29th Annual National Neurotrauma Symposium<br />Reconstructing the Injured Brain & Spinal Cord<br />with Stem Cells<br />J...
<ul><li>IN VIVO
Neurotraumatic Conditions (“The Chaperone Affect”)
 Stroke/hypoxic-ischemic injury
 Spinal Cord Injury / Traumatic Brain Injury
Not just diffusible factors but cell-cell contact via gap junctions, as well
 Improved “homing”/pathotropism
 IN VITRO --“Disease in a Dish”
Modeling development, injury, disease, ?therapy </li></ul>• hESCs & HIPSCs into “neural tubes” ==> ultimately into hNSCs)<...
Themes<br /><ul><li> Stem cells simply 1 component in a series of  			intrinsic developmental programs “designed” to</li><...
 including stem cell-mediated</li></ul>	‘rescue’/ ‘protection’ / ‘regeneration’ /<br />	altered gene expression /<br />	cr...
Host<br />NSC<br />NSC<br />
Host<br />NSC<br />influence<br />differentiation<br />fate<br />NSC<br />
…Or might be biased to<br />yield this neural cell type in a <br />neuron deficient environment<br />
Park et al, 2006<br />
Park et al, (2006)<br />
Park et al, Nature Biotech (2002)<br />
Neurons<br />(oligos)<br />(astros)<br />(NPs)<br />
Host<br />NSC<br />NSC<br />
G<br />G<br />GDNF<br />GDNF<br />N<br />N<br />N<br />Ret<br />Ret<br />GDNF<br />GDNF<br />GDNF<br />GDNF<br />GDNF<br /...
Host<br />NSC<br />NSC<br />
Host<br />NSC<br />NSC<br />Diffusible<br />Factors<br />
Host<br />NSC<br />NSC<br />Lysosomal<br />Enzymes<br />
Host<br />NSC<br />NSC<br />Anti-<br />Inflammation<br />
Host<br />NSC<br />NSC<br />Protection<br />
Host<br />NSC<br />NSC<br />Anti-<br />Scarring<br />
Host<br />NSC<br />NSC<br />Pro-<br />Mobilization<br />
Host<br />NSC<br />NSC<br />Pro-<br />Neurite <br />Outgrowth<br />
Host<br />NSC<br />NSC<br />Pro-<br />Angiogenic<br />
<ul><li> ALS / (?SMA)
 Parkinson’s Disease (Nat Biotech ‘02; PNAS ‘07; Stem Cells ‘09)
Neurogenetic degeneration (Nat Med ‘07; Stem Cells ‘09)
 Some aging-related degeneration
 Spinal cord injury & Head trauma (PNAS ‘02; PNAS’10)
 Stroke / Hypoxic-Ischemia (Nat Biotech ‘02; Exp Neurol ’06; PNAS’11)
Cerebellar Degeneration (J Neurosci ‘06; PNAS ‘06; PNAS’10)</li></ul>NSC-mediated‘rescue’/ ‘protection’ / ‘regeneration’/a...
(Infarcted)<br />(Intact)<br />Park et al, Nature Biotech (2002)<br />
Park et al, Nature Biotech (2002)<br />
Diminished<br />Scarring<br />in stroke<br />following<br />human<br />NSC <br />Implantation<br />(supported by scaffold)...
Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />
Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />(gap junctions)<br />
Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />(gap junctions)<br />
Early neuronal membrane properties emerge in NSCs during differentiation in slice<br />Ability to generate action potentia...
Spreading Calcium Waves<br />Jaderstad et al, PNAS (2010)<br />
Role of calcium fluctuations<br />Apoptosis<br />Astrocytic control of cerebral micro-circulation<br />Synaptic informatio...
Expression of first Connexin 43 & then Connexin 26 are important components in gap junction formation<br />DAP1<br />GFP<b...
Gap Junctions<br />
Functional Gap-junctional Intercellular Channels<br />Jaderstad et al, PNAS ‘10<br />
Pharmacological Suppression of Gap Junction Function(via carbenoxolone [CBX] or  18--glycyrrhetinic acid [18--GA]) Block...
Pharmacological Suppression of Gap Junction Function(via carbenoxolone) Blocks Glutamate-Initiated Calcium Influx in Host ...
Cell Death(i.e. % of Propidium Iodide+ host striatal cells)decreases when NSCs making gap junctions are present<br />Witho...
