Human‐Induced Pluripotent Stem Cells form Functional Neurons and Improve Recovery After Grafting in Stroke‐Damaged Brain

Reprogramming of adult human somatic cells to induced pluripotent stem cells (iPSCs) is a novel approach to produce patient‐specific cells for autologous transplantation. Whether such cells survive long‐term, differentiate to functional neurons, and induce recovery in the stroke‐injured brain are unclear. We have transplanted long‐term self‐renewing neuroepithelial‐like stem cells, generated from adult human fibroblast‐derived iPSCs, into the stroke‐damaged mouse and rat striatum or cortex. Recovery of forepaw movements was observed already at 1 week after transplantation. Improvement was most likely not due to neuronal replacement but was associated with increased vascular endothelial growth factor levels, probably enhancing endogenous plasticity. Transplanted cells stopped proliferating, could survive without forming tumors for at least 4 months, and differentiated to morphologically mature neurons of different subtypes. Neurons in intrastriatal grafts sent axonal projections to the globus pallidus. Grafted cells exhibited electrophysiological properties of mature neurons and received synaptic input from host neurons. Our study provides the first evidence that transplantation of human iPSC‐derived cells is a safe and efficient approach to promote recovery after stroke and can be used to supply the injured brain with new neurons for replacement. STEM CELLS2012;30:1120–1133

[1]  A. Björklund,et al.  Reformation of long axon pathways in adult rat central nervous system by human forebrain neuroblasts , 1990, Nature.

[2]  Seung-Up Kim,et al.  Anti-inflammatory mechanism of intravascular neural stem cell transplantation in haemorrhagic stroke. , 2008, Brain : a journal of neurology.

[3]  Yi Li,et al.  Intravenous Administration of Human Bone Marrow Stromal Cells Induces Angiogenesis in the Ischemic Boundary Zone After Stroke in Rats , 2003, Circulation research.

[4]  James A. Thomson,et al.  Induced pluripotent stem cells from a spinal muscular atrophy patient , 2009, Nature.

[5]  S. Dunnett,et al.  The “staircase test”: a measure of independent forelimb reaching and grasping abilities in rats , 1991, Journal of Neuroscience Methods.

[6]  A. Zechariah,et al.  Implications of Vascular Endothelial Growth Factor for Postischemic Neurovascular Remodeling , 2009, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[7]  Stephen B. Dunnett,et al.  The Corridor Task: A simple test of lateralised response selection sensitive to unilateral dopamine deafferentation and graft-derived dopamine replacement in the striatum , 2005, Brain Research Bulletin.

[8]  Su-Chun Zhang,et al.  Survival and differentiation of transplanted neural stem cells derived from human induced pluripotent stem cells in a rat stroke model. , 2013, Journal of stroke and cerebrovascular diseases : the official journal of National Stroke Association.

[9]  M. Hoehn,et al.  Stem Cell Mediation of Functional Recovery after Stroke in the Rat , 2010, PloS one.

[10]  Marta P Pereira,et al.  Transplanted Stem Cell‐Secreted Vascular Endothelial Growth Factor Effects Poststroke Recovery, Inflammation, and Vascular Repair , 2011, Stem cells.

[11]  M. Peschanski,et al.  Striatal progenitors derived from human ES cells mature into DARPP32 neurons in vitro and in quinolinic acid-lesioned rats , 2008, Proceedings of the National Academy of Sciences.

[12]  Joakim Bjorkdahl.  Reduced brain edema and infarction volume in mice lacking the neuronal isoform of nitric oxide synthase after transient MCA occlusion , 1996 .

[13]  D. Gershon.  Stem-cell research , 2003, Nature.

[14]  Anders Björklund,et al.  Characterisation of behavioural and neurodegenerative changes induced by intranigral 6‐hydroxydopamine lesions in a mouse model of Parkinson’s disease , 2010, The European journal of neuroscience.

[15]  Mark J. West,et al.  Stereological methods for estimating the total number of neurons and synapses: issues of precision and bias , 1999, Trends in Neurosciences.

[16]  B. Siesjö,et al.  The omega-conopeptide SNX-111, an N-type calcium channel blocker, dramatically ameliorates brain damage due to transient focal ischaemia. , 1994, Acta Physiologica Scandinavica.

[17]  I. Weissman,et al.  Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[18]  G. Comi,et al.  Delayed post-ischaemic neuroprotection following systemic neural stem cell transplantation involves multiple mechanisms. , 2009, Brain : a journal of neurology.

[19]  O. Lindvall,et al.  Regulation of Brain-Derived Neurotrophic Factor Gene Expression after Transient Middle Cerebral Artery Occlusion with and without Brain Damage , 1995, Experimental Neurology.

[20]  Matthew Trotter,et al.  Capture of Neuroepithelial-Like Stem Cells from Pluripotent Stem Cells Provides a Versatile System for In Vitro Production of Human Neurons , 2012, PloS one.

[21]  S. V. Anisimov,et al.  Transplantation of Human Embryonic Stem Cell‐Derived Cells to a Rat Model of Parkinson's Disease: Effect of In Vitro Differentiation on Graft Survival and Teratoma Formation , 2006, Stem cells.

