Introduction Of Stem Cell - University Of Cincinnati

Introduction of Stem CellMeifeng Xu, MD, PhDDepartment of Pathologyand Laboratory Medicine

1Stem CellsStem cells are unspecialized and capable of dividingand renewing themselves for long periods throughcell division. In addition, they can give rise tospecialized cell types through differentiation.

Unique Properties of Stem Cells1. Self-renewal: the ability to go throughnumerous cycles of cell division toproduce more stem cells.2.Differentiation: these cells candifferentiate into specialized cell types.Stem cell self-renew and differentiationA: stem cellsB: progenitor cellsC: differentiated cells

Progenitor CellsA progenitor cell likes a stem cell, has a tendency todifferentiate into a specific type of cell. They are in the“intermediate status” between stem cells and fullydifferentiated cells.1) progenitor cell is already more specific than a stem celland is pushed to differentiate into its "target" cell (e.g.endothelial progenitor cells).2) stem cells can replicate indefinitely, progenitor cells candivide only a limited number of times.3) they can be activated by damaged or dead cells. Growthfactors or cytokines can mobilize them toward thedamaged tissue.

Differentiation PotentialTotipotent can differentiate into all possible cell types.These cells are produced from the fusion of an egg andsperm cell, e.g. zygote formed at egg fertilization.Pluripotent are the descendants of totipotent cells and candifferentiate into nearly all cell types, e. g., embryonicstem cells.Multipotent can differentiate into those of a closely relatedfamily of cells, i.e. bone marrow stem cells.Unipotent can produce only their own, but have theproperty of self-renewal, which distinguishes them fromnon-stem cells, e.g., muscle stem cells.

Differentiation PotentialAll cells in the bodyCells of all 3 germ layersBone marrow stem cellsMultipotentMultiple cell types of aclosely related familyCommitted ProgenitorsOnly one cell type

Stem CellsIn mammals, there are two broad types of stem cells:embryonic stem cells which are isolated from the innercell mass of blastocysts, and adult stem cells which arefound in various tissues.Human embryonic stem cell colonyBone marrow stem cells

Embryonic Stem CellsEmbryonic stem (ES) cells are the cells of the innercell mass of a blastocyst, an early-stage embryo.Pluripotent Can contribute to all three germ layersDerived from blastocysts Derivation of ES cells from mouse blastocysts (1981) Human ES cells (Thomson, 1998)Blastocysts produced by in vitro fertilization (IVF)

The Nobel Prize in Physiology or Medicine 2007for their discoveries of principles for introducing specific genemodifications in mice by the use of embryonic stem cellsMario R. CapecchiBorn: 6 October 1937,Verona, ItalySir Martin J. EvansBorn: 1 January 1941,Stroud, United KingdomOliver SmithiesBorn: 23 June 1925,Halifax, United Kingdom9

ES Cells (human) culture

ES Cells (mouse) CultureLIF: Leukemia InhibitoryFactorRemoval of LIF pushes stemcells toward differentiation.

Utilizations of ES Cells Pluripotent stem cells have shown potential in treating anumber of varying conditions, including but not limitedto: spinal cord injuries, age related maculardegeneration, diabetes, neurodegenerative disorders(such as Parkinson's disease), AIDS, etc. Tissue/organs derived from ESCs can be madeimmunocompatible with the recipient. Embryonic stem cells can also serve as tools for theinvestigation of early human development, study ofgenetic disease and as in vitro systems for toxicologytesting.12

eGFP Transgenic Mice

Adverse Effect of ES Cells1. The ES cell controversy is the ethical debatecentered only with research involving the creation,usage, and destruction of human embryos.2. The major concern with the possibletransplantation of ESC into patients as therapies istheir ability to form tumors including teratoma.

