Skip to main content
NCBI home page
Brain Pathology logoLink to Brain Pathology
. 2006 Apr 5;10(3):330–341. doi: 10.1111/j.1750-3639.2000.tb00265.x

Differentiation and Histological Analysis of Embryonic Stem Cell‐Derived Neural Transplants in Mice

Yves Benninger 1, Silvia Marino 2, Roland Hardegger 1, Charles Weissmann 3, Adriano Aguzzi 1, Sebastian Brandner 1,✉
PMCID: PMC8098556  PMID: 10885652

Abstract

We report here that neural transplantation of in vitro‐differentiated embryonic stem (ES) cells provides a versatile strategy for gene transfer into the central nervous system. ES cells were subjected to an optimized in vitro differentiation protocol to obtain embryoid bodies. These aggregates were stereotaxically transplanted into the brain of recipient adult mice, where they followed a strictly controlled differentiation pattern and eventually formed mature neural grafts. A marker gene, introduced into the ROSA26 locus allowed for precise determination of the fate of the descendants of the transplanted embryoid bodies and revealed that not only neurons but also astrocytes, oligodendrocytes and even microglial cells were graft‐derived. Evaluation of long‐term experiments showed viable grafts with a stable transgene expression and proved that this approach provides a tool for reliable gene expression within a spatially delimited area of neural tissue.

Full Text

The Full Text of this article is available as a PDF (187.7 KB).