Even SmallSuppression of Connexin 43 Expression(via siRNA)--& henceGap Junction Formation--Abrogates a Component ofNSC’s B...
Jaderstad<br />et al,<br />PNAS ‘10<br />
<ul><li> ALS / (?SMA)
 Parkinson’s Disease (Nat Biotech ‘02; PNAS ‘07; Stem Cells ‘09)
Neurogenetic degeneration (Nat Med ‘07; Stem Cells ‘09)
 Some aging-related degeneration
 Spinal cord injury & Head trauma (PNAS ‘02; PNAS’10)
 Stroke / Hypoxic-Ischemia (Nat Biotech ‘02; Exp Neurol ’06; PNAS’11)
Cerebellar Degeneration (J Neurosci ‘06; PNAS ‘06; PNAS’10)</li></ul>NSC-mediated‘rescue’/ ‘protection’ / ‘regeneration’/a...
50 µm<br />0.5 cm<br />o.<br />Cerebellar <br />Purkinje Cell Neuron<br />Degeneration<br />Mutants<br />Li et al, PNAS & ...
Calbindin<br />Purkinje Cell Layer evident following early<br />transplantation of<br />neural stem cells<br />
Intercellular Contacts of NSCs with Nervous Mutant Purkinje Neurons<br />bgal (NSC)<br />Calb (PN)<br />Dapi (nuclei)<br /...
Host Nr Purkinje Neurons  (PNs) Remain At Nearly Wild Type Numbers<br />Following Early NSC Transplantation<br />Li, et al...
70<br />60<br />y = 0.5032x - 4.8333<br />50<br />2<br />R<br /> = 0.688 (n=8)<br />P<0.05<br />40<br />30<br />Rotarod te...
Influence of NSCs upon Nr PNs (via cell-cell contact) appears to be mediated by gap junctions (particularly those employin...
Blocking Gap Junction Formation(with Cx43 RNAi)abrogatesNSC-mediated neuronal rescue<br />Jaderstad et al, PNAS (2010)<br />
WHAT ARE THE GAP JUNCTIONS DOING?<br />
<ul><li>There is an ideal amount of  tissue plasminogen activator (TPA) Activity
 TPA in its role as upstream of a number of signal transduction pathways
 e.g., neurotrophic factor handling; mitochondrial function
Nervous mutant has abnormally high TPA levels
 Abnormalities emulated by  TPA
 NSC intercession (somehow) restores (lowers) level to normal</li></ul>A different way of viewing “gene therapy” -- homeos...
Nervous<br />Cerebellum<br />NSC<br />tPA <br />Plasminogen<br />Plasmin<br />Mitochondrial<br />VDAC<br />Neurotrophins...
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Strategies for enhancing neural cell efficacy in the host cns

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  • 4/30/11
  • Snyder, Evan

    1. 1. 29th Annual National Neurotrauma Symposium<br />Reconstructing the Injured Brain & Spinal Cord<br />with Stem Cells<br />July 13, 2011 – Hollywood Beach, Florida<br />Strategies for Enhancing Neural Stem Cell<br />Efficacy in the Host CNS <br />Evan Y. Snyder, MD, PhD, FAAP<br />Professor; Director, Program for Stem Cells & Regenerative Biology<br /> Director, Stem Cell Research Center<br /> Sanford-Burnham Institute for Medical Research<br />Depts. of Pediatrics (Newborn Intensive Care) & Neurology/Neuroscience, University of California, San Diego<br />Biomedical Sciences Graduate Program, University of California, San Diego<br />Steering Committee, Sanford (San Diego) Consortium for Regenerative Medicine<br />Associate Member, Sanford Pediatric Research Center<br />
    2. 2. <ul><li>IN VIVO
    3. 3. Neurotraumatic Conditions (“The Chaperone Affect”)
    4. 4. Stroke/hypoxic-ischemic injury
    5. 5. Spinal Cord Injury / Traumatic Brain Injury
    6. 6. Not just diffusible factors but cell-cell contact via gap junctions, as well
    7. 7. Improved “homing”/pathotropism
    8. 8. IN VITRO --“Disease in a Dish”
    9. 9. Modeling development, injury, disease, ?therapy </li></ul>• hESCs & HIPSCs into “neural tubes” ==> ultimately into hNSCs)<br />• neural-vascular obligate co-patterning/co-dependence – driven by neural crest<br />• high-throughput drug discovery<br />• large scale comprehensive profiling<br />
    10. 10. Themes<br /><ul><li> Stem cells simply 1 component in a series of intrinsic developmental programs “designed” to</li></ul>(a) construct & (b) maintain homeostasis<br /> (“cell replacement” is just 1 -- perhaps not even the most important mechanism by which equipoise is restored to a perturbed system)<br /><ul><li> Stem Cell-MediatedCross-TalkAdds Complexity but Also Perhaps Additional Therapeutic Opportunities
    11. 11. including stem cell-mediated</li></ul> ‘rescue’/ ‘protection’ / ‘regeneration’ /<br /> altered gene expression /<br /> creation of more supportive milieu<br />Complementing ‘cell replacement’<br />
    12. 12. Host<br />NSC<br />NSC<br />
    13. 13.
