Neural stem cell biology, plasticity and regeneration

Phil Horner, PhD has received a one-year, $33,000 award from the University of Washington Royalty Research Fund to study “Live Imaging of Stem Cells within a Neurogenic Niche” learn more about this >>>

Background

It is generally accepted that the adult central nervous system (CNS) does not spontaneously regenerate and neurons are not readily replaced. The limited recovery from traumatic or degenerative insults to the adult brain and spinal cord has devastating impact on patients and their loved ones. In addition, current treatments offer limited hope and provide inadequate improvement in quality of life. In the last decade, many of the molecular substrates that inhibit regeneration and cell replacement have begun to be elucidated.

In my lab we approach the problem of regeneration in two ways. The first is centered upon developing an understanding of stem cell biology as it pertains to the normal and injured CNS. The recent discovery of stem cells in the adult CNS has led to renewed hope for repair of debilitating neurological disorders. It is now accepted that stem cells routinely replace glia and neurons in specific regions of the brain throughout life. These observations have led to an exciting new perspective on brain plasticity but there is much to be discovered. The process of cell replacement is tightly regulated by endogenous signals that direct cell proliferation, migration and differentiation. My lab is actively engaged in discovering the factors that regulate these processes in the central nervous system.

The second major focus of the lab is neural regeneration in models of spinal cord injury and Glaucoma. Following injury to the central nervous system, a cascade of events leads to cellular proliferation and expression of molecules that act as physical and molecular inhibitors of axon growth. My lab is interested in utilizing gene therapy approaches to deliver growth promoting molecules that can stimulate axonal regeneration directly or block the effect of natural inhibitors. The ability of a neuron to regenerate its axon is determined by how the damaged axon responds to external factors found in the post-injury environment and to internal programs that govern growth. In previous work we have discovered that a small percentage of axons do re-grow in the adult CNS when presented with appropriate growth signals. We are currently trying to understand the genetic programs that allow one subset of axons to re-grow in an injury environment while another cannot.

Our goal is to unlock the potential of the adult nervous system by stimulating cell replacement via endogenous stem cells and to promote axonal regeneration through gene therapy.

Horner, P.J. and F.H.Gage. 2000. Regenerating the damaged central nervous system. Nature. 407:963-970

Research Question:

Stem cells and repair of the injured adult nervous system:
Can endogenous cells be recruited for self repair?

While it is now clear that the adult CNS contains stem cells and their progeny (e.g., the progenitor cell) there is a great deal of work that needs to be done. Future research must address the mechanisms underlying stem cell proliferation, migration and fate choice in order to approach translation of stem cell technologies into clinically relevant therapies. Recent experiments illustrate that there are important biologic roles being carried out by stem and progenitor cells resident in the embryonic and adult brain. Adult stem cells have similar capabilities to that of embryonic-derived cells in that they exhibit extensive self-renewal capacities and can differentiate into all of the major cell subtypes found in the adult CNS. This discovery suggests that the adult nervous system is not as "static" as previously thought. Hence, repair of the damaged CNS may be facilitated by activation of endogenous stem cells to participate in CNS regeneration. The presence of stem cells during adulthood suggests that they are participating in a state of homeostatic cell turnover. However, the importance of this process in health and disease is yet to be determined. Given the ethical and political complexities surrounding the use of embryonic tissue for treatment of neurological disorders, it is important to fully explore adult stem cells as a possible alternative source. Of course, with the observation of active stem cell populations within the CNS, we are led to ask an important question. If adult stem cells exist, why does spontaneous recovery and cell replacement not occur in the injured or degenerating brain? Are there natural inhibitors to cell replacement? Understanding the environmental signals in the intact and diseased CNS which influence stem cells is an important next step. Insight into these processes may help to elucidate the potential clinical utility of stem cells as it pertains to disorders affecting the CNS.

Lipson, A.C. and P.J. Horner. 2002. Potent possibilities: Endogenous stem cells in the adult spinal cord. Progress in Brain Research. Vol. 137, CH. 20, pg 283-297.

Research Highlights:

I. The Adult Spinal Cord Contains Multipotent Neural Stem Cells

II. Spinal Cord-Derived Progenitor Cells are Prevalent and Active

III. Fate Choice of Adult Neural Progenitor Cells can be Modulated after Spinal Cord Injury

IV. NT-3 Stimulates limited regeneration of the cortico-spinal tract following spinal contusion injury

Research Methods:

• Neural Stem Cell Cultures
  
• Fluorescence-Activated Cell Sorting
  
• Retrovirus production, purification and in vitro and in vivo cell transfection
  
• Quadruple immunofluorescent labeling of CNS tissue and confocal imaging
  
• Quantitative microscopy including stereological analysis of confocal and epifluorescent imaging
  
• CNS micro-surgery and stereotactic-transplantation
  
• CNS tract tracing and analysis
  
• Basic cell and molecular techniques
  
  • PCR, Western and Northern blot, mRNA and DNA purification, cell culture
• cDNA microarray.

Grants:

NIH R21:
Dr Philip Horner and Dr. Denise Inman have received a 2 year $275,000 NIH R21 award
to develop a "Murine Model for Regulation of Gliosis in Glaucoma".

Abstract:
Our knowledge of CNS glial cells now reveals that glia are much more than support cells for neurons. Glia provide structure to CNS tissue, guide migrating cells, regulate neurotransmitters in the extracellular milieu, produce signaling molecules, maintain synaptic connections, form the blood-brain barrier, monitor the environment and respond in myriad ways to injury and disease. Glial response to injury is called gliosis, and involves upregulation of intermediate filaments within the cell, changes in the complement of ion channels, secretion of signaling molecules and can also include proliferation. Gliosis occurs early in the chronic mouse model of glaucoma, and the magnitude of the response in all glial populations is significant. As of yet, the positive or negative impact of gliosis on RGC health and the progression of glaucoma has not been studied. This research will study the role gliosis plays in glaucoma by tempering the glial response in an animal model.

NIH R01:
"Regulation of Adult Progenitor Cells and Neural Repair".

Abstract:
Injury to the adult mammalian CNS leads to cell death and the severing of axonal connections that exhibit limited regeneration. Following spinal cord injury the functions of severed pathways, including locomotion, sensation and autonomic function are permanently lost leaving patients with a devastating level of disability. The failure to regenerate spinal pathways is due, in part, to the formation of physical and molecular barriers to axonal regrowth. This project seeks to define the plasticity of the NG2-expressing progenitor cell, its growth factor responsiveness and function in repair of the adult spinal cord. In addition, direct modification of endogenous progenitor cells using epidermal growth factor (EGF) is proposed to maintain NG2-expressing cells in a proliferative state and increase recovery from spinal cord injury. Direct activation of the endogenous NG2-cells with EGF will be used to increase the period of plasticity following injury and improve the regeneration of motor pathways. Overall, these experiments will better define the role and regulation of progenitor cells in the injured adult spinal cord and explore a novel approach to increase functional plasticity by regulation of endogenous proliferation.

Selected Bibliography:

SELECTED PUBLISHED AND ACCEPTED ARTICLES IN REFEREED JOURNALS:

Drew L. Sellers, Don O. Maris, and P.J. Horner. 2009. Post-injury niches direct temporal shifts in progenitor fate-choice to direct niche succession and repair after spinal cord injury. Journal of Neuroscience, 29(20):6722-33

Lasiene J, Matsui A, Sawa Y, Wong F, and P.J. Horner. Age-related myelin dynamicsrevealed by increased oligodendrogenesis and short internodes. Aging Cell. 2009. Apr;8(2):201-13.

Rhee, W. S. Ray, H. Yokoo, M.E. Hoane, C. C. Lee, A. M. Mikheev, P.J. Horner, R.C. Rostomily. 2009. Quantitative analysis of mitotic Olig2 cells in adult human brain and gliomas: implications for glioma histogenesis and biology. Glia, 2009 Apr 1;57(5):510-23

Calkins D, Horner PJ, Roberts R, Gradianu M, Berkowitz BA. 2008. Manganese-enhanced MRI of the DBA/2J Mouse Model of Hereditary Glaucoma. Invest Ophthalmol Vis Sci. 49(11):5083-8

Bosco, A., D.M.Inman, M.R.Steele, G.Wu, N.Marsh-Armstrong, D.J. Calkins, P.J. Horner, M.L.Vetter. 2008. Reduced retina microglial activation and improved optic nerve integrity with minocycline treatment in the DBA/2J mouse model of glaucoma. IOVS, 49(4):1437-46

Lasiene, J., L. Shupe, S. Perlmutter, P.J. Horner. 2008. No evidence for chronic demyelination in spared axons following spinal cord injury in a mouse. Journal of Neuroscience, 9;28(15):3887-96

Buckingham, B., D. Inman, W. Lambert, E. Oglesby, D.J. Calkins, M. Steele, M. Vetter, N. Marsh-Armstrong and P.J. Horner. 2008. Progressive Ganglion Cell Degeneration Precedes Neuronal Loss in a Mouse Model of Glaucoma. Journal of Neuroscience, 28(11):2735-44

Soto., I., E. Oglesby, B.P. Buckingham, J.L. Son, E.D.O. Roberson, M.R. Steele, D.M. Inman, M.L. Vetter, P.J. Horner and N. Marsh-Armstrong. 2007. Retinal Ganglion Cells Down-regulate Gene Expression and Lose their Axons within the Optic Nerve Head in a Mouse Glaucoma Model. Journal of Neuroscience, 9;28(2):548-61.

Petit, A., D.L. Sellers, D. J. Liebl, M. Tessier-Lavigne, T.E. Kennedy, P.J. Horner. 2007. Adult Spinal Cord Progenitor Cells are repelled by netrin-1 in the embryonic and injured adult spinal cord. Proc Natl Acad Sci U S A. Nov 6;104(45):17837-42.

Inman, D.M. and P.J. Horner. 2007. Reactive Non-Proliferative Gliosis Predominates in a Chronic Model of Glaucoma. Glia, 55(9):942-53

Park, I., J. Lasiene, S-H. Chou, P.J. Horner, S. H. Pun. 2007. Neuron-specific delivery of nucleic acids mediated by Tet1-modified poly(ethylenimine). Journal of Gene Medicine, 9(8):691-702

Bergen, J.M., I. Park, P.J. Horner, S. H. Pun. 2007. Nonviral Approaches for Neuronal Delivery of Nucleic Acids. Pharm Res. Oct 12

Horky, L.L., F. Galimi, F.H. Gage and P.J. Horner. 2006. Fate of Endogenous Stem/Progenitor Cells Following Spinal Cord Injury. J. Comp. Neurol. 498(4):525-538

Inman, D.M, R.M. Sappington, P.J. Horner and D.J. Calkins. 2005.Quantitative Correlation of Optic Nerve Pathology with Ocular Pressure and Corneal Thickness in the DBA/2J Mouse Model of Glaucoma. IOVS 47(3):986-96

Steele, M.R., D.M. Inman, D.J. Calkins, P.J. Horner, M.L. Vetter. 2005. Microarray Analysis of Retinal Gene Expression in the DBA/2J Model of Glaucoma. IOVS 47(3):977-85

Arvanian, V.L., W.J. Bowers, A.J. Anderson, P.J. Horner, H.J. Federoff, L.M. Mendell. 2005. Combined delivery of neurotrophin-3 and NMDA receptors 2D subunit strengthens synaptic transmission in contused and staggered double hemisected spinal cord of neonatal rat
Exp. Neurol. 197(2):347-52

Sellers D.L. and P.J. Horner. 2005. Instructive Niches: Environmental Instructions that Confound NG2 Proteoglycan Expression and the Fate-Restriction of CNS Progenitors. Journal of Anatomy. 207:727–734

Gaulke, L.J., P.J. Horner, A.J. Fink, C.L. McNamara and R.R. Hicks. 2005. Environmental Enrichment Increases Progenitor Cell Survival in the Dentate Gyrus following Lateral Fluid Percussion Injury in Rats. Mol. Brain Research. 141(2):138-50

Horner, P.J. and T.D. Palmer. 2003. La vida loca! The nightlife of an astrocyte. Trends in Neuroscience. 26(11): 597-603

Milatovic, D., S. Zala-Milatovic, K.S. Montine, P.J. Horner, and T.J. Montine. 2003. Pharmacologic suppression of neuronal oxidative damage and dendritic degeneration following direct activation of glial innate immunity in mouse cerebrum. Journal of Neurochemistry. 87(6): 1518-26

Arvanian, V.L., P.J. Horner, F.H.Gage and L.M.Mendell. 2003. Chronic neurotrophin-3 strengthens synaptic connections to motoneurons in the neonatal rat. J. Neurosci. 23:8706-8712

South, S., T. Kohno, B.K. Kaspar, D. Hegarty, B. Vissel, C. Drake, M. Ohata, S. Jenab, A. Sailer, S. Malkmus, T. Masuyama, P.J. Horner, J. Bogulavsky, S. Tonegawa, F.H. Gage, T.L.Yaksh, C.J.Woolf, S.F. Heinemann, and C.E. Inturrisi. 2003. A Conditional Deletion of the NMDAR1 Subunit in Adult Spinal Cord Dorsal Horn Reduces NMDA Currents and Injury-Induced Pain. J. Neurosci. 23(12):5031-5040

Horner, P.J., M. Thallmair and F.H. Gage. 2003. Defining the NG2 cell of the adult central nervous system. Journal of Neurocytology. 31(6/7):469-480.

