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Intravital Imaging of Cytotoxic T Lymphocytes

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Cytotoxic T-Cells

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1186))

Abstract

Intravital imaging approaches are proving to be essential to address new questions to better understand how the immune system operates. These approaches are especially valuable to characterize the complex organization of immune responses in vivo. Here, we examine how to take advantage of the cornea as a natural body window to apply noninvasive imaging techniques to assess cytotoxic T lymphocyte involvement in the immune rejection process in a model of intraocular allogeneic islet transplantation.

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References

  1. Monks CR, Freiberg BA, Kupfer H, Sciaky N, Kupfer A (1998) Three-dimensional segregation of supramolecular activation clusters in T cells. Nature 395(6697):82–86. doi:10.1038/25764

    Article  CAS  PubMed  Google Scholar 

  2. Cahalan MD, Parker I (2008) Choreography of cell motility and interaction dynamics imaged by two-photon microscopy in lymphoid organs. Annu Rev Immunol 26:585–626. doi:10.1146/annurev.immunol.24.021605.090620

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  3. Henrickson SE, Mempel TR, Mazo IB, Liu B, Artyomov MN, Zheng H, Peixoto A, Flynn M, Senman B, Junt T, Wong HC, Chakraborty AK, von Andrian UH (2008) In vivo imaging of T cell priming. Sci Signal 1(12):pt2. doi:10.1126/stke.112pt2

    Article  PubMed  Google Scholar 

  4. Gordon RE, Hanley PE, Shaw D, Gadian DG, Radda GK, Styles P, Bore PJ, Chan L (1980) Localization of metabolites in animals using 31P topical magnetic resonance. Nature 287 (5784):736–738

    Article  CAS  PubMed  Google Scholar 

  5. Kuhl DE, Phelps ME, Kowell AP, Metter EJ, Selin C, Winter J (1980) Effects of stroke on local cerebral metabolism and perfusion: mapping by emission computed tomography of 18FDG and 13NH3. Ann Neurol 8(1): 47–60. doi:10.1002/ana.410080108

    Article  CAS  PubMed  Google Scholar 

  6. Collaco AM, Geusz ME (2003) Monitoring immediate-early gene expression through firefly luciferase imaging of HRS/J hairless mice. BMC Physiol 3:8. doi:10.1186/1472-6793-3-8

    Article  PubMed Central  PubMed  Google Scholar 

  7. Ntziachristos V (2010) Going deeper than microscopy: the optical imaging frontier in biology. Nat Methods 7(8):603–614. doi: 10.1038/nmeth.1483

    Article  CAS  PubMed  Google Scholar 

  8. Denk W, Strickler JH, Webb WW (1990) Two-photon laser scanning fluorescence microscopy. Science 248(4951):73–76

    Article  CAS  PubMed  Google Scholar 

  9. Speier S, Nyqvist D, Cabrera O, Yu J, Molano RD, Pileggi A, Moede T, Kohler M, Wilbertz J, Leibiger B, Ricordi C, Leibiger IB, Caicedo A, Berggren PO (2008) Noninvasive in vivo imaging of pancreatic islet cell biology. Nat Med 14(5):574–578. doi:10.1038/nm1701

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  10. Nyqvist D, Speier S, Rodriguez-Diaz R, Molano RD, Lipovsek S, Rupnik M, Dicker A, Ilegems E, Zahr-Akrawi E, Molina J, Lopez-Cabeza M, Villate S, Abdulreda MH, Ricordi C, Caicedo A, Pileggi A, Berggren PO (2011) Donor islet endothelial cells in pancreatic islet revascularization. Diabetes 60(10):2571–2577. doi:10.2337/db10-1711

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  11. Abdulreda MH, Faleo G, Molano RD, Lopez-Cabezas M, Molina J, Tan Y, Echeverria OA, Zahr-Akrawi E, Rodriguez-Diaz R, Edlund PK, Leibiger I, Bayer AL, Perez V, Ricordi C, Caicedo A, Pileggi A, Berggren PO (2011) High-resolution, noninvasive longitudinal live imaging of immune responses. Proc Natl Acad Sci U S A 108(31):12863–12868. doi:10.1073/pnas.1105002108

  12. Pileggi A, Abdulreda M, Faleo G, Molano RD, Ricordi C, Caicedo A, Berggren PO (2012) The anterior chamber of the eye allows longitudinal, intravital imaging of CTLs to study islet immunobiology. J Clin Invest. doi:10.1172/JCI59285L1

    Google Scholar 

  13. Rodriguez-Diaz R, Speier S, Molano RD, Formoso A, Gans I, Abdulreda MH, Cabrera O, Molina J, Fachado A, Ricordi C, Leibiger I, Pileggi A, Berggren PO, Caicedo A (2012) Noninvasive in vivo model demonstrating the effects of autonomic innervation on pancreatic islet function. Proc Natl Acad Sci U S A 109 (52):21456–21461. doi:10.1073/pnas.1211659110

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  14. Rodriguez-Diaz R, Dando R, Huang YA, Berggren PO, Roper SD, Caicedo A (2012) Real-time detection of acetylcholine release from the human endocrine pancreas. Nat Protoc 7(6):1015–1023. doi:10.1038/nprot.2012.040