    14. 14. Host<br />NSC<br />influence<br />differentiation<br />fate<br />NSC<br />
    15. 15.
    16. 16. …Or might be biased to<br />yield this neural cell type in a <br />neuron deficient environment<br />
    17. 17.
    18. 18. Park et al, 2006<br />
    19. 19. Park et al, (2006)<br />
    20. 20. Park et al, Nature Biotech (2002)<br />
    21. 21.
    22. 22. Neurons<br />(oligos)<br />(astros)<br />(NPs)<br />
    23. 23.
    24. 24. Host<br />NSC<br />NSC<br />
    25. 25. G<br />G<br />GDNF<br />GDNF<br />N<br />N<br />N<br />Ret<br />Ret<br />GDNF<br />GDNF<br />GDNF<br />GDNF<br />GDNF<br />GDNF<br />Host<br />NSC<br />
    26. 26. Host<br />NSC<br />NSC<br />
    27. 27. Host<br />NSC<br />NSC<br />Diffusible<br />Factors<br />
    28. 28. Host<br />NSC<br />NSC<br />Lysosomal<br />Enzymes<br />
    29. 29. Host<br />NSC<br />NSC<br />Anti-<br />Inflammation<br />
    30. 30. Host<br />NSC<br />NSC<br />Protection<br />
    31. 31. Host<br />NSC<br />NSC<br />Anti-<br />Scarring<br />
    32. 32. Host<br />NSC<br />NSC<br />Pro-<br />Mobilization<br />
    33. 33. Host<br />NSC<br />NSC<br />Pro-<br />Neurite <br />Outgrowth<br />
    34. 34. Host<br />NSC<br />NSC<br />Pro-<br />Angiogenic<br />
    35. 35. <ul><li> ALS / (?SMA)
    36. 36. Parkinson’s Disease (Nat Biotech ‘02; PNAS ‘07; Stem Cells ‘09)
    37. 37. Neurogenetic degeneration (Nat Med ‘07; Stem Cells ‘09)
    38. 38. Some aging-related degeneration
    39. 39. Spinal cord injury & Head trauma (PNAS ‘02; PNAS’10)
    40. 40. Stroke / Hypoxic-Ischemia (Nat Biotech ‘02; Exp Neurol ’06; PNAS’11)
    41. 41. Cerebellar Degeneration (J Neurosci ‘06; PNAS ‘06; PNAS’10)</li></ul>NSC-mediated‘rescue’/ ‘protection’ / ‘regeneration’/anti-inflammation-- complementing ‘cell replacement’Neural stem cells display an<br />
    42. 42. (Infarcted)<br />(Intact)<br />Park et al, Nature Biotech (2002)<br />
    43. 43. Park et al, Nature Biotech (2002)<br />
    44. 44.
