Stem Cells & Neurological Disorders Said Ismail Faculty of Medicine University of Jordan

Stem Cells and Neurological Disorders Outline: Introduction Types & Potency of Stem Cells Embryonic Stem Cells Adult Stem Cells iPSCs Tissue Engineering and Regenerative Medicine Stem Cells & Neurological Disorders: - Neural Stem Cells - Examples of Therapeutic Applications - Conclusion Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Introduction Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Benefits of stem cell research : Treatment of complex diseases: Chronic Disorders: Diabetes Neurological Disorders: Alzhimer’s Parkinson’s Spinal Cord Injuries Heart disorders: MI Regenerative medicine (Spare parts !) Skin Cartilage Bone Cornea Heart Valves Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Definition: stem cells: renew itself indefinitely (ii) differentiate to multiple tissue types A stem cell is not committed to a specific function until it receives a signal to differentiate into a specialized cell Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Types & Potency Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Embryonic: - Blastomere (4-5 day embryo) - Pluripotent Adult: - Adult tissue - multi or uni potent Other : - Fetal: - Aborted embryos - Umbilical: - Umbilical cord blood - Multipotent Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Potency: 1.Totipotent (Fertilized egg) Generate: - all embryonic cells and tissues - supporting tissue like placenta and umbilical cord 2. Pluripotent - Give rise to cells of all 3 germ layers (ecto-, meso-, and endoderm - Come from embryos and fetal tissue - Have active telomerase (maintain long telomers) 3. Multipotent - Give rise to multiple different cell types 4. Unipotent - Cell differentiating along only one lineage Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Embryonic Stem Cell Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders The Embryonic Stem Cell Source: 1. IVF embryos 2. Aborted Fetus 3. Therapeutic cloning Stem Cells and Neurological Disorders

IVF embryos Thousands of frozen embryos are routinely destroyed when couples finish their treatment.

Somatic Cell Nuclear Transfer The nucleus of a donated egg is removed and replaced with the nucleus of a mature, "somatic cell" (a skin cell, for example).

Stem Cells and Neurological Disorders Embryonic Stem Cell First isolated and cultured in 1998 From inner cell mass of blastocyst (4-5 day embryo). Pluripotent with long-term self-renewal Capable of unlimited number of divisions without differentiation Can essentially live forever without forming tumors Maintain normal diploid complement of chromosomes (stable karyotype) Telomerase activity Clonogenic: give rise to genetically identical group of cells Expresses transcription factor Oct-4 (+ or – genes needed for proliferative state) Spend most of their time in S phase In-Vitro: 300 population doublings Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Human Blastocyst showing Inner Cell Mass Stem Cells and Neurological Disorders

GROWING HESC IN VITRO:

Stem Cells and Neurological Disorders Advantages: Immortal: supply endless amount of cells Flexible: can make any body cell Available: IVF clinics Disadvantages: Hard to control their differentiation Ethics Immune rejection Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Avoiding Immune rejection: Genetically engineering stem cell to: a. Express MHC antigens of recipient b. produces stem cells with deleted MHC genes 2. Therapeutic Cloning: Clone somatic Cell nucleus of recipient into egg develop into blastocyst and isolate ES cells Such ES cells have recipient immunological profile Co-transplantation with Hematopoitic Stem cells Stem Cells and Neurological Disorders

