腦中風 後 的 恢復與治療

1. 腦中風後的恢復與治療

2. Carbon14-dated age of cardiomyocytes Cardiomyocytes renewed with a gradual decrease from 1% turning over annually at the age of 25, to 0.45% at the age of 75. Fewer than 50% of cardiomyocytes are exchanged during a normal life span Young at heart Evidence for cardiomyocyte renewal in humans Bergmann et al. 2009, Science

3. Nuclear weapons and neurogenesisno more cortical neurons in normal adults How about after stroke ? Bhardwaj et al. 2006, PNAS

4. Barriers restrain CNS regeneration • Neuronal death • Glial cells inhibit nerve growth • Most neural stem cells are constrained • Few exceptions in lower vertebrates and in olfactory pathway and hippocampus in mammals

5. Plasticity of adult human brain Ward NS et al., Brain, 2003

6. Spontaneous changes in brain after stroke Cramer, 08, Annals of Neurology

7. Consequence of hypoxia-ischemia • Features of energy metabolism in brain • High metabolic rate • Limited intrinsic energy stores • Dependence on aerobic metabolism of glucose • Within seconds, ATP level fall, lactic acidosis  ion pumps dysfunction (Na/K ATPase)  rundown of transmembrane ion gradients  membrane depolarization  voltage-sensitive ion channels open  K efflux after a min, Na, Ca ion gradient lost and influx  Ca overload enhance glutamate release  cell death

9. Ischemic apoptosis in parallel with necrosis • Deprivation of growth factor support • Addition of NGF or bFGF reduce injury • Oxidative stress • Antioxidants: glutathione, superoxide dismutase (SOD)1 • Prolonged deficits in energy metabolism • Increased inflammatory cytokines • IL-1, TNF, TGF • Transgenic mice overexpressing bl2 had smaller infarcts

10. Penumbra Lo, 2008

11. Recent therapeutic developments of stroke • Prevention • Acute treatment • rt-PA thrombolysis • even for minor stroke (Gonzales et al. 2006) • Neuroprotection • Ebselen, anti-inflammatory selenocomound mimic glutathione • NXY-059, antioxidant nitrogen spin trap • Stem cell transplantation • G-CSF, endogenous stem cell-mobilizing agent • phase I for acute MCA stroke by Shyu et al., 2006 • Rehabilitation • Transcranial magnectic or direct current stimulation • Hummel & Cohen 2006 Review

12. tPA for brain attack ! • In US, ~ 50% pt arrive at ER < 1h In Taiwan, ~ 50% pt arrive at ER < 4h • NINCDS trial in 1990s: 13% more favorable • Symptomatic ICH rate (no protocol violation) ~4%; (violation) ~8% • < 3% of patients currently treated in US < 2% of patients currently treated in Taiwan • Increase likelihood of return to normal by 30%

13. Carotid endarterectomy or stent angioplasty • Severe (> 70%) carotid stenosis in symptomatic patients • Moderate (50-69%) stenosis: depend on age, comorbidities, symptoms • For asymptomatic patients, only recommend for > 80% carotid stenosis

14. Peripheral nerve grafts (Schawann cells) • Olfactory ensheathing glia • Stem/ progenitors cells • Adult bone marrow mesenchymal stem cells • Induced pluripotent stem (iPS) cells from skin fibroblasts • key pluripotency genes Oct-4, SOX2, c-Myc, Klf4 • Endogenous neural progenitor cells Types of cell therapy for CNS diseases

15. Critical issues for stem cell therapy • What is a good source of stem cells for neural repair ? (effective & obtainable) • What factors stimulate stem cells to home (migrate) to sites of injury? • What cues are needed for stem cells to differentiate into the desired cell type? • Limited graft survival • Avoid transplanting undifferentiated cells that cause tumors (purification)

16. “Astrocyte-like” stem cells in subventricular zone Ependymal cells Slowly dividing GFAP+ Johansson et al., Cell, 96:25-34, 1999. Doetsch et al., Cell, 97:703-16, 1999.

