R21 AI081010 to S.H.Y. and R01 EY14106 to C.P.L. and P01-AI041521 and P01-AI041521-S1 to T.B.S.) and the Juvenile Diabetes Research Foundation (JDRF 7-2005-1329 to T.B.S. and M.K.).

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Author notes

  1. Zhigang Fan and Joel A Spencer: These authors contributed equally to this work.

Authors and Affiliations

  1. Transplant Institute, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, USA

    Zhigang Fan, Yan Lu, Gurbakhshish Singh, Vasilis Toxavidis, Terry B Strom & Maria Koulmanda

  2. Advanced Microscopy Program, Center for Systems Biology and Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA

    Joel A Spencer, Costas M Pitsillides & Charles P Lin

  3. Department of Biomedical Engineering, Science and Technology Center, Tufts University, Medford, Massachusetts, USA

    Joel A Spencer

  4. Department of Biomedical Engineering, Boston University, Boston, Massachusetts, USA

    Costas M Pitsillides

  5. Department of Dermatology, Advanced Microscopy Program, Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA

    Pilhan Kim & Seok H Yun

Authors
  1. Zhigang Fan
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  9. Terry B Strom
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Contributions

Z.F. and J.A.S. designed the experiments, conducted research, collected and analyzed data and wrote the manuscript; Y.L., C.M.P., G.S. and V.T. helped conduct research and collected and analyzed data; P.K. and S.H.Y. developed and performed endoscopic microscopy; T.B.S., C.P.L. and M.K. designed the experiments, sponsored the project and wrote the manuscript.

Corresponding authors

Correspondence to Terry B Strom or Charles P Lin.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 and Supplementary Table 1 (PDF 3048 kb)

Supplementary Video 1

Z-stack reconstructed movie showing yellow iTreg (green + red), green nTreg and red Teff cells (red) in a tolerized islet allograft at week 2. Z axis step size is 2 ÎĽm. (AVI 10288 kb)

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Fan, Z., Spencer, J., Lu, Y. et al. In vivo tracking of 'color-coded' effector, natural and induced regulatory T cells in the allograft response. Nat Med 16, 718–722 (2010). https://doi.org/10.1038/nm.2155

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