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Large-scale expansion of mammalian neural stem cells: a review

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Abstract

A relatively new approach to the treatment of neurodegenerative diseases is the direct use of neural stem cells (NSCs) as therapeutic agents. The expected demand for treatment from the millions of afflicted individuals, coupled with the expected demand from biotechnology companies creating therapies, has fuelled the need to develop large-scale culture methods for these cells. The rapid pace of discovery in this area has been assisted through the use of animal model systems, enabling many experiments to be performed quickly and effectively. This review focuses on recent developments in expanding human and murine NSCs on a large scale, including the development of new serum-free media and bioreactor protocols. In particular, engineering studies that characterise important scale-up parameters are examined, including studies examining the effects of long-term culture of NSCs in suspension bioreactors. In addition, recent advances in the human NSC system are reviewed, including techniques for the evaluation of NSC characteristics.

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References

  • Alam, S. (2003): ‘Cell cycle kinetics of expanding populations of mammalian neural stem cells’. MSc thesis, University of Calgary, Calgary, Canada

    Google Scholar 

  • Auth, M. K. H., Okamoto, M., Ishida, Y., Keogh, A., Auth, S. H. G., Gerlach, J., Encke, A., McMaster, P., andStrain, A. J. (1998): ‘Maintained function of primary human hepatocytes by cellular interactions in coculture: implications for liver support systems’,Transpl. Int.,11, pp. S439-S443

    Article  Google Scholar 

  • Bjorklund, A., andGage, F. (2001): ‘Chimerical stem cells’,Trends Mol. Med.,7, pp. 144–145

    Google Scholar 

  • Bjorklund, L. M., Sanchez-Pernaute, R., Chung, S., Andersson, T., Chen, I. Y. C., McNaught, K. ST. P., Brownell, A. L., Jenkins, B. G., Wahlestedt, C., Kim, K. S., andIsacson, O. (2001): ‘Embryonic stem cells develop into functional dopaminergic neurons after transplantation in a Parkinson rat model’,Proc. Natl. Acad. Sci.,99, pp. 2344–2349

    Google Scholar 

  • Bjornson, C. R. R., Rietze, R. L., Reynolds, B. A., Magli, M. C., andVescovi, A. L. (1999): ‘Turning brain into blood: a hematopoietic fate adopted by adult neural stem cellsin vivo’,Science,283, pp. 534–537

    Article  Google Scholar 

  • Cao, Q., Benton, R. L., andWhittemore, S. R. (2002): ‘Stem cell repair of central nervous system injury’,J. Neurosci. Res.,68, pp. 501–510

    Article  Google Scholar 

  • Carpenter, M. K., Cui, X., Hu, Z., Jackson, J., Sherman, S., Seiger, A., andWahlberg, L. U. (1999): ‘In vitro expansion of a multipotent population of human neural progenitor cells’,Exp. Neurol.,158, pp. 265–278

    Article  Google Scholar 

  • Check, E. (2002): ‘Parkinson's patients show positive response to implants’,Nature,416, p. 666

    Google Scholar 

  • Cheshier, S. H., Morrison, S. J., Liao, X., andWeissman, I. L. (1999): ‘In vivo proliferation and cell cycle kinetics of long-term self-renewing hematopoietic stem cells’,Proc. Natl. Acad. Sci. USA,96, pp. 3120–3125

    Article  Google Scholar 

  • Chu, V. T., andGage, F. H. (2001): ‘Chipping away at stem cells’,Proc. Natl. Acad. Sci.,98, pp. 7652–7653

    Article  Google Scholar 

  • Darling, S. M., andAbbott, C. M. (1992): ‘Mouse models of human single gene disorders 1: non-transgenic mice’,BioEssays,14, pp. 359–366

    Article  Google Scholar 

  • Espinosa-Jeffrey, A., Becker-Catania, S. G., Zhao, P. M., Cole, R., Edmond, J., andDe Vellis, J. (2002): ‘Selective specification of CNS stem cells into oligodendroglial or neuronal cell lineage: cell culture and transplantation studies’,J. Neurosci. Res.,69, pp. 810–825

    Article  Google Scholar 

  • Flax, J. D., Aurora, S., Yang, C., Simonin, C., Wills, A. M., Billinghurst, L. L., Jendoubi, M., Sidman, R. L., Wolfe, S. U., andSnyder, E. Y. (1998): ‘Engraftable neural stem cells respond to developmental cues, replace neurons, and express foreign genes’,Nature Biotechnol.,16, pp. 1033–1039

    Google Scholar 

  • Forestell, S. P., Kalogerakis, N., Behie, L. A., andGerson, D. F. (1992a): ‘Development of the optimal inoculation conditions for microcarrier cultures’,Biotechnol. Bioeng.,39, pp. 305–313

    Article  Google Scholar 

  • Forestell, S. P., Milne, B. J., Kalogerakis, N., andBehie, L. A. (1992b): ‘A cellular automaton model for the growth of anchoragedependent mammalian cells used in vaccine production’,Chem. Eng. Sci.,47, pp. 2381–2386

    Google Scholar 

  • Freed, C. R., Greene, P. E., Breeze, R. E., Tsai, W. Y., Dumouchel, W., Kao, R., Dillon, S., Winfiled, H., Culver, S., Trojanowski, J. Q., Eidelberg, D., andFahn, S. (2001): ‘Transplantation of embryonic dopamine neurons for severe Parkinson's disease’,New Engl. J. Med.,344, pp. 710–719

    Article  Google Scholar 

  • Freyer, J. P., andSutherland, R. M. (1986): ‘Regulation of growth saturation and development of necrosis in EMT6/Ro multicellular spheroids by the glucose and oxygen supply’,Cancer Res.,46, pp. 3504–3512

    Google Scholar 

  • Fricker, R. A., Carpenter, M. K., Winkler, C., Greco, C., Gates, M. A., andBjorklund, A. (1999): ‘Site-specific migration and neuronal differentiation of human neural progenitor cells after transplantation in the adult rat brain’,J. Neurosci.,19, pp. 5990–6005

    Google Scholar 

  • Gage, F. H. (2000): ‘Mammalian neural stem cells’,Science,287, pp. 1433–1439

    Article  Google Scholar 

  • Galvin, K. A., andJones, D. G. (2002): ‘Adult human neural stem cells for cell-replacement therapies in the central nervous system’,Med. J. Aust.,177, pp. 316–318

    Google Scholar 

  • Gerlach, J. C. (1997): ‘Long-term liver cell cultures in bioreactors and possible application for liver support’,Cell Biol. Toxicol.,13, pp. 349–355

    Article  Google Scholar 

  • Gottlieb, D. I. (2002): ‘Large scale sources of neural stem cells’,Ann. Rev. Neurosci.,25, pp. 381–407

    Google Scholar 

  • Hu, W. S., andAunins, J. G. (1997): ‘Large-scale mammalian cell culture’,Curr. Opin. Biotechnol.,8, pp. 148–153

    Article  Google Scholar 

  • Ivanova, N. B., Dimos, J. T., Schaniel, C., Hackney, J. A., Moore, K. A., andLemischka, I. R. (2002): ‘A stem cell molecular signature’,Science,298, pp. 601–604

    Article  Google Scholar 

  • Kallos, M. S., Reynolds, B. A., Vescovi, A. L., andBehie, L. A. (1998): ‘High cell density growth of mammalian neural stem cells as aggregates in bioreactors’ inFan, L. S., andKnowlton, T. M. (Eds). ‘Fluidization IX’ (Engineering Foundation, New York, 1998), pp. 653–660

    Google Scholar 

  • Kallos, M. S., Vescovi, A. L., andBehie, L. A. (1999): ‘Extended serial passaging of mammalian neural stem cells in suspension bioreactors’,Biotechnol. Bioeng.,65, pp. 589–599

    Article  Google Scholar 

  • Kallos, M. S. (1999): ‘The development of bioreactor protocols for the large-scale expansion of mammalian neural stem cells in suspension bioreactors’. PhD thesis, University of Calgary, Calgary, Canada

    Google Scholar 

  • Kallos, M. S., andBehie, L. A. (1999): ‘Inoculation and growth conditions for high cell density expansion of mammalian neural stem cells’,Biotechnol. Bioeng.,63, pp. 473–483

    Article  Google Scholar 

  • Kordower, J. H., Chen, E. Y., Winkler, C., Fricker, R., Charles, V., Messing, A., Mufson, E. J., Wong, S. C., Rosenstein, J. R., Bjorklund, A., Emerich, D. F., Hammang, J., andCarpenter, M. K. (1997): ‘Grafts of EGF-responsive neural stem cells derived from GFAP-hNGF transgenic mice: trophic and tropic effects in a rodent model of Huntington's disease’,J. Compar: Neurol.,387, pp. 96–113

    Article  Google Scholar 

  • Lanza, R. P., Langer, R., andVacanti, J. (2000): ‘Principles of tissue engineering’, 2nd edn (Academic Press, San Diego, 2000)

    Google Scholar 

  • Leib, T. M., Pereira, C. J., andVilladsen, J. (2001): ‘Bioreactors: a chemical engineering perspective’,Chem. Eng. Sci.,56, pp. 5485–5497

    Google Scholar 

  • Lucchinetti, C. F., andRodriguez, M. (1997): ‘The controversy surrounding the pathogenesis of the multiple sclerosis lesion’,Mayo Clin. Proc.,72, pp. 665–678

    Google Scholar 

  • Majzoub, J. A., andMuglia, L. J. (1996): ‘Knockout mice’,New Engl. J. Med.,334, pp. 904–907

    Article  Google Scholar 

  • Madlambayan, G. J., Rogers, I., Casper, R. F., andZandstra, P. W. (2001): ‘Controlling culture dynamics for the expansion of hematopoietic stem cells’,J. Hematother. Stem Cell Res.,10, pp. 481–492

    Article  Google Scholar 

  • McKay, R. (1997): ‘Stem cells in the central nervous system’,Science,276, pp. 66–71

    Google Scholar 

  • Meisler, M. H. (1996): ‘The role of the laboratory mouse in the human genome project (review article)’,Am. J. Hum. Genet.,59, pp. 764–771

    Google Scholar 

  • Moreira, J. L., Santana, P. C., Feliciano, A. S., Cruz, P. E., Racher, A. J., Griffiths, J. B., andCarrondo, M. J. T. (1995a): ‘Effect of viscosity upon hydrodynamically controlled natural aggregates of animal cells grown in stirred vessels’,Biotechnol. Progr.,11, pp. 575–583

    Google Scholar 

  • Moreira, J. L., Alves, P. M., Aunins, J. G., andCarrondo, M. J. T. (1995b): ‘Hydrodynamic effects on BHK cells grown as suspended natural aggregates’,Biotechnol. Bioeng.,46, pp. 351–360

    Article  Google Scholar 

  • Morrison, S. J. (2001): ‘Stem cell potential: can anything make anything?’,Current Biol.,11, pp. R7-R9

    MathSciNet  Google Scholar 

  • Morshead, C. M., Reynolds, B. A., Craig, C. G., McBurney, M. W., Staines, W. M., Morassutti, D., Weiss, S., andvan der Kooy, D. (1994): ‘Neural stem cells in the adult mammalian forebrain: a relatively quiescent subpopulation of subependymal cells’,Neuron,13, pp. 1071–1082

    Article  Google Scholar 

  • Morshead, C. M., Benveniste, P., Iscove, N. N., andvan der Kooy, D. (2002): ‘Hematopoietic competence is a rare property of neural stem cells that may depend on genetic and epigenetic alterations’,Nature Med.,8, pp. 268–273

    Google Scholar 

  • Mouse Genome Sequencing Consortium (2002): ‘Initial sequencing and comparative analysis of the mouse genome’,Nature,420, pp. 520–562

    Google Scholar 

  • Nikkhah, G., Olsson, M., Eberhard, J., Bentlage, C., Cunningham, M. G., andBjorklund, A. (1994): ‘A microtransplantation approach for cell suspension grafting in the rat Parkinson model: a detailed account of the methodology’,Neuroscience,63, pp. 57–72

    Article  Google Scholar 

  • Park, K. I., Liu, S., Flax, J. D., Nissim, S., Stieg, P. E., andSnyder, E. Y. (1999): ‘Transplantation of neural progenitor and stem cells: developmental insights may suggest new therapies for spinal cord and other CNS dysfunction’,J. Neurotrauma,16, pp. 675–687

    Google Scholar 

  • Pellegrini, G., Bondanza, S., Guerra, L., andDe Luca, M. (1998): ‘Cultivation of human keratinocyte stem cells: current and future clinical applications’,Medical and Biological Engineering and Computing,36, pp. 778–790

    Google Scholar 

  • Reynolds, B. A., andWeiss, S. (1992): ‘Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system’,Science,255, pp. 1707–1710

    Google Scholar 

  • Reynolds, B. A., andWeiss, S. (1996): ‘Clonal and population analysis demonstrate that an EGF-responsive mammalian embryonic CNS precursor is a stem cell’,Dev. Biol.,175, pp. 1–13

    Article  Google Scholar 

  • Runge, D., Runge, D. M., Jager, D., Lubecki, K. A., Stoltz, D. B., Karathanasis, S., Kietzmann, T., Strom, S. C., Jungermann, K., Fleig, W. E., andMichalopoulos, G. K. (2000): ‘Serum-free, long-term cultures of human hepatocytes: maintenance of cell morphology, transcription factors, and liver-specific functions’,Biochem. Biophys. Res. Commun.,269, pp. 46–53

    Article  Google Scholar 

  • Ryder, E. F., Snyder, E. Y., andCepko, C. L. (1990): ‘Establishment and characterization of multipotent neural cell lines using retrovirus vector-mediated oncogene transfer’,J. Neurobiol.,21, pp. 356–375

    Article  Google Scholar 

  • Sen, A. (1998): ‘The development of media for thein vitro expansion of mammalian neural stem cells’. MSc thesis, University of Calgary, Calgary, Canada

    Google Scholar 

  • Sen, A., andBehie, L. A. (1999): ‘The development of a medium for thein vitro expansion of mammalian neural stem cells’,Can. J. Chem. Eng.,77, pp. 963–972

    Google Scholar 

  • Sen, A., Kallos, M. S., andBehie, L. A. (2001): ‘Hydrodynamic effects on extended cultures of mammalian neural stem cell aggregates in suspension culture’,Ind. Eng. Chem. Res.,40, pp. 5350–5357

    Article  Google Scholar 

  • Sen, A., Kallos, M. S., andBehie, L. A. (2002a): ‘Passaging protocols for mammalian neural stem cells in suspension bioreactors’,Biotechnol. Progr.,18, pp. 337–345

    Google Scholar 

  • Sen, A., Kallos, M. S., andBehie, L. A. (2002b): ‘Expansion of mammalian neural stem cells in bioreactors: effect of power input and medium viscosity’,Dev. Brain Res.,134, pp. 103–113

    Article  Google Scholar 

  • Shingo, T., Sorokan, S. T., Shimazaki, T., andWeiss, S. (2001): ‘Erythropoietin regulates thein vitro andin vivo production of neural progenitors by mammalian forebrain neural stem cells’,J. Neurosci.,21, pp. 9733–9743

    Google Scholar 

  • Snyder, E. Y., Deitcher, D. L., Walsh, C., Arnold-Aldea, S., Hartwieg, E. A., andCepko, C. L. (1992): ‘Multipotent neural cell lines can engraft and participate in development of mouse cerebellum’,Cell,68, pp. 33–51

    Article  Google Scholar 

  • Snyder, E. Y., Park, K. I., Flax, J. D., Liu, S., Rosario, C. M., Yandava, B. D., andAurora, S. (1997): ‘Potential of neural “stem-like” cells for gene therapy and repair of the degenerating central nervous system’,Adv. Neurol.,72, pp. 121–132

    Google Scholar 

  • Suslov, O. N., Kukekov, V. G., Ignatova, T. N., andSteindler, D. A. (2002): ‘Neural stem cell heterogeneity demonstrated by molecular phenotyping of clonal neurospheres’,Proc. Natl. Acad. Sci. USA,99, pp. 14506–14511

    Article  Google Scholar 

  • Svendsen, C. N., Skepper, J., Rosser, A. E., Ter Borg, M. G., Tyres, P., andRyken, T. (1997): ‘Restricted growth potential of rat neural precursors as compared to mouse’,Dev. Brain Res.,99, pp. 253–258

    Article  Google Scholar 

  • Svendsen, C. N., Ter Borg, M. G., Armstrong, R. J. E., Rosser, A. E., Chandran, S., Ostenfeld, T., andCaldwell, M. A. (1998): ‘A new method for the rapid and long term growth of human neural precursor cells’,J. Neurosci. Meth.,85, pp. 141–152

    Google Scholar 

  • Svendsen, C. N., Caldwell, M. A., andOstenfeld, T. (1999): ‘Human neural stem cells: isolation, expansion and transplantation’,Brain Pathol.,9, pp. 499–513

    Google Scholar 

  • Svendsen, C. N., andSmith, A. G. (1999): ‘New prospects for human stem-cell therapy in the nervous system’,Trends Neurosci.,22, pp. 357–364

    Article  Google Scholar 

  • Svendsen, C. N., andCaldwell, M. A. (2000): ‘Neural stem cells in the developing central nervous system: implications for cell therapy and transplantation’,Prog. Brain Res.,127, pp. 13–34

    Google Scholar 

  • Tamaki, S., Eckert, K., He, D., Sutton, R., Doshe, M., Jain, G., Tushinski, R., Reitsma, M., Harris, B., Tsukamoto, A., Gage, F., Weissman, I., andUchida, N. (2002): ‘Engraftment of sorted/expanded human central nervous system stem cells from fetal brain’,J. Neurosci. Res.,69, pp. 976–986

    Article  Google Scholar 

  • Teng, Y. D., Lavik, E. B., Qu, X., Park, K. I., Ourednik, J., Zurakowski, D., Langer, R., andSnyder, E. Y. (2002): ‘Functional recovery following traumatic spinal cord injury mediated by a unique polymer scaffold seeded with neural stem cells’,Proc. Natl. Acad. Sci. USA,99, pp. 3024–3029

    Article  Google Scholar 

  • Vescovi, A. L., andSnyder, E. Y. (1999): ‘Establishment and properties of neural stem cell clones: plasticityin vitro andin vivo’,Brain Pathol.,9, pp. 569–598

    Google Scholar 

  • Vessey, C. J., andHall, P. D. L. M. (2001): ‘Hepatic stem cells: a review’,Pathology,33, pp. 130–141

    Article  Google Scholar 

  • Villa, A., Snyder, E. Y., Vescovi, A. L., andMartinez-Serrano, A. (2000): ‘Establishment and properties of a growth factor-dependent, perpetual neural stem cell line from the human CNS’,Exp. Neurol.,161, pp. 67–84

    Article  Google Scholar 

  • Voigt, M., Schauer, M., Schaefer, D. J., Andree, C., Horch, R., andStark, G. B. (1999): ‘Cultured epidermal keratinocytes on a microspherical transport system are feasible to reconstitute the epidermis in full-thickness wounds’,Tissue Eng.,5, pp. 563–572

    Google Scholar 

  • Waymack, P., Duff, R. G., Sabolinski, M., andApligraf Burn Study Group (2000): ‘The effect of a tissue engineered bilayered living skin analog, over meshed split-thickness autografts on the healing of excised burn wounds’,Burns,26, pp. 609–619

    Article  Google Scholar 

  • Weissman, I. L., Anderson, D. J., andGage, F. (2001): ‘Stem and progenitor cells: origins, phenotypes, lineage commitments, and transdifferentiations’,Ann. Rev. Cell Dev. Biol.,17, pp. 387–403

    Google Scholar 

  • Yandava, B. D., Billinghurst, L. L., andSnyder, E. Y. (1999): ‘“Global” cell replacement is feasible via neural stem cell transplantation: evidence from the dysmyelinated shiverer mouse brain’,Proc. Natl. Acad. Sci. USA,96, pp. 7029–7034

    Article  Google Scholar 

  • Zacchi, V., Soranzo, C., Cortivo, R., Radice, M., Brun, P., andAbatangelo, G. (1998): ‘In vitro engineering of human skin-like tissue’,J. Biomed. Mater: Res.,40, pp. 187–194

    Article  Google Scholar 

  • Zandstra, P. W., andNagy, A. (2001): ‘Stem cell bioengineering’,Ann. Rev. Biomed. Eng.,3, pp. 275–305

    Article  Google Scholar 

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Kallos, M.S., Sen, A. & Behie, L.A. Large-scale expansion of mammalian neural stem cells: a review. Med. Biol. Eng. Comput. 41, 271–282 (2003). https://doi.org/10.1007/BF02348431

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