    45. 45. Diminished<br />Scarring<br />in stroke<br />following<br />human<br />NSC <br />Implantation<br />(supported by scaffold)<br />Kook In Park<br />
    46. 46. Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />
    47. 47. Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />(gap junctions)<br />
    48. 48. Host<br />NSC<br />NSC<br />Cell-Cell<br />Contact<br />(gap junctions)<br />
    49. 49. Early neuronal membrane properties emerge in NSCs during differentiation in slice<br />Ability to generate action potential<br />Jaderstad et al, PNAS ‘10<br />
    50. 50. Spreading Calcium Waves<br />Jaderstad et al, PNAS (2010)<br />
    51. 51. Role of calcium fluctuations<br />Apoptosis<br />Astrocytic control of cerebral micro-circulation<br />Synaptic information processing by astrocytes<br />Cell to cell signalling <br />Neuron ↔ neuron<br />Glia ↔ glia<br />Glia ↔ neuron<br />Cell cycle regulation<br />Cell migration timing<br />Differentiation<br />
    52. 52. Expression of first Connexin 43 & then Connexin 26 are important components in gap junction formation<br />DAP1<br />GFP<br />Tuj1<br />Cx26<br />Jaderstad et al, PNAS (2010)<br />
    53. 53. Gap Junctions<br />
    54. 54. Functional Gap-junctional Intercellular Channels<br />Jaderstad et al, PNAS ‘10<br />
    55. 55. Pharmacological Suppression of Gap Junction Function(via carbenoxolone [CBX] or 18--glycyrrhetinic acid [18--GA]) Blocks Dye Transfer to Host Cells in Engrafted Slice Cultures<br />Jaderstad et al, PNAS ‘10<br />
    56. 56. Pharmacological Suppression of Gap Junction Function(via carbenoxolone) Blocks Glutamate-Initiated Calcium Influx in Host Cells of Engrafted Slice Cultures<br />Jaderstad et al, PNAS (2010)<br />Eric Herlenius<br />
    57. 57. Cell Death(i.e. % of Propidium Iodide+ host striatal cells)decreases when NSCs making gap junctions are present<br />Without NSCs<br />With NSCs<br />PROPIDIUM IODIDE+ cells<br />(With<br />NSCs)<br />GFP/ DAPI / PROPIDIUM IODIDE<br />(No<br />NSCs)<br />Jaderstad et al, PNAS ‘10<br />
    58. 58. Even SmallSuppression of Connexin 43 Expression(via siRNA)--& henceGap Junction Formation--Abrogates a Component ofNSC’s Beneficial Impact on Host Cells(i.e., it’s anti-gliotic scar effect)<br />Jaderstad et al, PNAS ‘10<br />
    59. 59. Jaderstad<br />et al,<br />PNAS ‘10<br />
    60. 60. <ul><li> ALS / (?SMA)
    61. 61. Parkinson’s Disease (Nat Biotech ‘02; PNAS ‘07; Stem Cells ‘09)
    62. 62. Neurogenetic degeneration (Nat Med ‘07; Stem Cells ‘09)
    63. 63. Some aging-related degeneration
    64. 64. Spinal cord injury & Head trauma (PNAS ‘02; PNAS’10)
    65. 65. Stroke / Hypoxic-Ischemia (Nat Biotech ‘02; Exp Neurol ’06; PNAS’11)
    66. 66. Cerebellar Degeneration (J Neurosci ‘06; PNAS ‘06; PNAS’10)</li></ul>NSC-mediated‘rescue’/ ‘protection’ / ‘regeneration’/anti-inflammation-- complementing ‘cell replacement’Neural stem cells display an<br />
    67. 67. 50 µm<br />0.5 cm<br />o.<br />Cerebellar <br />Purkinje Cell Neuron<br />Degeneration<br />Mutants<br />Li et al, PNAS & J Neurosci 2006<br />
    68. 68. Calbindin<br />Purkinje Cell Layer evident following early<br />transplantation of<br />neural stem cells<br />
    69. 69. Intercellular Contacts of NSCs with Nervous Mutant Purkinje Neurons<br />bgal (NSC)<br />Calb (PN)<br />Dapi (nuclei)<br />Li, et al,<br />PNAS;<br />J. Neurosci.<br />(2006)<br />
    70. 70. Host Nr Purkinje Neurons (PNs) Remain At Nearly Wild Type Numbers<br />Following Early NSC Transplantation<br />Li, et al, PNAS; J. Neurosci. (2006)<br />
    71. 71. 70<br />60<br />y = 0.5032x - 4.8333<br />50<br />2<br />R<br /> = 0.688 (n=8)<br />P<0.05<br />40<br />30<br />Rotarod test (seconds)<br />20<br />10<br />0<br />0<br />20<br />40<br />60<br />80<br />100<br />Purkinje cell survival (%)<br />Positive Correlation between NSC-Rescued <br />Host PNs & Improved Motor Behavior <br />Li, et al,<br />PNAS;<br />J. Neurosci.<br />(2006)<br />
    72. 72. Influence of NSCs upon Nr PNs (via cell-cell contact) appears to be mediated by gap junctions (particularly those employing Cx 43)<br />Jaderstad et al,<br />PNAS ‘10<br />
    73. 73. Blocking Gap Junction Formation(with Cx43 RNAi)abrogatesNSC-mediated neuronal rescue<br />Jaderstad et al, PNAS (2010)<br />
    74. 74. WHAT ARE THE GAP JUNCTIONS DOING?<br />
    75. 75. <ul><li>There is an ideal amount of tissue plasminogen activator (TPA) Activity
    76. 76. TPA in its role as upstream of a number of signal transduction pathways
    77. 77. e.g., neurotrophic factor handling; mitochondrial function
    78. 78. Nervous mutant has abnormally high TPA levels
    79. 79. Abnormalities emulated by  TPA
    80. 80. NSC intercession (somehow) restores (lowers) level to normal</li></ul>A different way of viewing “gene therapy” -- homeostatic pressure to return an endogenous gene to equipoise -- not too high & not too low<br />Surviving Purkinje Neurons<br />(as % of wild type control<br />nr<br />NSCs<br />A logarithmic-parabolic relationship<br />between tPA activity & PN survival<br />in cerebellarorganotypic slice cultures<br />Li, et al, PNAS; J. Neurosci. (2006)<br />
    81. 81. Nervous<br />Cerebellum<br />NSC<br />tPA <br />Plasminogen<br />Plasmin<br />Mitochondrial<br />VDAC<br />Neurotrophins<br />BDNF, NT3<br />PKC<br />Energy <br /> Metabolism<br />Dendrite & Axon<br />Development<br />PN Survival<br />More Balanced<br />More Normal Motor Behavior<br />Li, et al, PNAS; J. Neurosci. (2006)<br />
    82. 82.
    83. 83. NSCs Restore/Improve Expression of Key Neurotrophic factors in Nr Cerebella<br />cultured NSCs<br />undifferentiated nsc <br />C<br />a<br />nt3/bgal/dapi<br />bdnf<br />undif.nsc<br />dif.nsc<br />b<br />BDNF<br />NT3<br />undifferentiated nsc<br />c<br />young<br />wt nr/nr nr+nsc<br />adult<br /> wt nr/nr nr+nsc<br /> trkc/dapi<br />trkb<br />BDNF<br />NT3<br /> wtnr<br />wt nr/nrnr + nsc<br />(downstream targets of TPA/plasmin<br />system)<br />A<br />EGL<br />BDNF/Calb / Dapi<br />NT-3/Calb / Dapi<br />B<br /> Wt nr/nrnr+nsc<br />BDNF<br />NT-3<br />NOTE: receptors notabnormal<br />Li, et al, PNAS; J. Neurosci. (2006)<br />
    84. 84.
    85. 85. wt<br />nr + NSC<br />nr/nr<br />256±63 nm diameter<br />274±82 nm diameter<br />585±95 nm diameter<br />Mitochondrial Morphological Abnormalities Reversed Following Early NSC Transplanation<br />
    86. 86. Nervous<br />Cerebellum<br />tPA Inhibitor<br />tPA<br />Plasminogen<br />Plasmin<br />Mitochondrial<br />VDAC<br />Neurotrophins<br />BDNF, NT3<br />PKC<br />Energy <br />Metabolism<br />Dendrite & Axon<br />Development<br />PN Survival<br />PKC Antagonist<br />More Normal Motor Behavior<br />Li, et al, PNAS; J. Neurosci. (2006)<br />
    87. 87. Jaderstad et al, PNAS (2010)<br />
    88. 88. Gap junction formation (Cx43) reciprocally between hNSC-derived neurons in cervical region of contused adult rat spinal cord associated with improved respiratory function<br />Jaderstad et al, PNAS (2010)<br />
    89. 89. Summary<br />• Direct cell-cell contact between stem cells & other neural cells, mediated by gap junction coupling, represents not only an early form of inter-cellular communication, establishing a template for more mature electrochemical synaptic interfaces, but also mediateshomeostasis-promoting actions (including metabolic/ mitochondrial)<br />• Exogenous stem cells may “rescue” endangered cells by this mechanism<br /> • Mechanismof rescue is not known<br />• Could involve the passage of beneficial molecules<br />• Could involve the containment of toxic molecules from inter-cellular spread<br />• Is likely a pervasive mechanism throughout the stem cell“world” & in development (& disease/therapy)<br />