[22]  R. Jaenisch,et al.  Neurons derived from reprogrammed fibroblasts functionally integrate into the fetal brain and improve symptoms of rats with Parkinson's disease , 2008, Proceedings of the National Academy of Sciences.

[23]  P. Greengard,et al.  DARPP-32, a dopamine- and adenosine 3':5'-monophosphate-regulated phosphoprotein enriched in dopamine-innervated brain regions. III. Immunocytochemical localization , 1984, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[24]  M. Moskowitz,et al.  Reduced Brain Edema and Infarction Volume in Mice Lacking the Neuronal Isoform of Nitric Oxide Synthase after Transient MCA Occlusion , 1996, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[25]  Alberto Martínez‐Serrano,et al.  Amelioration of ischaemia‐induced neuronal death in the rat striatum by NGF‐secreting neural stem cells , 1998, The European journal of neuroscience.

[26]  O. Lindvall,et al.  Upregulation of p75 Neurotrophin Receptor after Stroke in Mice Does Not Contribute to Differential Vulnerability of Striatal Neurons , 2001, Experimental Neurology.

[27]  Marta P Pereira,et al.  Human neural stem cells enhance structural plasticity and axonal transport in the ischaemic brain , 2011, Brain : a journal of neurology.

[28]  Tamar Licht,et al.  Reversible modulations of neuronal plasticity by VEGF , 2011, Proceedings of the National Academy of Sciences.

[29]  Anne-Lise D. Maag,et al.  Adherent Self-Renewable Human Embryonic Stem Cell-Derived Neural Stem Cell Line: Functional Engraftment in Experimental Stroke Model , 2008, PloS one.

[30]  R. Traystman.  Stem Cell Research in Stroke How Far From the Clinic ? , 2011 .

[31]  P. Gutin,et al.  Migration and differentiation of neural precursors derived from human embryonic stem cells in the rat brain , 2005, Nature Biotechnology.

[32]  M. Chopp,et al.  Contralesional Axonal Remodeling of the Corticospinal System in Adult Rats After Stroke and Bone Marrow Stromal Cell Treatment , 2008, Stroke.

[33]  O. Lindvall,et al.  Cell Number and Timing of Transplantation Determine Survival of Human Neural Stem Cell Grafts in Stroke-Damaged Rat Brain , 2011, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[34]  Anne-Lise D. Maag,et al.  Human Neural Stem Cell Grafts Modify Microglial Response and Enhance Axonal Sprouting in Neonatal Hypoxic–Ischemic Brain Injury , 2010, Stroke.

[35]  S. Yamanaka,et al.  Induction of Pluripotent Stem Cells from Mouse Embryonic and Adult Fibroblast Cultures by Defined Factors , 2006, Cell.

[36]  Hynek Wichterle,et al.  Induced Pluripotent Stem Cells Generated from Patients with ALS Can Be Differentiated into Motor Neurons , 2008, Science.

[37]  S. Dunnett,et al.  The staircase test of skilled reaching in mice , 2001, Brain Research Bulletin.

[38]  Hsin-Yang Li,et al.  Functional improvement of focal cerebral ischemia injury by subdural transplantation of induced pluripotent stem cells with fibrin glue. , 2010, Stem cells and development.

[39]  O. Brüstle,et al.  Human embryonic stem cell-derived neurons establish region-specific, long-range projections in the adult brain , 2011, Cellular and Molecular Life Sciences.

[40]  K. Reymann,et al.  Neuronal differentiation of transplanted embryonic stem cell-derived precursors in stroke lesions of adult rats. , 2006, Brain : a journal of neurology.

[41]  K. Abe,et al.  Tridermal Tumorigenesis of Induced Pluripotent Stem Cells Transplanted in Ischemic Brain , 2010, Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism.

[42]  O. Brüstle,et al.  A rosette-type, self-renewing human ES cell-derived neural stem cell with potential for in vitro instruction and synaptic integration , 2009, Proceedings of the National Academy of Sciences.

[43]  Z. Kokaia,et al.  Survival, migration and neuronal differentiation of human fetal striatal and cortical neural stem cells grafted in stroke‐damaged rat striatum , 2007, The European journal of neuroscience.

[44]  R. Jaenisch,et al.  Differentiated Parkinson patient-derived induced pluripotent stem cells grow in the adult rodent brain and reduce motor asymmetry in Parkinsonian rats , 2010, Proceedings of the National Academy of Sciences.

[45]  E. Hogan,et al.  A model of focal ischemic stroke in the rat: reproducible extensive cortical infarction. , 1986, Stroke.

[46]  Seung U. Kim,et al.  Human Neural Stem Cells Over-Expressing VEGF Provide Neuroprotection, Angiogenesis and Functional Recovery in Mouse Stroke Model , 2007, PloS one.

[47]  D. Corbett,et al.  Transplantation of human embryonic stem cell‐derived neural precursor cells and enriched environment after cortical stroke in rats: cell survival and functional recovery , 2009, The European journal of neuroscience.

[48]  R. Malenka,et al.  Molecular and magnetic resonance imaging of human embryonic stem cell-derived neural stem cell grafts in ischemic rat brain. , 2009, Molecular therapy : the journal of the American Society of Gene Therapy.

[49]  Q. Dong,et al.  Induction of pluripotent stem cells transplantation therapy for ischemic stroke , 2011, Molecular and Cellular Biochemistry.