Abstract

We grafted human spinal cord–derived neural progenitor cells (NPCs) into sites of cervical spinal cord injury in rhesus monkeys (Macaca mulatta). Under three-drug immunosuppression, grafts survived at least 9 months postinjury and expressed both neuronal and glial markers. Monkey axons regenerated into grafts and formed synapses. Hundreds of thousands of human axons extended out from grafts through monkey white matter and synapsed in distal gray matter. Grafts gradually matured over 9 months and improved forelimb function beginning several months after grafting. These findings in a 'preclinical trial' support translation of NPC graft therapy to humans with the objective of reconstituting both a neuronal and glial milieu in the site of spinal cord injury.

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References

  1. 1.

    & Spinal cord injury: plasticity, regeneration and the challenge of translational drug development. Trends Neurosci. 32, 41–47 (2009).

  2. 2.

    Overcoming inhibition in the damaged spinal cord. J. Neurotrauma 23, 371–383 (2006).

  3. 3.

    & CNS injury, glial scars, and inflammation: inhibitory extracellular matrices and regeneration failure. Exp. Neurol. 209, 294–301 (2008).

  4. 4.

    & The Nogo signaling pathway for regeneration block. Annu. Rev. Neurosci. 27, 341–368 (2004).

  5. 5.

    Nogo and axon regeneration. Curr. Opin. Neurobiol. 14, 118–124 (2004).

  6. 6.

    & Growth factors and combinatorial therapies for CNS regeneration. Exp. Neurol. 209, 313–320 (2008).

  7. 7.

    Recapitulate development to promote axonal regeneration: good or bad approach? Phil. Trans. R. Soc. Lond. B 361, 1565–1574 (2006).

  8. 8.

    et al. Combined intrinsic and extrinsic neuronal mechanisms facilitate bridging axonal regeneration one year after spinal cord injury. Neuron 64, 165–172 (2009).

  9. 9.

    & Axonal projections between fetal spinal cord transplants and the adult rat spinal cord: a neuroanatomical tracing study of local interactions. J. Comp. Neurol. 307, 311–334 (1991).

  10. 10.

    , , & Fetal cell grafts into resection and contusion/compression injuries of the rat and cat spinal cord. Exp. Neurol. 115, 177–188 (1992).

  11. 11.

    & Axon outgrowth from grafts of human embryonic spinal cord in the lesioned adult rat spinal cord. Neuroreport 3, 1045–1048 (1992).

  12. 12.

    et al. Axonal regeneration and functional recovery after complete spinal cord transection in rats by delayed treatment with transplants and neurotrophins. J. Neurosci. 21, 9334–9344 (2001).

  13. 13.

    et al. Human neural stem cells differentiate and promote locomotor recovery in spinal cord–injured mice. Proc. Natl. Acad. Sci. USA 102, 14069–14074 (2005).

  14. 14.

    , , , & Human neural stem cells differentiate and promote locomotor recovery in an early chronic spinal cord injury NOD-scid mouse model. PLoS One 5, e12272 (2010).

  15. 15.

    et al. Grafted neural progenitors integrate and restore synaptic connectivity across the injured spinal cord. J. Neurosci. 31, 4675–4686 (2011).

  16. 16.

    et al. Recovery of supraspinal control of stepping via indirect propriospinal relay connections after spinal cord injury. Nat. Med. 14, 69–74 (2008).

  17. 17.

    et al. Long-distance growth and connectivity of neural stem cells after severe spinal cord injury. Cell 150, 1264–1273 (2012).

  18. 18.

    et al. Long-distance axonal growth from human induced pluripotent stem cells after spinal cord injury. Neuron 83, 789–796 (2014).

  19. 19.

    et al. Spinal cord reconstitution with homologous neural grafts enables robust corticospinal regeneration. Nat. Med. 22, 479–487 (2016).

  20. 20.

    et al. Extensive spinal decussation and bilateral termination of cervical corticospinal projections in rhesus monkeys. J. Comp. Neurol. 513, 151–163 (2009).

  21. 21.

    in Handbook of Physiology (eds. Brooks, V.B., Brookhart, J.M. & Mountcastle, V.B.). Anatomy of the descending pathways (The American Physiological Society, 1981).

  22. 22.

    et al. Bilateral corticospinal projections arise from each motor cortex in the macaque monkey: a quantitative study. J. Comp. Neurol. 473, 147–161 (2004).

  23. 23.

    & Manual dexterity and corticospinal connectivity following unilateral section of the cervical spinal cord in the macaque monkey. J. Comp. Neurol. 381, 307–319 (1997).

  24. 24.

    et al. Large animal and primate models of spinal cord injury for the testing of novel therapies. Exp. Neurol. 269, 154–168 (2015).

