Skip to main content
SpringerLink
Log in

Zebrafish Xenografts for the In Vivo Analysis of Healthy and Malignant Human Hematopoietic Cells

  • Protocol
  • First Online:
Stem Cell Mobilization

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2017))

Abstract

The zebrafish is a powerful vertebrate model for genetic studies on embryonic development and organogenesis. In the last decades, zebrafish were furthermore increasingly used for disease modeling and investigation of cancer biology. Zebrafish are particularly used for mutagenesis and small molecule screens, as well as for live imaging assays that provide unique opportunities to monitor cell behavior, both on a single cell and whole organism level in real time. Zebrafish have been also used for in vivo investigations of human cells transplanted into embryos or adult animals; this zebrafish xenograft model can be considered as an intermediate assay between in vitro techniques and more time-consuming and expensive mammalian models.

Here, we present a protocol for transplantation of healthy and malignant human hematopoietic cells into larval zebrafish; transplantation into adult zebrafish and possible advantages and limitations of the zebrafish compared to murine xenograft models are discussed.

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 199.99
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. Mullins MC, Nüsslein-Volhard C (1993) Mutational approaches to studying embryonic pattern formation in the zebrafish. Curr Opin Genet Dev 3:648–654

    Article  CAS  Google Scholar 

  2. Granato M, Nüsslein-Volhard C (1996) Fishing for genes controlling development. Curr Opin Genet Dev 6:461–468

    Article  CAS  Google Scholar 

  3. Kari G, Rodeck U, Dicker AP (2007) Zebrafish: an emerging model system for human disease and drug discovery. Clin Pharmacol Ther 82:70–80

    Article  CAS  Google Scholar 

  4. Feitsma H, Cuppen E (2008) Zebrafish as a cancer model. Mol Cancer Res 6:685–694

    Article  CAS  Google Scholar 

  5. Amatruda JF, Patton EE (2008) Genetic models of cancer in zebrafish. Int Rev Cell Mol Biol 271:1–34

    Article  CAS  Google Scholar 

  6. den Hertog J (2005) Chemical genetics: drug screens in zebrafish. Biosci Rep 25:289–297

    Article  CAS  Google Scholar 

  7. Lam SH, Chua HL, Gong Z, Lam TJ, Sin YM (2004) Development and maturation of the immune system in zebrafish, Danio rerio: a gene expression profiling, in situ hybridization and immunological study. Dev Comp Immunol 28:9–28

    Article  CAS  Google Scholar 

  8. Krauss J, Astrinides P, Frohnhöfer HG, Walderich B, Nüsslein-Volhard C (2013) transparent, a gene affecting stripe formation in Zebrafish, encodes the mitochondrial protein Mpv17 that is required for iridophore survival. Biol Open 2:703–710

    Article  Google Scholar 

  9. White RM, Sessa A, Burke C, Bowman T, LeBlanc J, Ceol C, Bourque C, Dovey M, Goessling W, Burns CE, Zon LI (2008) Transparent adult zebrafish as a tool for in vivo transplantation analysis. Cell Stem Cell 2:183–189

    Article  CAS  Google Scholar 

  10. Theocharides AP, Rongvaux A, Fritsch K, Flavell RA, Manz MG (2016) Humanized hemato-lymphoid system mice. Haematologica 101:5–19

    Article  CAS  Google Scholar 

  11. Paczulla AM, Dirnhofer S, Konantz M, Medinger M, Salih HR, Rothfelder K, Tsakiris DA, Passweg JR, Lundberg P, Lengerke C (2017) Long-term observation reveals high-frequency engraftment of human acute myeloid leukemia in immunodeficient mice. Haematologica 102:854–864

    Article  CAS  Google Scholar 

  12. Konantz M, Balci TB, Hartwig UF, Dellaire G, André MC, Berman JN, Lengerke C (2012) Zebrafish xenografts as a tool for in vivo studies on human cancer. Ann N Y Acad Sci 1266:124–137