Moore, E.M., S. Presnell, U Garrigues, A. Guibot, E. LeGuern, D. Smith, L. Yao, T.E. Whitmore, T. Gilbert, T.D. Palmer, P.J. Horner, R.E. Kuestner. 2002. Expression of IL-17B in neurons and evaluation of its possible role in the chromosome 5q-linked form of Charcot-Marie-Tooth disease. Neuromuscular Disorders. 12(2):141-50

Lipson, A.C. and P.J. Horner. 2002. Potent possibilities: Endogenous stem cells in the adult spinal cord. Progress in Brain Research. 137:283-297

Horner, P.J. and F.H. Gage. 2002. Regeneration in the adult and aging brain. Archives of Neurology. 59(11):1717-1720

Horner, P.J. and F.H.Gage. 2000. Regenerating the damaged central nervous system. Nature. 407:963-970

Shihabuddin, L., P.J. Horner, J. Ray and F.H.Gage. 2000. Adult spinal cord stem cells generate neurons after transplantation in the adult dentate gyrus. J.Neurosci. 20(23):8727-8735

Horner, P.J., A.E.Power, G.Kempermann, H.G.Kuhn, T.D.Palmer, J. Winkler, L.J.Thal, and F.H. Gage. 2000. Existence of progenitor cells throughout the intact adult rat spinal cord. J.Neurosci. 20(6): 2218-2228

McTigue, D. M., P. J. Horner, B. T. Stokes, and F. H. Gage.1998. Neurotrophin-3 and brain-derived neurotrophic factor induce oligodendrocyte proliferation, migration, and myelination in the contused adult rat spinal cord. J.Neurosci. 18(14):5354-5365

Popovich, P.G., P.J. Horner, B.B. Mullin, and B.T. Stokes. 1996. A Quantitative Spatial Analysis of the Blood-Spinal Cord Barrier I. Permeability after Spinal Contusion Injury. Exp. Neurol. 142:258-275

Horner, P.J., P.G. Popovich, B.B. Mullin, and B.T. Stokes. 1996. A Quantitative Spatial Analysis of the Blood-Spinal Cord Barrier II. Permeability after Intraspinal Fetal Transplantation. Exp. Neurol. 142:226-243

Horner, P.J., P.J. Reier, and B.T. Stokes. 1996. Quantitative Analysis of Vascularization and Cytochrome Oxidase Following Fetal Transplantation in the Contused Rat Spinal Cord. J. Comp. Neurol. 364(4):690-703

Horner, P.J. and B.T. Stokes. 1995. Fetal Transplantation Following Spinal Contusion Injury Results in Chronic Alterations in CNS Glucose Metabolism. Exp. Neurol.133(2):231-243

BOOK CHAPTERS:

Jacobson, B.E. and P.J. Horner. 2008. Gliogenesis in the Adult Spinal Cord. Postnatal and Adult Neurogenesis, Research Signpost, L. Bonfanti editor, Ch 6, pg 83-100.

McDonald, J.W. and the APA Spinal Cord Injury Consortium. 1999. Spinal Cord Injury: New Insights for Novel Treatments. Scientific American. August, pg65-73.

Senut, M-C., I. Aubert, P.J. Horner, and F.H. Gage. 1997. Gene Transfer for Adult CNS Regeneration and Aging. Gene Transfer and Therapy for Neurobiological Disorders, Umana Press, E.A. Choicca and X.O. Brakefield editors, Ch 17, pg 341-371.

Stokes, B.T., P.J. Horner, and M. Akino. 1996. Spinal Cord Injury Modeling and Functional Assessment. Central Nervous System Trauma: Research Techniques, CRC Press, S.T. Ohnishi and T. Ohnishi editors, Ch 20.

Stokes, B.T. and P.J. Horner. 1996. Spinal Cord Injury Modeling and Outcome Assessment. Neurotrauma: A Comprehensive Textbook on Head and Spinal Injury, McGraw-Hill, Inc., R.K. Narayan, J.E. Wilberger, and J.T. Povlishock editors, Ch 103, pg 1395-1402.

Horner, P.J., P.G. Popovich, P.J. Reier and B.T. Stokes. 1994. Fetal Spinal Transplant Vascularity: Metabolic and Immunologic Mechanisms. Neural Transplantation, CNS Neuronal Injury, and Regeneration - Recent Advances , CRC press, Joe Marwah, Herman Teitelbaum, and Kedar N. Prasad editors, ch 9, pg 119-140.

Stokes, B.T., M. Kim-Lee and P.J. Horner. 1993. Calcium Paradox: A Novel Mode of Secondary CNS Injury. Neurosurgical Topics Series, 14 Spinal Trauma: Current Evaluation and Management, Gary Rea and Carole Miller editors, ch 13, pg 207-212.

OTHER PUBLICATIONS:

Horner, P.J. and F.H. Gage. 1997. Experimental Spinal Cord Regeneration: Promising New Approaches. Brain Work Vol. 7 No. 2.