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  15. Abdulreda MH, Caicedo A, Berggren PO (2013) A natural body window to study human pancreatic islet cell function and survival. CellR4 1(2):111–122

    Google Scholar 

  16. Mojibian M, Harder B, Hurlburt A, Bruin JE, Asadi A, Kieffer TJ (2013) Implanted islets in the anterior chamber of the eye are prone to autoimmune attack in a mouse model of diabetes. Diabetologia 56(10):2213–2221. doi:10.1007/s00125-013-3004-z

    Article  CAS  PubMed  Google Scholar 

  17. Schmidt-Christensen A, Hansen L, Ilegems E, Fransen-Pettersson N, Dahl U, Gupta S, Larefalk A, Hannibal TD, Schulz A, Berggren PO, Holmberg D (2013) Imaging dynamics of CD11c cells and Foxp3 cells in progressive autoimmune insulitis in the NOD mouse model of type 1 diabetes. Diabetologia. doi:10.1007/s00125-013-3024-8

    PubMed  Google Scholar 

  18. Rodriguez-Diaz R, Caicedo A (2013) Novel approaches to studying the role of innervation in the biology of pancreatic islets. Endocrinol Metab Clin North Am 42(1):39–56. doi:10.1016/j.ecl.2012.11.001

  19. Unutmaz D, Xiang W, Sunshine MJ, Campbell J, Butcher E, Littman DR (2000) The primate lentiviral receptor Bonzo/STRL33 is coordinately regulated with CCR5 and its expression pattern is conserved between human and mouse. J Immunol 165(6):3284–3292

    Article  CAS  PubMed  Google Scholar 

  20. Pileggi A, Molano RD, Berney T, Ichii H, San Jose S, Zahr E, Poggioli R, Linetsky E, Ricordi C, Inverardi L (2005) Prolonged allogeneic islet graft survival by protoporphyrins. Cell Transplant 14(2–3):85–96

    Article  PubMed  Google Scholar 

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Acknowledgements

We thank the Diabetes Research Institute’s Cores (“Preclinical Cell Processing and Translational Models,” “Imaging,” “Flow Cytometry,” and “Histology Laboratory”) for the invaluable technical expertise provided.

Financial support for these studies was provided by the Diabetes Research Institute Foundation (to POB and AP); the National Institute of Health (NIH) through the National Institute of Digestive and Kidney Disease (5U19AI050864-10 to AP; U01DK089538 to POB and AP; 5U42 RR016603; 1R01EB008009 to AP; F32DK083226; R56DK084321, R01DK084321, and R03DK075487, the Juvenile Diabetes Research Foundation International (4-2004-361 to POB and AP; 4-2008-811 to AP; 17-2010-5 and 17-2012-361 to AP); the American Diabetes Association (7-13-IN-32, to AP); and the Larry L. Hillblom Foundation (to GF). Additional research support was provided through funds (to POB) from Karolinska Institutet, the Swedish Research Council, the Swedish Diabetes Foundation, the Family Erling-Persson Foundation, the Family Knut and Alice Wallenberg Foundation, the Skandia Insurance Company Ltd., VIBRANT (FP7-228933-2), Strategic Research Program in Diabetes at Karolinska Institutet, the NovoNordisk Foundation, the Stichting af Jochnick Foundation, and the Berth von Kantzow’s Foundation.

Disclosures: P-OB is co-founder and CEO of BioCrine AB who is using the anterior chamber of the eye in vivo imaging platform for commercial purposes.

Author information

Authors and Affiliations

  1. Transplantation Research Laboratory, Department of Surgery, University of California, 513 Parnassus Ave., HSE-520, San Francisco, CA, 94143, USA

    Gaetano Faleo Ph.D.

  2. Hospital Stockholm, Karolinska Institutet, Stockholm, Sweden

    Per-Olof Berggren Ph.D.

  3. Diabetes Research Institute, Miller School of Medicine, University of Miami, Miami, FL, 33136, USA

    Per-Olof Berggren Ph.D. & Antonello Pileggi M.D., Ph.D.

  4. Division of Cellular Transplantation, DeWitt-Daughtry Family Department of Surgery, Miller School of Medicine, University of Miami, Miami, FL, 33136, USA

    Antonello Pileggi M.D., Ph.D.

  5. Department of Microbiology and Immunology, Miller School of Medicine, University of Miami, Miami, FL, 33136, USA

    Antonello Pileggi M.D., Ph.D.

  6. Department of Biomedical Engineering, University of Miami, Miami, FL, USA

    Antonello Pileggi M.D., Ph.D.

Authors
  1. Gaetano Faleo Ph.D.
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  2. Per-Olof Berggren Ph.D.
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  3. Antonello Pileggi M.D., Ph.D.
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Corresponding author

Correspondence to Gaetano Faleo Ph.D. .

Editor information

Editors and Affiliations

  1. Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy

    Elena Ranieri

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Faleo, G., Berggren, PO., Pileggi, A. (2014). Intravital Imaging of Cytotoxic T Lymphocytes. In: Ranieri, E. (eds) Cytotoxic T-Cells. Methods in Molecular Biology, vol 1186. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1158-5_9

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  • DOI: https://doi.org/10.1007/978-1-4939-1158-5_9

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1157-8

  • Online ISBN: 978-1-4939-1158-5

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