    Article  Google Scholar 

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

    Article  Google Scholar 

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

    Article  CAS  Google Scholar 

  15. Pruvot B, Jacquel A, Droin N, Auberger P, Bouscary D, Tamburini J, Muller M, Fontenay M, Chluba J, Solary E (2011) Leukemic cell xenograft in zebrafish embryo for investigating drug efficacy. Haematologica 96:612–616

    Article  CAS  Google Scholar 

  16. von Mässenhausen A, Sanders C, Brägelmann J, Konantz M, Queisser A, Vogel W, Kristiansen G, Duensing S, Schröck A, Bootz F, Brossart P, Kirfel J, Lengerke C, Perner S (2016) Targeting DDR2 in head and neck squamous cell carcinoma with dasatinib. Int J Cancer 139:2359–2369

    Article  Google Scholar 

  17. Queisser A, Hagedorn S, Wang H, Schaefer T, Konantz M, Alavi S, Deng M, Vogel W, von Mässenhausen A, Kristiansen G, Duensing S, Kirfel J, Lengerke C, Perner S (2017) Ecotropic viral integration site 1, a novel oncogene in prostate cancer. Oncogene 36:1573–1584

    Article  CAS  Google Scholar 

  18. Schaefer T, Wang H, Mir P, Konantz M, Pereboom TC, Paczulla AM, Merz B, Fehm T, Perner S, Rothfuss OC, Kanz L, Schulze-Osthoff K, Lengerke C (2015) Molecular and functional interactions between AKT and SOX2 in breast carcinoma. Oncotarget 6:43540–43556

    Article  Google Scholar 

  19. Wang H, Schaefer T, Konantz M, Braun M, Varga Z, Paczulla AM, Reich S, Jacob F, Perner S, Moch H, Fehm TN, Kanz L, Schulze-Osthoff K, Lengerke C (2017) Prominent oncogenic roles of EVI1 in breast carcinoma. Cancer Res 77:2148–2160

    Article  CAS  Google Scholar 

  20. He S, Lamers GE, Beenakker JW, Cui C, Ghotra VP, Danen EH, Meijer AH, Spaink HP, Snaar-Jagalska BE (2012) Neutrophil-mediated experimental metastasis is enhanced by VEGFR inhibition in a zebrafish xenograft model. J Pathol 227:431–445

    Article  CAS  Google Scholar 

  21. Nicoli S, Presta M (2007) The zebrafish/tumor xenograft angiogenesis assay. Nat Protoc 2:2918–2923

    Article  CAS  Google Scholar 

  22. Lee SL, Rouhi P, Dahl Jensen L, Zhang D, Ji H, Hauptmann G, Ingham P, Cao Y (2009) Hypoxia-induced pathological angiogenesis mediates tumor cell dissemination, invasion, and metastasis in a zebrafish tumor model. Proc Natl Acad Sci U S A 106:19485–19490

    Article  CAS  Google Scholar 

  23. Zhao C, Wang X, Zhao Y, Li Z, Lin S, Wei Y, Yang Y (2011) A novel xenograft model in zebrafish for high-resolution investigating dynamics of neovascularization in tumors. PLoS One 6:e21768

    Article  CAS  Google Scholar 

  24. Stoletov K, Kato H, Zardouzian E, Kelber J, Yang J, Shattil S, Klemke R (2010) Visualizing extravasation dynamics of metastatic tumor cells. J Cell Sci 123:2332–2341

    Article  CAS  Google Scholar 

  25. Jacob F, Alam S, Konantz M, Liang CY, Kohler RS, Everest-Dass AV, Huang YL, Rimmer N, Fedier A, Schötzau A, Núñez López M, Packer N, Lengerke C, Heinzelmann-Schwarz V (2018) Transition of mesenchymal and epithelial cancer cells depends on α1-4 galactosyltransferase-mediated glycosphingolipids. Cancer Res 78(11):2952–2965

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  27. Bentley VL, Veinotte CJ, Corkery DP, Pinder JB, LeBlanc MA, Bedard K, Weng AP, Berman JN, Dellaire G (2015) Focused chemical genomics using zebrafish xenotransplantation as a pre-clinical therapeutic platform for T-cell acute lymphoblastic leukemia. Haematologica 100:70–76

    Article  CAS  Google Scholar 

  28. Liu Y, Asnani A, Zou L, Bentley VL, Yu M, Wang Y, Dellaire G, Sarkar KS, Dai M, Chen HH, Sosnovik DE, Shin JT, Haber DA, Berman JN, Chao W, Peterson RT (2014) Visnagin protects against doxorubicin-induced cardiomyopathy through modulation of mitochondrial malate dehydrogenase. Sci Transl Med 6:266ra170

    Article  Google Scholar 

  29. Zhang B, Shimada Y, Hirota T, Ariyoshi M, Kuroyanagi J, Nishimura Y, Tanaka T (2016) Novel immunologic tolerance of human cancer cell xenotransplants in zebrafish. Transl Res 170:89–98.e83

    Article  CAS  Google Scholar 

  30. Mizgirev IV, Revskoy S (2010) A new zebrafish model for experimental leukemia therapy. Cancer Biol Ther 9:895–902

    Article  CAS  Google Scholar 

  31. Staal FJ, Spaink HP, Fibbe WE (2016) Visualizing human hematopoietic stem cell trafficking in vivo using a zebrafish xenograft model. Stem Cells Dev 25:360–365

    Article  CAS  Google Scholar 

  32. Hess I, Iwanami N, Schorpp M, Boehm T (2013) Zebrafish model for allogeneic hematopoietic cell transplantation not requiring preconditioning. Proc Natl Acad Sci U S A 110:4327–4332

    Article  CAS  Google Scholar 

  33. Shayegi N, Alakel N, Middeke JM, Schetelig J, Mantovani-Löffler L, Bornhäuser M (2015) Allogeneic stem cell transplantation for the treatment of refractory scleromyxedema. Transl Res 165:321–324

    Article  Google Scholar 

  34. Moore JC, Tang Q, Yordán NT, Moore FE, Garcia EG, Lobbardi R, Ramakrishnan A, Marvin DL, Anselmo A, Sadreyev RI, Langenau DM (2016) Single-cell imaging of normal and malignant cell engraftment into optically clear prkdc-null SCID zebrafish. J Exp Med 213:2575–2589

    Article  CAS  Google Scholar 

  35. Iwanami N, Hess I, Schorpp M, Boehm T (2017) Studying the adaptive immune system in zebrafish by transplantation of hematopoietic precursor cells. Methods Cell Biol 138:151–161

    Article  CAS  Google Scholar 

  36. de Jong JL, Burns CE, Chen AT, Pugach E, Mayhall EA, Smith AC, Feldman HA, Zhou Y, Zon LI (2011) Characterization of immune-matched hematopoietic transplantation in zebrafish. Blood 117:4234–4242

    Article  Google Scholar 

  37. Langenau DM, Ferrando AA, Traver D, Kutok JL, Hezel JP, Kanki JP, Zon LI, Look AT, Trede NS (2004) In vivo tracking of T cell development, ablation, and engraftment in transgenic zebrafish. Proc Natl Acad Sci U S A 101:7369–7374

    Article  CAS  Google Scholar 

  38. Tenente IM, Tang Q, Moore JC, Langenau DM (2014) Normal and malignant muscle cell transplantation into immune compromised adult zebrafish. J Vis Exp. https://doi.org/10.3791/52597

  39. Tang Q, Moore JC, Ignatius MS, Tenente IM, Hayes MN, Garcia EG, Torres Yordán N, Bourque C, He S, Blackburn JS, Look AT, Houvras Y, Langenau DM (2016) Imaging tumour cell heterogeneity following cell transplantation into optically clear immune-deficient zebrafish. Nat Commun 7:10358

    Article  CAS  Google Scholar 

  40. Stachura DL, Svoboda O, Campbell CA, Espín-Palazón R, Lau RP, Zon LI, Bartunek P, Traver D (2013) The zebrafish granulocyte colony-stimulating factors (Gcsfs): 2 paralogous cytokines and their roles in hematopoietic development and maintenance. Blood 122:3918–3928

    Article  CAS  Google Scholar 

  41. Svoboda O, Stachura DL, Machoňová O, Pajer P, Brynda J, Zon LI, Traver D, Bartůněk P (2014) Dissection of vertebrate hematopoiesis using zebrafish thrombopoietin. Blood 124:220–228

    Article  CAS  Google Scholar 

  42. Svoboda O, Stachura DL, Machoňová O, Zon LI, Traver D, Bartůněk P (2016) Ex vivo tools for the clonal analysis of zebrafish hematopoiesis. Nat Protoc 11:1007–1020

    Article  CAS  Google Scholar 

  43. Santos MD, Yasuike M, Hirono I, Aoki T (2006) The granulocyte colony-stimulating factors (CSF3s) of fish and chicken. Immunogenetics 58:422–432

    Article  CAS  Google Scholar 

  44. Wehmas LC, Tanguay RL, Punnoose A, Greenwood JA (2016) Developing a novel embryo-larval zebrafish xenograft assay to prioritize human glioblastoma therapeutics. Zebrafish 13:317–329

    Article  CAS  Google Scholar 

  45. Harfouche R, Basu S, Soni S, Hentschel DM, Mashelkar RA, Sengupta S (2009) Nanoparticle-mediated targeting of phosphatidylinositol-3-kinase signaling inhibits angiogenesis. Angiogenesis 12:325–338

    Article  CAS  Google Scholar 

  46. Cheng J, Gu YJ, Wang Y, Cheng SH, Wong WT (2011) Nanotherapeutics in angiogenesis: synthesis and in vivo assessment of drug efficacy and biocompatibility in zebrafish embryos. Int J Nanomedicine 6:2007–2021

    Article  CAS  Google Scholar 

  47. Ellett F, Pase L, Hayman JW, Andrianopoulos A, Lieschke GJ (2011) mpeg1 promoter transgenes direct macrophage-lineage expression in zebrafish. Blood 117:e49–e56

    Article  CAS  Google Scholar 

  48. Choi J, Dong L, Ahn J, Dao D, Hammerschmidt M, Chen JN (2007) FoxH1 negatively modulates flk1 gene expression and vascular formation in zebrafish. Dev Biol 304:735–744

    Article  CAS  Google Scholar 

  49. Progatzky F, Dallman MJ, Lo Celso C (2013) From seeing to believing: labelling strategies for in vivo cell-tracking experiments. Interface Focus 3:20130001

    Article  Google Scholar 

Download references

Acknowledgments

We thank Prof. Dr. Sven Perner for the help with histopathological analyses of tumorlike structures. This work was funded by grants from the Swiss National Science Foundation (164200 and 149735).

Author information

Authors and Affiliations

  1. Department of Biomedicine, University of Basel and University Hospital Basel, Basel, Switzerland

    Martina Konantz, Joëlle S. Müller & Claudia Lengerke

Authors
  1. Martina Konantz
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Joëlle S. Müller
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Claudia Lengerke
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Claudia Lengerke .

Editor information

Editors and Affiliations

  1. Department of Internal Medicine II, Center for Medical Research, University of Tübingen, Tübingen, Germany

    Gerd Klein

  2. Institute of Transfusion Medicine and Immunology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany

    Patrick Wuchter

Rights and permissions

Reprints and permissions

Copyright information

© 2019 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Konantz, M., Müller, J.S., Lengerke, C. (2019). Zebrafish Xenografts for the In Vivo Analysis of Healthy and Malignant Human Hematopoietic Cells. In: Klein, G., Wuchter, P. (eds) Stem Cell Mobilization. Methods in Molecular Biology, vol 2017. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9574-5_16

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-9574-5_16

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9573-8

  • Online ISBN: 978-1-4939-9574-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation