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Cell Migration pp 195-211 | Cite as

Using the Zebrafish Embryo to Dissect the Early Steps of the Metastasis Cascade

  • Gautier Follain
  • Naël Osmani
  • Cédric Fuchs
  • Guillaume Allio
  • Sébastien Harlepp
  • Jacky G. Goetz
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1749)

Abstract

Most cancers end up with the death of patients caused by the formation of secondary tumors, called metastases. However, how these secondary tumors appear and develop is only poorly understood. A fine understanding of the multiple steps of the metastasis cascade requires in vivo models allowing high spatiotemporal analysis of the behavior of metastatic cells. Zebrafish embryos combine several advantages such as transparency, small size, stereotyped anatomy, and easy handling, making it a very powerful model for cell and cancer biology, and in vivo imaging analysis. In the following chapter, we describe a complete procedure allowing in vivo imaging methods, at high throughput and spatiotemporal resolution, to assess the behavior of circulating tumor cells (CTCs) in an experimental metastasis assay. This protocol provides access, for the first time, to the earliest steps of tumor cell seeding during metastasis formation.

Key words

Zebrafish Circulating tumor cells (CTCs) Metastasis Injection Live imaging 

Notes

Acknowledgments

We thank all members of the Goetz Lab for helpful discussions throughout the development of this technology. We are grateful to Sofia AZEVEDO and Nina FEKONJA for their help in various aspects of this method. We are very much grateful to Francesca PERI (EMBL) and Kerstin RICHTER (EMBL) for providing zebrafish embryos. This work has been funded by Plan Cancer (OptoMetaTrap, to J.G. and S.H) and CNRS IMAG’IN (to S.H., J.G., and C.P.) and by institutional funds from INSERM and University of Strasbourg. G.F. is supported by La Ligue Contre le Cancer. N.O is supported by Plan Cancer. G.A. was supported by FRM (Fondation pour la Recherche Médicale).

Supplementary material

429754_1_En_15_MOESM1_ESM.zip (0 kb)
Supp_matlab_script (ZIP 58 kb)

