Skip to main content
SpringerLink
Log in

Imaging of Human Cancer Cell Proliferation, Invasion, and Micrometastasis in a Zebrafish Xenogeneic Engraftment Model

  • Protocol
  • First Online:
Zebrafish

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1451))

Abstract

The xenograft model, using the early life stages of the zebrafish, allows imaging of tumor cell behavior both on a single cell and whole organism level, over time, within a week. This robust and reproducible assay can be used as an intermediate step between in vitro techniques and the expensive, and time consuming, murine models of cancer invasion and metastasis.

In this chapter, a detailed protocol to inject human cancer cells into the blood circulation of a zebrafish embryo is described; the engraftment procedure is then followed by visualization and quantification methods of tumor cell proliferation, invasion, and micrometastasis formation during subsequent larval development. Interaction with the host microenvironment is also considered.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 99.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 129.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 179.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. White R, Rose K, Zon L (2013) Zebrafish cancer: the state of the art and the path forward. Nat Rev Cancer 9:624–636

    Article  Google Scholar 

  2. Lieschke GJ, Trede NS (2009) Fish immunology. Curr Biol 19(16):R678–R682

    Article  CAS  PubMed  Google Scholar 

  3. Jagannathan-Bogdan M, Zon LI (2013) Hematopoiesis. Development 140(12):2463–2467

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Amatruda JF, Patton EE (2008) Genetic models of cancer in zebrafish. In: Jeon KW (ed) International review of cell and molecular biology, vol 271. Elsevier, Amsterdam, pp 1–34

    Google Scholar 

  5. He S, Lamers GE, Beenakker JW et al (2012) Neutrophil-mediated experimental metastasis is enhanced by VEGFR inhibition in a zebrafish xenograft model. J Pathol 227(4):431–445

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Feng Y, Santoriello C, Mione M et al (2010) Live imaging of innate immune cell sensing of transformed cells in zebrafish larvae: parallels between tumor initiation and wound inflammation. PLoS Biol. doi:10.1371/journal.pbio.1000562

    Google Scholar 

  7. Amatruda JF, Shepard JL, Stern HK et al (2002) Zebrafish as a cancer model system. Cancer Cell. doi:10.1016/S1535-6108(02)00052-1

    PubMed  Google Scholar 

  8. Zon LI, Peterson RT (2005) In vivo drug discovery in the zebrafish. Nat Rev Drug Discov 4(1):35–44

    Article  CAS  PubMed  Google Scholar 

  9. Veinotte CJ, Dellaire G, Berman JN (2014) Hooking the big one: the potential of zebrafish xenotransplantation to reform cancer drug screening in the genomic area. Dis Model Mech 7(7):745–754

    Article  PubMed  PubMed Central  Google Scholar 

  10. Murphy JB (1913) Transplantability of tissues to the embryo of foreign species. Its bearing on questions of tissue specificity and tumor immunity. J Exp Med 17:482–493

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Ribatti D (2014) The chick embryo chorioallantoic membrane as a model for tumor biology. Exp Cell Res 328(2):314–324

    Article  CAS  PubMed  Google Scholar 

  12. Rygaard J, Povlsen CO (1969) Heterotransplantation of a human malignant tumour to “Nude” mice. Acta Pathol Microbiol Scand 77:758–760

    Article  CAS  PubMed  Google Scholar 

  13. Cekanova M, Rathore K (2014) Animal models and therapeutic molecular targets of cancer: utility and limitations. Drug Des Devel Ther 8:1911–1922

    Article  PubMed  PubMed Central  Google Scholar 

  14. Haldi M, Ton C, Seng WL et al (2006) Human melanoma cells transplanted into zebrafish proliferate, migrate, produce melanin, form masses and stimulate angiogenesis in zebrafish. Angiogenesis 9(3):139–151

    Article  PubMed  Google Scholar 

  15. Corkey DP, Dellaire G, Berman JN (2011) Leukaemia xenotransplantation in zebrafish-chemotherapy response assay in vivo. Br J Haematol 153:786–789

    Article  Google Scholar 

  16. van der Ent W, Jochemsen AG, Teunisse AF et al (2014) Ewing sarcoma inhibition by disruption of EWSR1-FLI1 transcriptional activity and reactivation of p53. J Pathol 233(4):415–424

    Article  PubMed  Google Scholar 

  17. van der Ent W, Burrello C, Teunisse AF et al (2014) Modeling of human uveal melanoma in zebrafish xenograft embryos. Invest Ophthalmol Vis Sci 55(10):6612–6622

    Article  PubMed  Google Scholar 

  18. Nicoli S, Ribatti D, Cotelli F et al (2007) Mammalian tumor xenografts induce neovascularization in zebrafish embryos. Cancer Res 67(7):2927–2931

    Article  CAS  PubMed  Google Scholar 

  19. Zhao C, Wang X, Zhao Y et al (2011) A novel xenograft model in zebrafish for high resolution investigating dynamics of neovascularization in tumors. PLoS One 6:e21768

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Lee SL, Rouhi P, Dahl JL et al (2009) Hypoxia-induced pathological angiogenesis mediates tumor cell dissemination, invasion, and metastasis in a zebrafish tumor model. Proc Natl Acad Sci U S A 106(46):19485–19490

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Stoletov K, Kato H, Zardouzian E et al (2010) Visualizing extravasation dynamics of metastatic tumor cells. J Cell Sci 123(Pt 13):2332–2341

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Tobia C, Gariano G, De Sena G et al (2013) Zebrafish embryo as a tool to study tumor/endothelial cell cross-talk. Biochim Biophys Acta 1832(9):1371–1377

    Article  CAS  PubMed  Google Scholar 

  23. Drabsch Y, He S, Zhang L et al (2013) Transforming growth factor-β signalling controls human breast cancer metastasis in a zebrafish xenograft model. Breast Cancer Res 15(6):R106

    Article  PubMed  PubMed Central  Google Scholar 

  24. Wetterwald A, van der Pluijm G, Que I et al (2002) Optical imaging of cancer metastasis to bone marrow: a mouse model of minimal residual disease. Am J Pathol 160(3):1143–1153

    Article  PubMed  PubMed Central  Google Scholar 

  25. Pettaway CA, Pathak S, Greene G et al (1996) Selection of highly metastatic variants of different human prostatic carcinomas using orthotopic implantationin nude mice. Clin Cancer Res 2(9):1627–1636

    CAS  PubMed  Google Scholar 

  26. Wang Y, Kaiser MS, Larson JD et al (2010) Moesin1 and Ve-cadherin are required in endothelial cells during in vivo tubulogenesis. Development 137(18):3119–3128

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Jin SW, Beis D, Mitchell T et al (2005) Cellular and molecular analyses of vascular tube and lumen formation in zebrafish. Development 132:5199–5209

    Article  CAS  PubMed  Google Scholar 

  28. Lawson N, Weinstein B (2002) In vivo imaging of embryonic vascular development using transgenic zebrafish. Dev Biol 248:307–318

    Article  CAS  PubMed  Google Scholar 

  29. Renshaw SA, Loynes CA, Trushell DMI et al (2006) A transgenic zebrafish model of neutrophilic inflammation. Blood 108:3976–3978

    Article  CAS  PubMed  Google Scholar 

  30. Ellett F, Pase L, Hayman JW et al (2011) Mpeg1 promoter transgenes direct macrophage-lineage expression in zebrafish. Blood 117(4):e49–e56

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Bernut A, Herrmann JL, Kissa K et al (2014) Mycobacterium abscessus cording prevents phagocytosis and promotes abscess formation. Proc Natl Acad Sci U S A 111:E943–E952

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Cui C, Benard EL, Kanwal Z et al (2011) Infectious disease modeling and innate immune function in zebrafish embryos. Methods Cell Biol 105:273–308

    Article  CAS  PubMed  Google Scholar 

  33. Loynes CA, Martin JS, Robertson A et al (2010) Pivotal advance: pharmacological manipulation of inflammation resolution during spontaneously resolving tissue neutrophilia in the zebrafish. J Leukoc Biol 87:203–212

    Article  CAS  PubMed  Google Scholar 

  34. Ghotra VP, He S, de Bont H et al (2012) Automated whole animal bio-imaging assay for human cancer dissemination. PLoS One 7(2):e31281

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgments

The cell lines reported in this chapter A673 GFP, MDA-MB-231-B DsRed, and PC3 Pro4 were kindly provided by C. Schleitoff and U. Dirksten (University of Münster), P. ten Dijke (LUMC), and G. van der Pluijm (LUMC), respectively. We thank H. de Bont for assistance with the Nikon A1R confocal microscope and Image-Pro analysis support.

The work was supported by the Netherlands Organization for Scientific Research (TOP GO Grant: 854.10.012).

Author information

Authors and Affiliations

  1. Institute of Biology, Leiden University, Einsteinweg 55, 2333 CC, Leiden, The Netherlands

    Claudia Tulotta, Shuning He, Lanpeng Chen, Arwin Groenewoud, Wietske van der Ent, Annemarie H. Meijer, Herman P. Spaink & B. Ewa Snaar-Jagalska

Authors
  1. Claudia Tulotta
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Shuning He
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Lanpeng Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Arwin Groenewoud
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wietske van der Ent
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Annemarie H. Meijer
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Herman P. Spaink
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. B. Ewa Snaar-Jagalska
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to B. Ewa Snaar-Jagalska .

Editor information

Editors and Affiliations

  1. Division of Molecular and Developmental Biology, National Institute of Genetics, Mishima, Japan

    Koichi Kawakami

  2. Institute of Genetics and Molecular Medicine, MRC Human Genetics Unit & The University of Edinburgh Cancer Research UK Centre, Edinburgh, United Kingdom

    E. Elizabeth Patton

  3. Champalimaud Centre for the Unknown, Champalimaud Neuroscience Programme Champalimaud Centre for the Unknown, Lisbon, Portugal

    Michael Orger

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Tulotta, C. et al. (2016). Imaging of Human Cancer Cell Proliferation, Invasion, and Micrometastasis in a Zebrafish Xenogeneic Engraftment Model. In: Kawakami, K., Patton, E., Orger, M. (eds) Zebrafish. Methods in Molecular Biology, vol 1451. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3771-4_11

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3771-4_11

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3769-1

  • Online ISBN: 978-1-4939-3771-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation