Clinical & Experimental Metastasis

, Volume 29, Issue 4, pp 371–380 | Cite as

Distinct patterns of human medulloblastoma dissemination in the developing chick embryo nervous system

  • Tene A. Cage
  • Jonathan D. Louie
  • Sharon R. Liu
  • Arturo Alvarez-Buylla
  • Nalin Gupta
  • Jeanette Hyer
Technical Note


Medulloblastoma (MB) is the most common malignant primary brain tumor in children. Aggressive tumors that disseminate along the leptomeninges carry extremely poor prognoses. Mechanisms that predict dissemination are poorly understood. Our objective was to develop a reliable and reproducible model to study MB dissemination. We have created a chicken-human xenograft to study features of MB with leptomeningeal dissemination. Human MB cell lines (D283, Daoy), primary human MB cells (SF8113), and primary genetic mouse model (Math1cre:SmoM2 flox/flox) MB cells were either transfected to express green fluorescent protein (GFP) or were labeled with a membrane permeable green fluorescent probe. Cells were then injected as aggregates or implanted as pellets into the developing chicken brain immediately after neural tube closure at embryonic day 2 (E2). Most embryos were harvested three days after implantation (E5) though some were harvested up to E15. The developing brain was analyzed via whole mount fluorescent imaging and tissue section immunohistochemistry. Human and mouse MBs survived in the developing chicken central nervous system (CNS). They exhibited distinct patterns of incorporation and dissemination into the CNS that were consistent with observed phenotypes of the corresponding human patient or mouse host. Specifically, metastatic D283 cells disseminated along the leptomeninges whereas Daoy, primary mouse MB, and primary human MB cells did not. This work supports an avian-human xenograft as a successful model to study patterns of MB dissemination. Our model provides a basis for manipulating cell signaling mechanisms to understand critical targets involved in MB dissemination.


Brain Tumor Chicken embryo Dissemination Medulloblastoma Pediatric Xenograft 



Grant support: This study was primarily supported by a research grant from the Sami Disharoon Brain Tumor Research Foundation and Katie’s Kids for the Cure and form. Additional support from: American Brain Tumor Foundation Medical Student Summer Fellowship (TAC); Genentech Foundation Fellowship (TAC); Howard Hughes Medical Institute Medical Fellows Fellowship (TAC); NIH/NINDS K08 NS05506 (NG). Materials: David Rowitch, M.D., Ph.D. and Vivi Heine, Ph.D. for providing Math1cre:SmoM2 flox/flox mice; William Weiss, Ph.D. and Fredrik Johansson Swartling, Ph.D. for providing SF8113 cells; The Neurological Surgery Tissue Bank at University of California, San Francisco for providing information on the medulloblastoma tumor sample.