References

  1. 1.
    Massagué J, Obenauf AC (2016) Metastatic colonization by circulating tumour cells. Nature 529:298–306CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Katt ME, Placone AL, Wong AD, Xu ZS, Searson PC (2016) In vitro tumor models: advantages, disadvantages, variables, and selecting the right platform. Front Bioeng Biotechnol 4:12CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Cheung KJ, Gabrielson E, Werb Z, Ewald AJ (2013) Collective invasion in breast cancer requires a conserved basal epithelial program. Cell 155:1639–1651CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Karreman MA, Hyenne V, Schwab Y, Goetz JG (2016) Intravital correlative microscopy: imaging life at the nanoscale. Trends Cell Biol 26:848–863CrossRefPubMedGoogle Scholar
  5. 5.
    Kienast Y et al (2010) Real-time imaging reveals the single steps of brain metastasis formation. Nat Med 16:116–122CrossRefPubMedGoogle Scholar
  6. 6.
    Cheung KJ et al (2016) Polyclonal breast cancer metastases arise from collective dissemination of keratin 14-expressing tumor cell clusters. Proc Natl Acad Sci U S A 113:E854–E863CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Cheon D-J, Orsulic S (2011) Mouse models of cancer. Annu Rev Pathol 6:95–119CrossRefPubMedGoogle Scholar
  8. 8.
    Deryugina EI, Kiosses WB (2017) Intratumoral cancer cell intravasation can occur independent of invasion into the adjacent stroma. Cell Rep 19:601–616CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Leong HS et al (2014) Invadopodia are required for cancer cell extravasation and are a therapeutic target for metastasis. Cell Rep 8:1558–1570CrossRefPubMedGoogle Scholar
  10. 10.
    Amatruda JF, Shepard JL, Stern HM, Zon LI (2002) Zebrafish as a cancer model system. Cancer Cell 1:229–231CrossRefPubMedGoogle Scholar
  11. 11.
    Stoletov K, Klemke R (2008) Catch of the day: zebrafish as a human cancer model. Oncogene 27:4509–4520CrossRefPubMedGoogle Scholar
  12. 12.
    Stoletov K, Montel V, Lester RD, Gonias SL, Klemke R (2007) High-resolution imaging of the dynamic tumor cell–vascular interface in transparent zebrafish. Proc Natl Acad Sci U S A 104:17406–17411CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Stoletov K et al (2010) Visualizing extravasation dynamics of metastatic tumor cells. J Cell Sci 123:2332–2341CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    White RM et al (2008) Transparent adult zebrafish as a tool for in vivo transplantation analysis. Cell Stem Cell 2:183–189CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Heilmann S et al (2015) A quantitative system for studying metastasis using transparent zebrafish. Cancer Res 75:4272–4282CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Kaufman CK et al (2016) A zebrafish melanoma model reveals emergence of neural crest identity during melanoma initiation. Science 351:aad2197CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Tang Q et al (2016) Imaging tumour cell heterogeneity following cell transplantation into optically clear immune-deficient zebrafish. Nat Commun 7:10358CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Kim IS et al (2017) Microenvironment-derived factors driving metastatic plasticity in melanoma. Nat Commun 8:14343CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Corey DR, Abrams JM (2001) Morpholino antisense oligonucleotides: tools for investigating vertebrate development. Genome Biol 2:reviews1015CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Zhang Y, Huang H, Zhang B, Lin S (2016) TALEN- and CRISPR-enhanced DNA homologous recombination for gene editing in zebrafish. Methods Cell Biol 135:107–120CrossRefPubMedGoogle Scholar
  21. 21.
    De Santis F, Di Donato V, Del Bene F (2016) Clonal analysis of gene loss of function and tissue-specific gene deletion in zebrafish via CRISPR/Cas9 technology. Methods Cell Biol 135:171–188CrossRefPubMedGoogle Scholar
  22. 22.
    Ablain J, Zon LI (2016) Tissue-specific gene targeting using CRISPR/Cas9. Methods Cell Biol 135:189–202CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Lawson ND, Weinstein BM (2002) In vivo imaging of embryonic vascular development using transgenic zebrafish. Dev Biol 248:307–318CrossRefPubMedGoogle Scholar
  24. 24.
    Follain G, Osmani N, Azevedo S, Allio G, Mercier L, Karreman MA, Solecki G, Garcia-Leon MJ, Lefebvre O, Fekonja N, Hille C, Chabannes V, Dollé G, Metivet T, Der Hovsepian F, Prudhomme C, Ruthensteiner B, Kemmling A, Siemonsen S, Schneider T, Fiehler J, Glatzel M, Winkler F, Schwab Y, Pantel K, Harlepp S, Goetz JG (2017) Hemodynamic forces tune the arrest, adhesion and extravasation of circulating tumor cells. Dev Cell. https://doi.org/10.1101/183046
  25. 25.
    Goetz JG et al (2014) Endothelial cilia mediate low flow sensing during zebrafish vascular development. Cell Rep 6:799–808CrossRefPubMedGoogle Scholar
  26. 26.
    Goetz JG, Monduc F, Schwab Y, Vermot J (2015) Using correlative light and electron microscopy to study zebrafish vascular morphogenesis. Methods Mol Biol 1189:31–46CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2018

Authors and Affiliations

  • Gautier Follain
    • 1
    • 2
    • 3
    • 4
  • Naël Osmani
    • 1
    • 2
    • 3
    • 4
  • Cédric Fuchs
    • 1
    • 2
    • 3
    • 4
  • Guillaume Allio
    • 1
    • 2
    • 3
    • 4
    • 5
  • Sébastien Harlepp
    • 2
    • 3
    • 4
    • 6
    • 7
  • Jacky G. Goetz
    • 1
    • 2
    • 3
    • 4
  1. 1.INSERM UMR_S1109, Tumor BiomechanicsInstitut d’Hématologie et d’ImmunologieStrasbourg CedexFrance
  2. 2.Université de StrasbourgStrasbourgFrance
  3. 3.LabEx MedalisUniversité de StrasbourgStrasbourgFrance
  4. 4.Fédération de Médecine Translationnelle de Strasbourg (FMTS)StrasbourgFrance
  5. 5.Centre de Biologie du DéveloppementUMR 5547 CNRS/Université Paul SabatierToulouseFrance
  6. 6.IPCMS, UMR7504StrasbourgFrance
  7. 7.LabEx NIEUniversité de StrasbourgStrasbourgFrance

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