References

  • 1. Axelrod HR (1984) Embryonic stem cell lines derived from blastocysts by a simplified technique. Dev Biol 101: 225–228. [DOI] [PubMed] [Google Scholar]
  • 2. Bain G, Kitchens D, Yao M, Huettner JE, Gottlieb DI (1995) Embryonic stem cells express neuronal properties in vitro. Dev Biol 168: 342–357. [DOI] [PubMed] [Google Scholar]
  • 3. Bain G, Ray WJ, Yao M, Gottlieb DI (1996) Retinoic acid promotes neural and represses mesodermal gene expression in mouse embryonic stem cells in culture. Biochem Biophys Res Commun 223: 691–694. [DOI] [PubMed] [Google Scholar]
  • 4. Bjornson CR, Rietze RL, Reynolds BA, Magli MC, Vescovi AL (1999) Turning brain into blood: a hematopoietic fate adopted by adult neural stem cells in vivo. Science 283: 534–537. [DOI] [PubMed] [Google Scholar]
  • 5. Bottenstein JE (1985) Cell Culture ion Neurosciences In: Bottenstein JE, Harvey, A.L. (ed) Cell Culture ion Neurosciences. Plenum Press, New York , London , pp 3. [Google Scholar]
  • 6. Brewer GJ, Torricelli JR, Evege EK, Price PJ (1993) Optimized survival of hippocampal neurons in B27‐supplemented Neurobasal, a new serum‐free medium combination. J Neurosci Res 35: 567–576. [DOI] [PubMed] [Google Scholar]
  • 7. Brüstle O (1999) Building brains: neural chimeras in the study of nervous system development and repair. Brain Pathol 9: 527–545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8. Brüstle O, Jones KN, Learish RD, Karram K, Choudhary K, Wiestler OD, Duncan ID, McKay RD (1999) Embryonic stem cell‐derived glial precursors: A source of myelinating transplants. Science 285: 754–6. [DOI] [PubMed] [Google Scholar]
  • 9. Brüstle O, Maskos U, McKay RD (1995) Host‐guided migration allows targeted introduction of neurons into the embryonic brain. Neuron 15: 1275–1285. [DOI] [PubMed] [Google Scholar]
  • 10. Brüstle O, McKay RD (1996) Neuronal progenitors as tools for cell replacement in the nervous system. Curr Opin Neurobiol 6: 688–695. [DOI] [PubMed] [Google Scholar]
  • 11. Brüstle O, Spiro AC, Karram K, Choudhary K, Okabe S, McKay RD (1997) In vitro‐generated neural precursors participate in mammalian brain development. Proc Natl Acad Sci USA 94: 14809–14814. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12. Castro‐Obregon S, Covarrubias L (1996) Role of retinoic acid and oxidative stress in embryonic stem cell death and neuronal differentiation. FEBS Lett 381: 93–97. [DOI] [PubMed] [Google Scholar]
  • 13. Deacon T, Dinsmore J, Costantini LC, Ratliff J, Isacson O (1998) Blastula‐stage stem cells can differentiate into dopaminergic and serotonergic neurons after transplantation. Exp Neurol 149: 28–41. 9454612 [Google Scholar]
  • 14. Dinsmore J, Ratliff J, Deacon T, Pakzaban P, Jacoby D, Galpern W, Isacson O (1996) Embryonic stem cells differentiated in vitro as a novel source of cells for transplantation. Cell Transplant 5: 131–143. [DOI] [PubMed] [Google Scholar]
  • 15. Friedrich G, Soriano P (1991) Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice. Genes Dev 5: 1513–1523. [DOI] [PubMed] [Google Scholar]
  • 16. Hardy RJ, Friedrich VL, Jr. (1996) Oligodendrocyte progenitors are generated throughout the embryonic mouse brain, but differentiate in restricted foci. Development 122: 2059–2069. [DOI] [PubMed] [Google Scholar]
  • 17. Isenmann S, Brandner S, Kuhne G, Boner J, Aguzzi A (1996) Comparative in vivo and pathological analysis of the blood‐brain barrier in mouse telencephalic transplants. Neuropathol Appl Neurobiol 22: 118–128. [PubMed] [Google Scholar]
  • 18. Isenmann S, Brandner S, Sure U, Aguzzi A (1996) Telencephalic transplants in mice: characterization of growth and differentiation patterns. Neuropathol Appl Neurobiol 22: 108–117. [PubMed] [Google Scholar]
  • 19. Kalcheim C, Barde YA, Thoenen H, Le Douarin NM (1987) In vivo effect of brain‐derived neurotrophic factor on the survival of developing dorsal root ganglion cells. Embo J 6: 2871–2873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20. Koutouzis TK, Emerich DF, Borlongan CV, Freeman TB, Cahill DW, Sanberg PR (1994) Cell transplantation for central nervous system disorders. Crit Rev Neurobiol 8: 125–162. [PubMed] [Google Scholar]
  • 21. Lachyankar MB, Condon PJ, Quesenberry PJ, Litofsky NS, Recht LD, Ross AH (1997) Embryonic precursor cells that express Trk receptors: induction of different cell fates by NGF, BDNF, NT‐3, and CNTF. Exp Neurol 144: 350–360. [DOI] [PubMed] [Google Scholar]
  • 22. Lindvall O, Bjorklund A (1989) Transplantation strategies in the treatment of Parkinson's disease: experimental basis and clinical trials. Acta Neurol Scand Suppl 126: 197–210. [DOI] [PubMed] [Google Scholar]
  • 23. Marino S, Kretschmer C, Brandner S, Cavard C, Zider A, Briand P, Isenmann S, Wagner EF, Aguzzi A (1995) Activation of HIV transcription by human foamy virus in transgenic mice. Lab Invest 73: 103–110. [PubMed] [Google Scholar]
  • 24. Martinez Serrano A, Fischer W, Soderstrom S, Ebendal T, Bjorklund A (1996) Long‐term functional recovery from age‐induced spatial memory impairments by nerve growth factor gene transfer to the rat basal forebrain. Proc Natl Acad Sci USA 93: 6355–6360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25. McDonald JW, Liu XZ, Qu Y, Liu S, Mickey SK, Turetsky D, Gottlieb DI, Choi DW (1999) Transplanted embryonic stem cells survive, differentiate and promote recovery in injured rat spinal cord. Nat Med 5: 1410–1412. [DOI] [PubMed] [Google Scholar]
  • 26. Milner RJ, Lai C, Nave KA, Lenoir D, Ogata J, Sutcliffe JG (1985) Nucleotide sequences of two mRNAs for rat brain myelin proteolipid protein. Cell 42: 931–939. [DOI] [PubMed] [Google Scholar]
  • 27. Paulin D, Jakob H, Jacob F, Weber K, Osborn M (1982) In vitro differentiation of mouse teratocarcinoma cells monitored by intermediate filament expression. Differentiation 22: 90–99. [DOI] [PubMed] [Google Scholar]
  • 28. Ridet JL, Privat A (1995) Gene therapy in the central nervous system: direct versus indirect gene delivery. J Neurosci Res 42: 287–293. [DOI] [PubMed] [Google Scholar]
  • 29. Robertson EJ (1987) Teratocarcinomas and embryonic stem cells ‐ a practical approach In: D R and BD H (eds) Practical approach series. IRL Press, Oxford , vol 1. [Google Scholar]
  • 30. Rohrer DC, Nilaver G, Nipper V, Machida CA (1996) Genetically modified PC12 brain grafts: survivability and inducible nerve growth factor expression. Cell Transplant 5: 57–68. [DOI] [PubMed] [Google Scholar]
  • 31. Rosenstein JM (1995) Why do neural transplants survive? An examination of some metabolic and pathophysiological considerations in neural transplantation. Exp Neurol 133: 1–6. [DOI] [PubMed] [Google Scholar]
  • 32. Sabate O, Horellou P, Vigne E, Colin P, Perricaudet M, Buc Caron MH, Mallet J (1995) Transplantation to the rat brain of human neural progenitors that were genetically modified using adenoviruses. Nat Genet 9: 256–260. [DOI] [PubMed] [Google Scholar]
  • 33. Soriano P (1999) Generalized lacZ expression with the ROSA26 Cre reporter strain [letter]. Nat Genet 21: 70–71. [DOI] [PubMed] [Google Scholar]
  • 34. Spector DL, D. GR. L.A. L (1998) Culture and biochemical analysis of cells. Cold Spring Harbour Laboratory Press. [Google Scholar]
  • 35. Strubing C, Ahnert‐Hilger G, Shan J, Wiedenmann B, Hescheler J, Wobus AM (1995) Differentiation of pluripotent embryonic stem cells into the neuronal lineage in vitro gives rise to mature inhibitory and excitatory neurons. Mech Dev 53: 275–287. [DOI] [PubMed] [Google Scholar]
  • 36. Takahashi J, Palmer TD, Gage FH (1999) Retinoic acid and neurotrophins collaborate to regulate neurogenesis in adult‐derived neural stem cell cultures. J Neurobiol 38: 65–81. [PubMed] [Google Scholar]
  • 37. Tsuda M, Yuasa S, Fujino Y, Sekikawa M, Ono K, Tsuchiya T, Kawamura K (1990) Retrovirus‐mediated gene transfer into mouse cerebellar primary culture and its application to the neural transplantation. Brain Res Bull 24: 787–792. [DOI] [PubMed] [Google Scholar]
  • 38. Van Esseveldt KE, Liu R, Hermens WT, Verhaagen J, Boer GJ (1997) Adenoviral vector‐mediated gene transfer and neurotransplantation: possibilities and limitations in grafting of the fetal rat suprachiasmatic nucleus. J Neurosci Methods 71: 113–23. [DOI] [PubMed] [Google Scholar]
  • 39. Wobus AM, Grosse R, Schoneich J (1988) Specific effects of nerve growth factor on the differentiation pattern of mouse embryonic stem cells in vitro. Biomed Biochim Acta 47: 965–973. [PubMed] [Google Scholar]
  • 40. Yao M, Bain G, Gottlieb DI (1995) Neuronal differentiation of P19 embryonal carcinoma cells in defined media. J Neurosci Res 41: 792–804. [DOI] [PubMed] [Google Scholar]
  • 41. Zambrowicz BP, Imamoto A, Fiering S, Herzenberg LA, Kerr WG, Soriano P (1997) Disruption of overlapping transcripts in the ROSA beta geo 26 gene trap strain leads to widespread expression of beta‐galactosidase in mouse embryos and hematopoietic cells. Proc Natl Acad Sci USA 94: 3789–3794. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Brain Pathology are provided here courtesy of Wiley

RESOURCES