  1. 1.
    Louis DN, Ohgaki H, Wiestler OD, Cavenee W (2007) Classification of Tumors of the Central Nervous System, 4th edn. WHO, GenevaGoogle Scholar
  2. 2.
    Sutton LN, Phillips PC, Molloy PT (1996) Surgical management of medulloblastoma. J Neurooncol 29(1):9–21PubMedCrossRefGoogle Scholar
  3. 3.
    Cretu A, Fotos JS, Little BW, Galileo DS (2005) Human and rat glioma growth, invasion, and vascularization in a novel chick embryo brain tumor model. Clin Exp Metastasis 22(3):225–236PubMedCrossRefGoogle Scholar
  4. 4.
    Fontaine-Perus J, Cheraud Y (2005) Mouse-chick neural chimeras. Int J Dev Biol 49(2):349–353PubMedCrossRefGoogle Scholar
  5. 5.
    Mitsiadis TA, Cheraud Y, Sharpe P, Fontaine-Perus J (2003) Development of teeth in chick embryos after mouse neural crest transplantations. Proc Natl Acad Sci USA 100(11):6541–6545PubMedCrossRefGoogle Scholar
  6. 6.
    Hatton BA, Knoepfler PS, Kenney AM, Rowitch DH, de Alboran IM, Olson JM, Eisenman RN (2006) N-myc is an essential downstream effector of Shh signaling during both normal and neoplastic cerebellar growth. Cancer Res 66(17):8655–8661PubMedCrossRefGoogle Scholar
  7. 7.
    Kenney AM, Cole MD, Rowitch DH (2003) Nmyc upregulation by sonic hedgehog signaling promotes proliferation in developing cerebellar granule neuron precursors. Development 130(1):15–28PubMedCrossRefGoogle Scholar
  8. 8.
    Lelievre V, Seksenyan A, Nobuta H, Yong WH, Chhith S, Niewiadomski P, Cohen JR, Dong H, Flores A, Liau LM, Kornblum HI, Scott MP, Waschek JA (2008) Disruption of the PACAP gene promotes medulloblastoma in ptc1 mutant mice. Dev Biol 313(1):359–370PubMedCrossRefGoogle Scholar
  9. 9.
    Zurawel RH, Allen C, Chiappa S, Cato W, Biegel J, Cogen P, de Sauvage F, Raffel C (2000) Analysis of PTCH/SMO/SHH pathway genes in medulloblastoma. Genes Chrom Cancer 27(1):44–51PubMedCrossRefGoogle Scholar
  10. 10.
    Raffel C (2004) Medulloblastoma: molecular genetics and animal models. Neoplasia 6:310–322PubMedCrossRefGoogle Scholar
  11. 11.
    Ellison D (2002) Classifying the medulloblastoma: insights from morphology and molecular genetics. Neuropathol Appl Neurobiol 28(4):257–282PubMedCrossRefGoogle Scholar
  12. 12.
    Ramaswamy V, Northcott PA, Taylor MD (2011) FISH and chips: the recipe for improved prognostication and outcomes for children with medulloblastoma. Cancer genetics 204:577–588PubMedCrossRefGoogle Scholar
  13. 13.
    McManamy CS, Pears J, Weston CL, Hanzely Z, Ironside JW, Taylor RE, Grundy RG, Clifford SC, Ellison DW (2007) Nodule formation and desmoplasia in medulloblastomas-defining the nodular/desmoplastic variant and its biological behavior. Brain Pathol 17(2):151–164PubMedCrossRefGoogle Scholar
  14. 14.
    Crawford JR, MacDonald TJ, Packer RJ (2007) Medulloblastoma in childhood: new biological advances. Lancet Neurol 6(12):1073–1085PubMedCrossRefGoogle Scholar
  15. 15.
    Scheurlen WG, Schwabe GC, Joos S, Mollenhauer J, Sorensen N, Kuhl J (1998) Molecular analysis of childhood primitive neuroectodermal tumors defines markers associated with poor outcome. J Clin Oncol 16(7):2478–2485PubMedGoogle Scholar
  16. 16.
    Gajjar A, Hernan R, Kocak M, Fuller C, Lee Y, McKinnon PJ, Wallace D, Lau C, Chintagumpala M, Ashley DM, Kellie SJ, Kun L, Gilbertson RJ (2004) Clinical, histopathologic, and molecular markers of prognosis: toward a new disease risk stratification system for medulloblastoma. J Clin Oncol 22(6):984–993PubMedCrossRefGoogle Scholar
  17. 17.
    Hernan R, Fasheh R, Calabrese C, Frank AJ, Maclean KH, Allard D, Barraclough R, Gilbertson RJ (2003) ERBB2 up-regulates S100A4 and several other prometastatic genes in medulloblastoma. Cancer Res 63(1):140–148PubMedGoogle Scholar
  18. 18.
    MacDonald TJ, Brown KM, LaFleur B, Peterson K, Lawlor C, Chen Y, Packer RJ, Cogen P, Stephan DA (2001) Expression profiling of medulloblastoma: PDGFRA and the RAS/MAPK pathway as therapeutic targets for metastatic disease. Nat Genet 29(2):143–152PubMedCrossRefGoogle Scholar
  19. 19.
    Stearns D, Chaudhry A, Abel TW, Burger PC, Dang CV, Eberhart CG (2006) c-myc overexpression causes anaplasia in medulloblastoma. Cancer Res 66(2):673–681PubMedCrossRefGoogle Scholar
  20. 20.
    Hatton BA, Villavicencio EH, Tsuchiya KD, Pritchard JI, Ditzler S, Pullar B, Hansen S, Knoblaugh SE, Lee D, Eberhart CG, Hallahan AR, Olson JM (2008) The Smo/Smo model: hedgehog-induced medulloblastoma with 90% incidence and leptomeningeal spread. Cancer Res 68(6):1768–1776PubMedCrossRefGoogle Scholar
  21. 21.