Induced Pluripotent Stem CellsInduced pluripotent stem cells (iPScells or iPSCs) are artificially derivedfrom the non-pluripotent cells byforcing to express the specific genes(Oct-4, c-Myc, Kif4, and Sox2).iPS cells are first produced in 2006(Yamanaka et al. Cell) from mousecells and in 2007 (Yu et al, Science)from human cells.iPS cells are similar to natural pluripotent stem cells (e.g., ES cells),such as the expression of certain stem cell genes and proteins,doubling time, embryoid body formation, potency ofdifferentiation, and teratoma formation.

The Nobel Prize in Physiology or Medicine 2012In 2012, Dr. Yamanaka was awarded theNobel Prize in Physiology or Medicine forhis discovery that adult somatic cells canbe reprogrammed into pluripotent cells. Byintroducing the genes for four factors, heinduced the skin cells of adult mice tobecome like embryonic stem cells, whichhe called induced pluripotent stem (iPS)cells. This iPS cell technology represents anentirely new platform for fundamentalstudies of developmental biology.

Adult Stem CellsAdult stem cells are undifferentiated cells, found throughoutthe body among differentiated cells in a tissue or organ afterdevelopment.They remain in a quiescent or non-dividing state for yearsuntil activated by disease or tissue injury. They can divide orself-renew to generate a range of cell types from theoriginating organ to replenish dying cells and regeneratedamaged tissues or entire original organ.Most adult stem cells are lineage-restricted and are generallyreferred to by their tissue origin, but they can alsodifferentiate to become other cell types.

Adult Stem CellsAdult stem cells are undifferentiated cells which are foundthroughout the body after development. Most adult stem cellsare multipotent or progenitors.

Sources of Adult Stem Cells1) The bone marrow stroma contain mesenchymal stem cells(MSCs), also called marrow stromal cells.2) Adipose tissue (lipid cells), which requires extraction byliposuction.3) Blood is drawn from the donor (similar to a blooddonation), passed through a machine that extracts thestem cells and returns other portions of the blood to thedonor.4) In addition, stem cells can also be taken from umbilicalcord blood, amniotic fluid, adult muscle or the dental pulpof deciduous baby teeth.

2Stem Cell TherapyRegeneration of damaged tissue andorgan by stem cells or the therapeuticgenes carried by stem cells.

Selection of Stem Cells1. The capacity of differentiation: Embryonic stem cells can become all cell typesof the body. Adult stem cells are limited to differentiating into the cell types oftheir tissue of origin.2. Grow in culture: Embryonic stem cells can be grown relatively easily in culture.Adult stem cells are rare in mature tissues, so isolating these cells from anadult tissue is challenging.3. ES cells, injected directly into another body, ES cells will differentiate into manydifferent types of cells, causing a teratoma.4. Immune rejection: Identical matches between donor and recipient must bemade for successful transplantation treatments, but matches are uncommon.5. Ethical considerations: many nations currently have limitations on eitherhuman ES cell research or the production of new human ES cell lines. The useof adult stem cells does not require the destruction of an embryo.21

Clinic Sources of Stem Cell TherapyCell typeAbbreviation OriginBone marrow derived stem cellBMSCBone marrowSkeletal myoblastSMAdult skeletal muscleCardiomyocyte progenitorcell/Cardiac stem cellCMPCAdult or fetal heartEndothelial progenitorcell/endothelial precursor cellEPCBone marrow/peripheralbloodEmbryonic stem cellESCBlastocyst stage embryosInduced pluripotent stem celliPSCAny somatic cell22

Source of Stem Cells Allogenic Donor with matchingtissue type Immune systemsuppression therapy Autologous Extraction and storage ofstem cells from patient(bone marrow or adiposetissue) Risk of graft-versus-hostdisease & infection Lower risk ofinfection/rejection23