  25. 25.

    in Translational Neuroscience 1st edn. (ed. Tuszynski, M.H.) Conclusion (Springer US, 2016).

  26. 26.

    , , , & Preclinical efficacy failure of human CNS-derived stem cells for use in the pathway study of cervical spinal cord injury. Stem Cell Rep. 8, 249–263 (2017).

  27. 27.

    et al. Extensive spontaneous plasticity of corticospinal projections after primate spinal cord injury. Nat. Neurosci. 13, 1505–1510 (2010).

  28. 28.

    et al. Functional recovery in rats with ischemic paraplegia after spinal grafting of human spinal stem cells. Neuroscience 147, 546–560 (2007).

  29. 29.

    , , , & Characterization of a human fetal spinal cord stem cell line, NSI-566RSC, and its induction to functional motoneurons. J. Tissue Eng. Regen. Med. 4, 181–193 (2010).

  30. 30.

    et al. Lumbar intraspinal injection of neural stem cells in patients with amyotrophic lateral sclerosis: results of a phase I trial in 12 patients. Stem Cells 30, 1144–1151 (2012).

  31. 31.

    et al. SOX9 Is an astrocyte-specific nuclear marker in the adult brain outside the neurogenic regions. J. Neurosci. 37, 4493–4507 (2017).

  32. 32.

    et al. Ependymal cell contribution to scar formation after spinal cord injury is minimal, local and dependent on direct ependymal injury. Sci. Rep. 7, 41122 (2017).

  33. 33.

    et al. Identification of Tmem10/Opalin as an oligodendrocyte enriched gene using expression profiling combined with genetic cell ablation. Glia 56, 1176–1186 (2008).

  34. 34.

    et al. Prolonged human neural stem cell maturation supports recovery in injured rodent CNS. J. Clin. Invest. 127, 3287–3299 (2017).

  35. 35.

    , , , & Molecular logic of neocortical projection neuron specification, development and diversity. Nat. Rev. Neurosci. 14, 755–769 (2013).

  36. 36.

    , & Neurotransmitters in nociceptive modulatory circuits. Annu. Rev. Neurosci. 14, 219–245 (1991).

  37. 37.

    & The role of serotonin in reflex modulation and locomotor rhythm production in the mammalian spinal cord. Brain Res. Bull. 53, 689–710 (2000).

  38. 38.

    et al. Methods for functional assessment after C7 spinal cord hemisection in the rhesus monkey. Neurorehabil. Neural Repair 26, 556–569 (2012).

  39. 39.

    et al. Animal models of neurologic disorders: a nonhuman primate model of spinal cord injury. Neurotherapeutics 9, 380–392 (2012).

  40. 40.

    et al. A unilateral cervical spinal cord contusion injury model in non-human primates (Macaca mulatta). J. Neurotrauma 33, 439–459 (2016).

  41. 41.

    et al. Pronounced species divergence in corticospinal tract reorganization and functional recovery after lateralized spinal cord injury favors primates. Sci. Transl. Med. 7, 302ra134 (2015).

  42. 42.

    et al. Fetal and adult human oligodendrocyte progenitor cell isolates myelinate the congenitally dysmyelinated brain. Nat. Med. 10, 93–97 (2004).

  43. 43.

    & Glial progenitor cell-based treatment of the childhood leukodystrophies. Exp. Neurol. 283 Part B, 476–488 (2016).

  44. 44.

    , , , & Characterization of ectopic colonies that form in widespread areas of the nervous system with neural stem cell transplants into the site of a severe spinal cord injury. J. Neurosci. 34, 14013–14021 (2014).

  45. 45.

    et al. Neural stem cell dissemination after grafting to CNS injury sites. Cell 156, 388–389 (2014).

  46. 46.

    , , & Reliable activation of immature neurons in the adult hippocampus. PLoS One 4, e5320 (2009).

  47. 47.

    et al. Low excitatory innervation balances high intrinsic excitability of immature dentate neurons. Nat. Commun. 7, 11313 (2016).

  48. 48.

    , , , & Induction of corticospinal regeneration by lentiviral trkB-induced Erk activation. Proc. Natl. Acad. Sci. USA 106, 7215–7220 (2009).

  49. 49.

    et al. Extensive cell migration, axon regeneration, and improved function with polysialic acid-modified Schwann cells after spinal cord injury. Glia 60, 979–992 (2012).

  50. 50.

    et al. PTEN deletion enhances the regenerative ability of adult corticospinal neurons. Nat. Neurosci. 13, 1075–1081 (2010).

  51. 51.

    et al. Restoration of skilled locomotion by sprouting corticospinal axons induced by co-deletion of PTEN and SOCS3. Nat. Commun. 6, 8074 (2015).

  52. 52.

    , , & Human spinal cord retains substantial structural mass in chronic stages after injury. J. Neurotrauma 16, 523–531 (1999).

  53. 53.

    , & in Translational Neuroscience 1st edn. (ed. Tuszynski, M.H.) Neural Stem Cells for Spinal Cord Injury (Springer US, 2016).

  54. 54.

    , , , & PTEN inhibition to facilitate intrinsic regenerative outgrowth of adult peripheral axons. J. Neurosci. 30, 9306–9315 (2010).

  55. 55.

    , , & Bisperoxovanadium compounds are potent PTEN inhibitors. FEBS Lett. 566, 35–38 (2004).

  56. 56.

    et al. Programmed cell death and the control of cell survival: lessons from the nervous system. Science 262, 695–700 (1993).

  57. 57.

    Size control: the regulation of cell numbers in animal development. Cell 86, 173–175 (1996).

  58. 58.

    , & Programmed cell death in animal development. Cell 88, 347–354 (1997).

  59. 59.

    & Neurite pruning and neuronal cell death: spatial regulation of shared destruction programs. Curr. Opin. Neurobiol. 23, 990–996 (2013).

  60. 60.

    Directional statistical decisions. Psychol. Rev. 67, 160–167 (1960).

  61. 61.

    The scree test for the number of Factors. Multivariate Behav. Res. 1, 245–276 (1966).

  62. 62.

    & Relation of sample size to the stability of component patterns. Psychol. Bull. 103, 265–275 (1988).

  63. 63.

    et al. Derivation of multivariate syndromic outcome metrics for consistent testing across multiple models of cervical spinal cord injury in rats. PLoS One 8, e59712 (2013).

  64. 64.

    & Statistical Analysis with Missing Data 2nd edn (John Wiley & Sons, Inc., 2002).

  65. 65.

    , & Breaking free of sample size dogma to perform innovative translational research. Sci. Transl. Med. 3, 87ps24 (2011).

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Acknowledgements

Human 566RSC-UBQT neural stem cells were a gift from NeuralStem, Inc. This work was supported by the Veterans Administration (Gordon Mansfield Spinal Cord Injury Collaborative Consortium, RR&D B7332R, MHT; RR&D RX001045, JHB), the National Institutes of Health (R01 NS042291, MHT; R01 NS104442, MHT), the Department of Defense (W81XWH-12-1-0592; E.S.R.), the Craig H. Neilsen Foundation (M.H.T.), the Bernard and Anne Spitzer Charitable Trust (M.H.T.), and the Dr. Miriam and Sheldon G. Adelson Medical Research Foundation (M.H.T.).

Author information

Affiliations

  1. Department of Neurosciences, University of California, San Diego, La Jolla, California, USA.

    • Ephron S Rosenzweig
    • , John H Brock
    • , Paul Lu
    • , Hiromi Kumamaru
    • , Ken Kadoya
    • , Janet L Weber
    • , Justine J Liang
    •  & Mark H Tuszynski
  2. Veterans Administration Medical Center, La Jolla, California, USA.

    • John H Brock
    • , Paul Lu
    •  & Mark H Tuszynski
  3. California National Primate Research Center, University of California, Davis, Davis, California, USA.

    • Ernesto A Salegio
    • , Rod Moseanko
    •  & Stephanie Hawbecker
  4. Department of Orthopaedic Surgery, Hokkaido University, Sapporo, Japan.

    • Ken Kadoya
  5. Department of Neurosurgery, University of California, San Francisco, San Francisco, California, USA.

    • J Russell Huie
    • , Adam R Ferguson
    • , Michael S Beattie
    •  & Jacqueline C Bresnahan
  6. Department of Neurology, University of California, Los Angeles, Los Angeles, California, USA.

    • Leif A Havton
  7. College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, USA.

    • Yvette S Nout-Lomas
  8. Veterans Administration Medical Center, San Francisco, California, USA.

    • Adam R Ferguson

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Contributions

E.S.R., J.H.B., P.L., E.A.S., K.K., L.A.H., Y.S.N.-L., A.R.F., M.S.B., J.C.B., and M.H.T. designed the experiments. E.S.R., J.H.B., P.L., H.K., E.A.S., K.K., R.M., S.H., Y.S.N.-L., J.C.B., and M.H.T. carried out the experiments. E.S.R., J.H.B., H.K., E.A.S., J.L.W., J.J.L., J.R.H., L.A.H., A.R.F., and M.H.T. analyzed the data. E.S.R. and M.H.T. wrote the manuscript. E.S.R., J.H.B., P.L., H.K., E.A.S., K.K., J.L.W., J.J.L., R.M., S.H., J.R.H., L.A.H., Y.S.N.-L., A.R.F., M.S.B., J.C.B., and M.H.T. edited the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Mark H Tuszynski.

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https://doi.org/10.1038/nm.4502