    Li Y, Guessous F, Johnson EB, Eberhart CG, Li XN, Shu Q, Fan S, Lal B, Laterra J, Schiff D, Abounader R (2008) Functional and molecular interactions between the HGF/c-Met pathway and c-Myc in large-cell medulloblastoma. Lab Invest 88(2):98–111PubMedCrossRefGoogle Scholar
  22. 22.
    Schuller U, Heine VM, Mao J, Kho AT, Dillon AK, Han YG, Huillard E, Sun T, Ligon AH, Qian Y, Ma Q, Alvarez-Buylla A, McMahon AP, Rowitch DH, Ligon KL (2008) Acquisition of granule neuron precursor identity is a critical determinant of progenitor cell competence to form Shh-induced medulloblastoma. Cancer Cell 14(2):123–134PubMedCrossRefGoogle Scholar
  23. 23.
    Friedman HS, Burger PC, Bigner SH, Trojanowski JQ, Wikstrand CJ, Halperin EC, Bigner DD (1985) Establishment and characterization of the human medulloblastoma cell line and transplantable xenograft D283 Med. J Neuropathol Exp Neurol 44(6):592–605PubMedCrossRefGoogle Scholar
  24. 24.
    Jacobsen PF, Jenkyn DJ, Papadimitriou JM (1985) Establishment of a human medulloblastoma cell line and its heterotransplantation into nude mice. J Neuropathol Exp Neurol 44(5):472–485PubMedCrossRefGoogle Scholar
  25. 25.
    Ponten J, Macintyre EH (1968) Long term culture of normal and neoplastic human glia. Acta pathol microbiol Scand 74(4):465–486PubMedCrossRefGoogle Scholar
  26. 26.
    Kuhlman J, Niswander L (1997) Limb deformity proteins: role in mesodermal induction of the apical ectodermal ridge. Development 124(1):133–139PubMedGoogle Scholar
  27. 27.
    Dahmane N, Sanchez P, Gitton Y, Palma V, Sun T, Beyna M, Weiner H, Ruiz i Altaba A (2001) The Sonic Hedgehog-Gli pathway regulates dorsal brain growth and tumorigenesis. Development 128(24):5201–5212PubMedGoogle Scholar
  28. 28.
    Enguita-German M, Schiapparelli P, Rey JA, Castresana JS (2010) CD133 + cells from medulloblastoma and PNET cell lines are more resistant to cyclopamine inhibition of the sonic hedgehog signaling pathway than CD133-cells. Tumour Biol 31(5):381–390PubMedCrossRefGoogle Scholar
  29. 29.
    Shu Q, Antalffy B, Su JM, Adesina A, Ou CN, Pietsch T, Blaney SM, Lau CC, Li XN (2006) Valproic Acid prolongs survival time of severe combined immunodeficient mice bearing intracerebellar orthotopic medulloblastoma xenografts. Clin Cancer Res 12(15):4687–4694PubMedCrossRefGoogle Scholar
  30. 30.
    Yang WQ, Senger D, Muzik H, Shi ZQ, Johnson D, Brasher PM, Rewcastle NB, Hamilton M, Rutka J, Wolff J, Wetmore C, Curran T, Lee PW, Forsyth PA (2003) Reovirus prolongs survival and reduces the frequency of spinal and leptomeningeal metastases from medulloblastoma. Cancer Res 63(12):3162–3172PubMedGoogle Scholar
  31. 31.
    Cretu A, Brooks PC (2007) Impact of the non-cellular tumor microenvironment on metastasis: Potential therapeutic and imaging opportunities. J Cell Physiol 213:391–402PubMedCrossRefGoogle Scholar
  32. 32.
    Kulesa PM, Kasemeier-Kulesa JC, Teddy JM, Margaryan NV, Seftor EA, Seftor RE, Hendrix MJ (2006) Reprogramming metastatic melanoma cells to assume a neural crest cell-like phenotype in an embryonic microenvironment. Proc Natl Acad Sci USA 103(10):3752–3757PubMedCrossRefGoogle Scholar
  33. 33.
    Shu Q, Wong KK, Su JM, Adesina AM, Yu LT, Tsang YT, Antalffy BC, Baxter P, Perlaky L, Yang J, Dauser RC, Chintagumpala M, Blaney SM, Lau CC, Li XN (2008) Direct orthotopic transplantation of fresh surgical specimen preserves CD133 + tumor cells in clinically relevant mouse models of medulloblastoma and glioma. Stem cells 26(6):1414–1424PubMedCrossRefGoogle Scholar
  34. 34.
    Awatramani R, Soriano P, Rodriguez C, Mai JJ, Dymecki SM (2003) Cryptic boundaries in roof plate and choroid plexus identified by intersectional gene activation. Nat Genet 35(1):70–75PubMedCrossRefGoogle Scholar
  35. 35.
    Jung W, Castren E, Odenthal M (1994) Vande Woude GF, Ishii T, Dienes HP, Lindholm D, Schirmacher P. Expression and functional interaction of hepatocyte growth factor-scatter factor and its receptor c-met in mammalian brain. J Cell Biol 126(2):485–494PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Tene A. Cage
    • 1
  • Jonathan D. Louie
    • 2
  • Sharon R. Liu
    • 1
  • Arturo Alvarez-Buylla
    • 1
    • 3
  • Nalin Gupta
    • 1
    • 4
  • Jeanette Hyer
    • 1
  1. 1.Department of Neurological SurgeryUniversity of California, San FranciscoSan FranciscoUSA
  2. 2.Faculty of ScienceUniversity of CaliforniaBerkeleyUSA
  3. 3.Department of Neurological Surgery, Broad Center of Regeneration Medicine and Stem Cell ResearchUniversity of California, San FranciscoSan FranciscoUSA
  4. 4.Department of Neurological Surgery, Brain Tumor Research CenterUniversity of California, San FranciscoSan FranciscoUSA

Personalised recommendations