Avoiding Immune rejection

Stem Cells and Neurological Disorders Laboratory tests to identify ESC : 1. Immortality: Sub-culturing stem cells for many months (long-term self-renewal) 2. Morphology: Inspecting culture by microscope (for undifferentiation) 3. Surface markers & Stemnss genes: (e.g. Oct-4) 4. Karyotype stability: Examining the state of chromosomes 5. Telomerase Activity 6. Pluripotency: testing differentiation potential into diff. cells types Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Ethics and ESCs: Here Here or Here When is it OK….when is it NOT Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Group of cells or Human life Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Adult Stem Cells Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders The Adult Stem Cell Undifferentiated cell found in a specialized tissue in adult. Capable of self-renewal Become specialized to cell types of the tissue from which it originated. Properties: Somatic Long-term self-renewal give rise to mature cell types Generate intermediate cell (progenitors) “committed” Can migrate whenever needed Uni- or Multipotent Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Sources of adult stem cells : Bone marrow Blood stream Umbilical cord blood Dental pulp of the tooth Cornea and retina Skeletal muscle Liver Skin (epithelia) Gastrointestinal tract Pancreas Brain & spinal cord Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Bone marrow Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders umbilical cord blood Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Dental Pulp Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Adult stem cell plasticity stem cell from one adult tissue can generate the differentiated cell types of another tissue: “unorthodox differentiation” or “transdifferentiation” EX. Hematopoietic stem cell Neurons Possible under specific conditions Stem Cells and Neurological Disorders

Plasticity of adult stem cells Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Advantages : No immune rejection Available: eg HSC Partly specialized: easier to control differentiation Flexible: under the right conditions Disadvantages : Scarce (Rare): True for many Adult SCs Unavailable: Some are difficult to isolate like Neural stem cells Vanishing: Don’t live in culture as long as ES cells Questionable quality: more prone to DNA abnormalities Stem Cells and Neurological Disorders

Induced Pluripotent Stem Cells Stem Cells and Neurological Disorders (iPSCs) Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Induced Pluripotent Stem Cells (iPSCs): = Retro-differentiation = Re-programming Producing stem cells from differentiated cells !!! Pluripotent embryonic like stem cells are produced Reversal of normal process Does Not require human embryos No donor…..No rejection Less expensive No Ethical issues Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Main Key Genes: iPSCs are derived from adult somatic cells by inducing expression of certain Stemness genes: (usually by viral vectors: risk !!!) - eg: Master transcriptional regulators: Oct-4 Sox2 Nonog - other genes: c-Myc (oncogene: cancer risk !!!!) Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Pluripotency: Believed to be identical to embryonic stem (ES) cells in many respects: - expression of certain stemness genes - chromatin methylation patterns - doubling time - embryoid body formation - teratoma formation - viable chimera formation - potency and differentiability Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Generation of induced pluripotent stem (iPS) cells Isolate and culture donor cells. (2) Transfect stemness genes into cells by viral vectors. Red cells express those genes (3) Harvest and culture the cells according to ES cell culture, on feeder cells (light gray) (4) A subset of the transfected cells become iPS cells and generate ES-like colonies    Stem Cells and Neurological Disorders

Neurogenesis of iPS Pluripotent Neuronal Stem Cells derived from Adult Leukocytes

Stem Cells and Neurological Disorders Potential target disorders for Stem Cell Therapy: Leukemia Heart damage Anemia Cornea damage Retinal damage Parkinson’s Alzhimer’s Diabetes Spinal Cord Injury Kidney Failure Skin grafts Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders leukemia Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Heart damage Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Diabetes Stem Cells and Neurological Disorders

Regenerative Medicine Stem Cells and Neurological Disorders Tissue Engineering & Regenerative Medicine Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Bone Repair Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Skin graft grown from stem cells Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Cornea Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders trachea from stem cells Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders A grown ear seeded with cartilage cells Stem Cells and Neurological Disorders

Neurological Disorders Stem Cells and Neurological Disorders & Neurological Disorders Stem Cells and Neurological Disorders

Which Stem Cell: Delivery Strategy: Graft type: 1. Neural stem cells 2. Other Adult SC (HSCs & MSCs) 3. Cord Blood SC 3. Embryonic SC 4. iPSCs Delivery Strategy: Injection into brain Into Blood stream (Homing + immobilization by cytokines) Graft type: Stem cells + Biomaterial Stem Cells + Gene therapy All have been shown to generate neural tissue (Adult SCs are the mostly used in clinical trials) Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Comparison: Stem Cell   Embryonic Pluirpotent Ethics Fetal Pluripotent Ethics Cord Blood Potent Rejection Available Adult Neural / Autologus Self low Numbers Same tissue Isolation Adult (HSCs, MSCs,…) Easy isolation rejection (if allo.) Easy culture Plasticity ?!! iPCs Pluripotent vector safety Self Stem Cells and Neurological Disorders