    90. 90. Summary of<br />Stem Cell-Mediated Therapeutic Actions<br />TISSUE<br />(e.g., CNS)<br />Damage or Degeneration<br />Stem Cell Engraftment<br />(e.g., NSC)<br />Paracrine support by secreted factors<br />Replacement of lost Cells<br />Induction of beneficial [Ca++]i signaling patterns<br />Intercellular exchange of ions & molecules<br />RESCUE<br />REPLACEMENT<br />
    91. 91. Host<br />NSC<br />NSC<br />
    92. 92. Ischemia<br /> Tumor<br /> Trauma<br /> Neuronal degeneration<br /> Amyloid plaques<br /> Lysosomal storage diseases<br /> Demyelination<br />NSC<br />Pathology<br /><ul><li> Migration toward injury = 1st critical step</li></ul>in somatic stem cell engagement<br />during regeneration<br /><ul><li> In CNS, ? molecular mechanisms</li></ul>that mobilize & direct precise<br />homing of even distant NSCs,<br />thru parenchyma,<br />along non-stereotypical routes,<br />towards regions of pathology <br />where they might engage niches<br />harboring local reparative signals<br /><ul><li> NSCs seem to home similarly to:
    93. 93. These pathologic sites</li></ul> all have disparate<br /> etiologies<br /><ul><li> What do they have</li></ul> in common?<br />An Inflammatory Signature<br />Imitola et al, PNAS (2004)<br />
    94. 94. <ul><li>Proof of concept:
    95. 95. Examined role of prototypical inflammatory chemokinestromal-derived factor-1a(SDF-1a)</li></ul> in directing human NSC mobilization & homing<br /> in a representative CNS injury<br /><ul><li> unilateral focal cerebral stroke (hypoxia-ischemia)</li></ul> in mouse<br />NSC<br />CX<br />SDF1a<br />CXCR4<br />Pathology<br />Imitola et al, PNAS (2004)<br />
    96. 96. Host<br />NSC<br />Molecules<br />That effect<br />HOST NICHE<br />NSC<br />
    97. 97. <ul><li>IN VIVO
    98. 98. Neurotraumatic Conditions (“The Chaperone Affect”)
    99. 99. Stroke/hypoxic-ischemic injury
    100. 100. Spinal Cord Injury / Traumatic Brain Injury
    101. 101. Not just diffusible factors but cell-cell contact via gap junctions, as well
    102. 102. Improved “homing”/pathotropism
    103. 103. IN VITRO --“Disease in a Dish”
    104. 104. Modeling development, injury, disease, ?therapy </li></ul>• hESCs & HIPSCs into “neural tubes” ==> ultimately into hNSCs)<br />• neural-vascular obligate co-patterning/co-dependence – driven by neural crest<br />• high-throughput drug discovery<br />• large scale comprehensive profiling<br />
    105. 105. Stem Cells as “Instructional Tools”:<br />Models of <br />Embryogenesis,<br />Organogenesis, <br />Homeostasis-preserving Mechanisms <br />Intrinsic Developmental Programs<br />
    106. 106. PluripotentStem Cells as “Diseases-in-a-Dish”<br />Blastocyst<br />Blastocyst<br />(can be from a diseased embryo diagnosed<br />by PGD)<br />Embryonic stem (ES) cells<br />Neural <br />stem cell <br />(NSC)<br />Morphogens<br /> ECM<br />Fibroblasts<br />Transcription factors;<br />Proteins; Plasmids; RNA; <br />Small molecules; microRNA<br />Patient<br />skin <br />cells<br />Induced pluripotent stem (iPS) cells<br />
    107. 107. Stem Cells As In Vitro Models of Disease<br />“Disease-in-a-Dish”<br />* “diseased” cells, i.e., cells obtained from patients with particular diseases* “normal” cells that are stressed or perturbed in prescribed ways<br />- To study underlying mechanisms of disease- To identify drug targets diagnostics prognostics- To identify protective agents and/or mechanisms- To identify & test drugs or “leads”-to-drugs<br />hIPSCs particularly appealing for studying cell types that are difficult to obtain from a living patient<br />
    108. 108. -<br />-<br />Screening Drug-like Small Molecules, in a Robotic High-Throughput Manner, to Reveal Mechanisms, Identify Novel Therapeutic Targets, and/or Discover New Drugs<br />Add<br />compounds<br />Incubate<br />plates<br />Score hits on automated microscope<br />