Stem Cell Therapyhttp://en.wikipedia.org/wiki/Stem cell treatments24

Stem Cell TherapyThe effect of stem cells includes: protecting risky cells,regenerating damaged tissues or organs, replacing lostcells, stimulating growth of new blood vessels torepopulate damaged tissue.Therapy success is highly dependent on: survival oftransplanted cells in the recipient; integration within thetargeted tissue and restoration of function; proliferationand differentiation in a site-specific manner.25

HSC TransplantationHSC transplantation is the transplantation of multipotent HSC,usually derived from bone marrow, peripheral blood, orumbilical cord blood. It may be autologous, allogeneic, orsyngeneic.It is most often performed for patients with certain cancers ofthe blood or bone marrow, such as multiple myeloma orleukemia.However, HSC transplantation remains a dangerous procedurewith many possible complications, including infection andgraft-versus-host disease. It is reserved for patients with lifethreatening diseases.1

Myocardial RegenerationSeveral clinical trials targeting heart disease have shown thatadult stem cell therapy is safe, effective, and equally efficientin treating old and recent infarcts. Stem cell therapy fortreatment of myocardial infarction usually makes use ofautologous bone marrow stem cells (a specific type or all).However other types of adult stem cells may be used, suchas adipose-derived stem cells. Adult stem cell therapy fortreating heart disease was commercially available in at leastfive continents at the last count.27

Cells Being Tested for Heart TherapyRangappa, et al, J Cardiovasc Pharmacol Ther. 2010, 15:338-43.28

3Mechanism1. (Trans)-differentiationCytoprotection2. Paracrine Effect AngiogenesisRecruiting other cells

Transdifferentiation of Adult Stem Cells

Transdifferentiation of Stem CellsStem cells (MSCs) were GFP (green) and co-cultured withcardiomyocytes (CM, red) for 3 days. MSCs are beating with CM.Contrast (MSC CM)CM (red)MSC (Green)Merge (MSC CM)Li. et al. Cytotherapy. 2011;13:1057-65

Immunostaining of Co-cultured CellsGreen—stem cells; Red---CM (α-actinin positive)α-actininGFPMerge50 mXu, et al. Circulation 2004;110:2658-6532

Mechanism--Paracrine Effect3. 2Stem cells secrete paracrine factors into theimmediate extracellular environment. These factors thentravel to nearby cells to result in a response, through1)activating a specific receptor, and2)triggering a biochemical chain of events inside the cell.

Stem Cells Secrete Paracrine FactorsMirotsou, et al., JMCC, 2011; 50:280-9.

Mechanism of Paracrine EffectCytoprotection2. Paracrine Effect AngiogenesisRecruiting other cells

Hypoxia CdMLDH#p 0.05 vs Normoxia* p 0.05 vs HypoxiaNormoxia Hypoxia Hypoxia CdM% of Normoxia% of NormoxiaHypoxiaNormoxiaMSCCdM Decrease Hypoxic InjuryMTS#p 0.05 vs Normoxia* p 0.05 vs Hypoxia#*Normoxia Hypoxia Hypoxia CdMYu et al.: Plos One

Capillary-like Tube Formation37

Capillary-like Tube FormationHUVECsCdMCONHUVECs (labeled withPKH67)CdMNullCdM Conditioned mediumCdMGATA-4Li, et al. AJP. 2010;299:H1772-81

Administration of Stem Cell Therapy1. Introduce locally2. Cell Patch3. Intravenously4. MobilizationNiall. et al. J. of Cardiovasc. Trans. Res. 2012; 5:713–2639

Stem Cell Mobilization1. Some cytokines and chemokines can promote stem cellsmobilization.2. The damaged cells or tissues release some cytokines orchemokine which can mobilize stem cells to start repairprocession.3. Mobilized stem cells highly expressed cardiac transcriptionfactors, e.g. GATA-4 and anti-apoptotic proteins. Therefore,mobilized cells are more beneficial for repair of damagedtissue.4. Mobilizing stem cells is an optimal way for repairing thetissues or organs which are difficult to directly transplantstem cells.40