Ongoing clinical Trials in US and the world 2012 Sanberg et al. February 2012

Stem Cells and Neurological Disorders Different strategies for stem cell delivery to repair degenerated tissue Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Neural stem cells: Generate new neural cells throughout the lifetime Can migrate and replace dying neurons Give rise to all types of neurons, astrocytes and oligodendrocytes, … Capable of only Minor repairs Their activity is up-regulated following injury - Found in: - Sub-ventricular zone of lateral ventricles (Most neurogenic area) - Dentate gyrus of Hippocampus (2nd) fewer in: - Cerebellum    - Spinal Cord Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Therapeutic Applications: Main target disorders: - Parkinson’s: localized degeneration (in substantia nigra) easier cell therapy Huntington’s: clear etiology, single gene disorders (Gene/Cell Therapy) Alzheimer : damage is less defined, widespread neuro-degeneration Spinal Cord injuries: very promising prospects Other: - Multiple Sclerosis (Siatskas and Bernard, 2009) Ischemia / stroke Epilepsy (Naegele et al., 2010) Amyotrophic Lateral Sclerosis (ALS) (Wolfson et al., 2009). Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Parkinson’s: Main Strategy: - Replacing degenerated neurons with dopamine-producing cells Site: - Substantia nigra: area were most degeneration occurs in PD - Source of SCs: - Pieces of fetal midbrain tissue (Mendez et al., 2005) - Autologous adult neural stem/progenitor cells (Michel et al., 2009) - Embryonic SCs (Friling et al. 2009) Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Huntington’s: Good Model: well characterized single gene disorder Main Strategy: Blocking neuronal cell death & replacing lost neurons in striatum - Source of SCs: - SCs of fetal striatal primordium into striatum of HD patients (Bachoud-Lévi et al., 2006) - Autologous adult neural stem/progenitor cells (Yu and Silva, 2008; Visnyei et al., 2006). Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Alzheimer’s: Neuro-genesis in hippocampus deteriorates in AD patients Example approaches: (Lunn et al., 2011) 1. Implanting Neural Stem Cells: - Replace lost neurons - Delay degeneration by producing Brain-Derived Neurotrophic Factor (BDNF) 2. Nerve growth factor (NGF) production: - Genetically engineered patient fibroblasts that produce NGF …!!! - Integration of NGF fibroblasts into a major cholinergic center of the basal forebrain provided some benefit to AD patients Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Spinal cord injuries: (Salewski et al., 2010; Hu et al., 2010, Mathai et al 2008). Stem cells can: 1. Replace neurons that died from injury 2. Generate supporting cells to re-form the myelin sheath & stimulate re- growth of damaged nerves 3. Protect cells at injury site from further damage, by releasing protective factors Stem cells under trials: - Embryonic SCs - Umbilical cord SCs - Adult neural SCs - Mesenchymal / bone marrow SCs - induced pluripotent Scs Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Christopher Reeve 1952 - 2004 Stem Cells and Neurological Disorders

Paralyzed Patients Walking Again Stem Cells and Neurological Disorders http://www.youtube.com/watch?v=KGUAyKQKmmY http://www.youtube.com/watch?v=-kygF2leZCE http://www.youtube.com/watch?v=ZgI4tm8Tr5M Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders Conclusion: Very promising clinical trial results in the last few years More research needed to optimize diff. SC replacement protocols: - Cell type - Route - No. of cells - Single or multiple cell doses Choice between ESCs / ASCs / iPSCs: yet to be resolved Ethics (ESCs and Fetal tissue): Each Country has to decide Stem Cells and Neurological Disorders

Stem Cells and Neurological Disorders THANK YOU Stem Cells and Neurological Disorders