    109. 109. High-content / High-throughputDrug Discovery <br />Proof-of-Concept:<br />Screening a random library in an unbiased manner for neuroprotective small molecules<br />IlyasSingec, Evan Snyder, Susanne Heynen, Michael Jackson<br />
    110. 110. In the fully developed human body plan, the vascular & nervous systems are (must) be co-patterned<br />How does that come about?<br />?An early program in embryogenesis that is sustained throughout development & growth<br />
    111. 111. Some Implications<br />Prototype of how 2 germ layers co-pattern (in an obligatory manner?) to give rise to the “real” body plan where lineages do not exist in isolation but must coordinate with each other<br />Ectoderm & mesoderm& endoderm all must talk to each other<br />“Cross-talk” as one of “Nature’s strategies” for organogenesis<br />Important to recognize when thinking about repair<br />?Help explain “mysteries” in “normal development”<br />e.g., brain vascularized before blood flow -- is this how?<br />Developmental anomalies -- ?Help explain associations in some syndromes<br />Neural with vascular / vascular with neural (e.g., Sturge-Weber? T-S? NF?)<br />Reinforces notion that “repair strategies” may need to reinvoke “developmental strategies”<br />When thinking about “regeneration” must be cognizant of “all lineages”<br />Many pathologic entities injure multiple systems -- e.g., stroke, trauma, infection, inflammation<br />
    112. 112. POTENTIAL ACTIONS of STEM CELLS in<br />NEUROTRAUMATIC DISORDERS<br /><ul><li>MECHANISMS INHERENT TO STEM CELL BIOLOGY
    113. 113. Trophic Support(endogenous or following genetic enhancement)
    114. 114. Neuroprotection(e.g., against oxidative stress; apoptosis)
    115. 115. Reducing inflammation
    116. 116. Restore intracellular mitochondrial-mediated homeostasis
    117. 117. Promote intercellular transfer of molecules via gap junctions
    118. 118. e.g., aid degradation of protein aggregates
    119. 119. Secrete or transfer other yet-to-be identified molecules
    120. 120. that might alter gene expression within damaged host cell
    121. 121. AS VEHICLES FOR EXOGENOUS GENE DELIVERY
    122. 122. ????CELL REPLACEMENT</li></li></ul><li><ul><li> Protection of established circuits may be as important & more tractablethan establishing new ones</li></li></ul><li>Conclusions<br />Stem cells can model developmental processes<br />Repair likely involves re-invoking developmental processes, particularly those that underlie plasticity & adaptation (exogenous as well as endogenous stem cells)<br />Entails multiple mechanisms & cell types (as well as cross- talk between these cell types:<br />Diffusible factors; cell-cell contact via gap junctions; altering niche (ECM; vasculature; toxins)<br />Disease & injuries that can be addressed by moleculartherapies or cell protection may be the “low hanging fruit” for stem cell-mediated interventions<br />Modeling “Diseases-in-a-Dish” for purposes of drug discovery= another near-term stem cell application<br />Addressing chronic/established injury remains a challenge<br />
    123. 123. Thanks<br />IlyasSingec Ted Teng<br />Eric Herlineus Richard Sidman<br />Andrew CrainHelen Blau & Renee Reijo-Pera<br />Brian TobeMassimo Pandolfo & SatyanChintawar<br />Kook In Park ZeweiHuang<br />NejmiDilmac David Cheresh<br />Mahesh Lachyankar Peter Black & XandraBreakefield<br />Jochen MaurerSteve Ashwal & Andy Obenaus<br />SaharNissim Jeff Macklis<br />Runquan Zhang Ralph Weissleder & Khalid Shah<br />Karen AboodySamiaKhoury & Jaime Imitola<br />Dustin Wakeman Jeff Rothstein & Bob Brown<br />Jean-Pyo Lee Bill Weiss & Anders Persson<br />Jitka & Vaclav Ourednik Gene Redmond & John Sladek<br />Jeff Lindquist Bob Langer & Erin Lavik<br />Xuejun (Jun) Parsons Fran Platt & Tom Seyfried<br />Franz-Josef Muller Ernest Arenas & Carmen Salto<br />Jianxue Li Tony Atala & Rene Yiou<br />

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