17. Endogenous neuronal replacement after stroke in adult rats No neurogenesis in the cortex! Arvidsson et al., Nat Med 2002

18. Expanding endogenous stem cells • Adult NG2+ oligodendrocyte precursors • generate functional neurons and glia in vitro • After selective elimination of cortical neurons, some endogenous neural precursors proliferated and incorporated BrdU, migrated to the injury site, most are glia • Infusion of growth factors • Epidermal growth factor • Sonic hedgehog • Antagonize inhibitory micro-environment

19. De-differentiation: reprogram to youth • Irreversible differentiation and germ layer restriction can be broken by extracellular cues, especially following injury and in cell culture • First evidence of de-differentiation in vertebrate cells came from studies of urodeles and avians • Regenerated limb following limb amputation in urodeles (Brockes 1997, Brockes & Kumar 2002) • Newly formed lens after resection in salamander (Eguchi & Kodama 1993, Stone 1967)

20. De-differentiation in mammals • Schwann cells de-differentiated into precursors, proliferate, and redifferentiate after peripheral nerve injury (Brockes & Kumar 2002) • Oligodendrocyte precursor cells can produce neurons, astrocytes, and oligodendrocytes in vitro(Kondo & Raff 2000) • Precursor-to-stem cell conversion • In intestinal crypt (Marshman et al. 2002) • EGF convert transit-amplifying precursors in the adult brain into stem cells (Doetsch et al. 2002)

21. Bone marrow stem cells not restricted to the blood cell lineage • Adult bone marrow stem cells (BMSC) give rise to cells of all three germ layers, including neural cells in vitro • Some transplanted BMSC in the brain had characteristics of macrophage/microglia, astrocytes, neurons? • Tissue injury may recruit BMSC to additional cell types • mdx muscular dystrophy mice: BMSC myocytes

22. Immunosuppressive properties of mesenchymal stem cells Nauta and Fibbe. 2007, Blood

23. CNS entry of peripherally injected umbilical cord blood cells is not required for neuroprotection in stroke Borlongan et al., 2004, Stroke

24. Brain cells may be derived from non-neural stem cells ? • “Transdifferentiated” cells from one primary germ lineage to another ? • Bone marrow SC give rise to astrocytes • Umbilical cord SC form neurons and glia • Adult NSC reconstitute hematopoietic system • Controversies concerning plasticity of bone marrow and hematopoietic SC (Castro et al., 2002; Wagers et al., 2002) • Few differentiated cells present in the graft, most derived from cell fusion; most suggest trophic supply that facilitate endogenous repair process

25. Neural stem cells differ from those in other systems • Unlike hematopoietic system, the nervous system is largely formed during early development • Unlike epithelial system, neurons form intricate connections over long distances

26. Neuron-like morphology? • Reevaluation of in vitro differentiation of bone marrow stromal cells: Disruption of actin cytoskeleton induces rapid morphological changes and mimics neuronal phenotype (Neuhuber et al., 2004) • It’s unclear if neurons can be differentiated from any source other than neural stem cells and ES cells in vivo

27. Grafted neural stem cells develop into functional pyramidal neurons Englund et al., 2002

28. Induced Pluripotent Stem Cells (iPS) • Promising alternative of embryonic stem cells (ES), Overexpression of OCT4, SOX2, KLF4 and C-MYC, or Nanog and Lin 28(Takahashi and Yamanaka, 2006; Okita et al., 2007; Wernig et al., 2007; Yu et al., 2007) • Chimera or differentiate into 3 germ layers (Park et al., 2007; Takahashi et al., 2007) • Disease-specific iPS provide resources for disease investigation and drug development (Park et al., 2008) • iPS cells from patients with ALS can be differentiated into motor neurons (Dimos et al., 2008) • Transplantation of neural precursor cells derived from iPS cells: functional integration in the fetal rat brain, enhance functional recovery in Parkinsonism rats (Wernig et al., 2008)

29. A fresh look of iPS Yamanaka, 2009, Cell

30. Mechanisms of cell therapy-mediated recovery • Trophic factors and reduce death of host cells • VEGF, GDNF, FGF, BDNF • Increased neovascularization • Attenuation of inflammation • T- cells inhibition • Induce host plasticity • Neurogenesis • Recruitment of endogenous progenitors • Neuronal replacement and functional integration

31. Adverse effects of cell therapy • Cell-related: • Tumorigenic • Graft rejection • Allodynia (Hofstetter et al. 2005) • Procedure-related: • Intracranial hemorrhage during stereotactic transplantation without general anesthesia • Infection

32. Post-stroke neuroplasticity Swayne et al., Cerebral cortex, 2008 Corticospinal excitability decreased at the lesion Weakened intracortical inhibition and facilitation in the ipsilesional hemisphere

33. Strategies of brain stimulation after stroke Hummel and Cohen, 2006, Lancet Neurology

34. Short-term effects (minutes) • Shifting ionic balance • Short-term synaptic plasticity • Neurotransmitter release • Long-lasting effects (hours) • Long-term depression (LTD) and potentiation (LTP) • Induced gene expression • Regulation of postsynaptic receptors (ex: AMPA, NMDA, GABA) After-effects of repetitive transcranial stimulation Ridding & Rothwell, Nature Rev, 2007