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Stem Cell：The Promise and The Challenge

ChiaChia--Cheng Chang, Ph.D.Cheng Chang, Ph.D.

Department of PediatricsDepartment of Pediatricsand Human Developmentand Human DevelopmentMichigan State UniversityMichigan State University

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Stem cells are undifferentiated cells Stem cells are undifferentiated cells with the capacity for unlimited or with the capacity for unlimited or

prolonged selfprolonged self--renewal and the ability renewal and the ability to give rise to differentiated cells.to give rise to differentiated cells.

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Differentiation can be defined as qualitative changes in the cellular phenotype that are a consequence of the onset of synthesis of new gene products, i.e. the non-cyclic changes in gene expression that ultimately lead to functional competence.

Bach, Renehan and PottenCarcinogenesis 21:469-476, 2000

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New Excitement About Stem Cells

Successful cultivation of Human embryonic Successful cultivation of Human embryonic stem cell lines.stem cell lines.Adult stem cells can differentiate into Adult stem cells can differentiate into developmentally unrelated cell types.developmentally unrelated cell types.

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Characteristics of Human ES Cells

• Normal Karyotypes• Express high telomerase activity• Express cell surface markers of primate embryonic stem cells• Maintained undifferentiated proliferation for 4-5 months and

developmental potential to form trophoblasts and gut epithelium (endoderm), cartilage, bone, smooth and striated muscle (mesoderm), neural epithelium, stratified squamous epithelium (ectoderm)

• Developed teratomas in immune-deficient mice

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ESES1. High malleability1. High malleability2. Potential for undesired2. Potential for undesired

development (development (teratomasteratomas))3. Infinite lifespan, 3. Infinite lifespan,

unlimited supplyunlimited supply4. High ethical burden4. High ethical burden

Uncertain legal statusUncertain legal status

Adult Stem CellsAdult Stem CellsLimited developmental potentialLimited developmental potentialBetter behaved, easier to manageBetter behaved, easier to manage

Lose their ability to proliferate/Lose their ability to proliferate/differentiate after a time in culturedifferentiate after a time in cultureLess moral ambiguityLess moral ambiguityLess legal controversyLess legal controversy

Advantage and Drawbacks of Embryonic Stem Cells and Stem Cells from the Adult

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Can Old Cells Learn New Tricks?

Stem cells found in adults show surprising versatility, but it’s not yet clear whether they can match the power of cells from embryos.

G. Vogel

Science 287:1418,2000

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Adult Stem Cells Can Differentiate into Developmentally Unrelated Cell Types

1.1. Neural stem cells produced myeloid and lymphoid cells as Neural stem cells produced myeloid and lymphoid cells as

well as early well as early hematopoietichematopoietic cells.cells.

2.2. Stem cells from mouse muscle could repopulate the blood Stem cells from mouse muscle could repopulate the blood

stream and rescue lethally irradiated mice.stream and rescue lethally irradiated mice.

3.3. Bone marrow stem cells can become brain cells and liver Bone marrow stem cells can become brain cells and liver

cell precursors, plus heart skeletal and smooth muscle.cell precursors, plus heart skeletal and smooth muscle.

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Location of Adult Stem Cells

HematopoieticHematopoietic —— yolk sac blood island, fetal liveryolk sac blood island, fetal liverand spleen, bone marrow, circulating umbilical cord and spleen, bone marrow, circulating umbilical cord BloodBloodLiver Liver —— portal zone near bile ductportal zone near bile ductIntestine Intestine —— cryptcryptEpidermal Epidermal —— basal layerbasal layerRetinal Retinal —— pigmented pigmented cilliary cilliary marginmarginBreast epithelial Breast epithelial —— end buds(Cap cells) and basal layer of end buds(Cap cells) and basal layer of

mammary glandmammary glandPancreas Pancreas —— pancreatic pancreatic islats islats and ductsand ductsMesenchymalMesenchymal —— bone marrow bone marrow stromastroma, adipose tissue, adipose tissue

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Retinal Stem Cell

•• Localized to Localized to ciliaryciliary marginmargin

•• Can differentiate into retinalCan differentiate into retinal--specific specific

neuronal and neuronal and glialglial progeny ( rod photoprogeny ( rod photo--

receptors, bipolar neurons and receptors, bipolar neurons and muller gliamuller glia).).

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Clonogenic stem cells in small and large intestine

Roberts et al., Radiation Res. 141:303-308, 1995

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Mouse Endbuds and Budding Structures Formed by Two Types of HBEC

In Vivo In Vitro

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Lobule Type 1 and Organoid Formed in Matrigel

In Vivo In Vitro

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Some Characteristics of Stem Cells

1. Undifferentiated cells with the capacity for unlimited or prolonged self-renewal that can give rise to differentiated cells.

2. Unipotent and multipotente.g. Intestinal stem cells produce one or more of the 4 classes of mature differentiated cells (absorptive, goblet, Paneth and enteroendocrine cells).

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Some Characteristics of Stem Cells3. Metaplasia-The formation of one differentiated cell

type from another.e.g. Patches of intestinal epithelium in the stomach; foci of hepatocytes in the regenerating pancreas.

4. Slow cycling in cell division.

5. Contact-insensitive;deficient in gap-junctional intercellular communication.

6. Specific gene expression.

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Specific Gene Expression in Stem Cells

• Human hematopoietic stem cells--CD34, Thy-1• Epidermal stem cells--β1integrin (α2β1, α3β1)• Small intestinal stem cells--β-catenin• Hair follicle stem cells--cytokeratin-15• Retinal stem cells--CHX10, Nestin• Neural crest stem cells--p75• Liver stem cells--AFP,GGT

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Methods for Isolating Stem Cells from Adult Tissue

1. Flow 1. Flow cytometrycytometryHematopoieticHematopoietic stem cellsstem cells----CD34CD34++, HLA, HLA--DRDR--

Neural crest stem cellsNeural crest stem cells----p75p75++

Epidermal stem cellsEpidermal stem cells----ββ1 1 integrinintegrin

2. 2. MicrodissectionMicrodissection/colony/colony--forming assayforming assayRetinal stem cellsRetinal stem cells

Hair follicle stem cellsHair follicle stem cells

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Methods for Isolating Stem Cells from Adult Tissue

3. Cell adhesion or lack of adhesion3. Cell adhesion or lack of adhesion----Human epidermal stem cells adhered most rapidly to Human epidermal stem cells adhered most rapidly to

Type IV collagen, Type IV collagen, fibronectinfibronectin or or keratinocyteskeratinocytes ECM.ECM.----Late attachment of human breast epithelial stem cellsLate attachment of human breast epithelial stem cells

on plastic.on plastic.

4. Contact insensitive growth on cell mat4. Contact insensitive growth on cell mat

Putative human kidney epithelial stem cells.Putative human kidney epithelial stem cells.

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Alternative Models for Stem Cell Deployment

1. 1. Invariant Asymmetry Invariant Asymmetry

2. 2. PopulationalPopulational AsymmetryAsymmetry

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Regulation of Stem Cell roliferation/Differentiation

β-integrin/ECM ----epidermal Required for cell survival and Keratinocyte differentiation through MAP Kinase.β-catenin(Tcf/Lef)----intestine-Tcf-null mice lack stem cells in the small intestine-Overexpression of β-catenin increases the proportion of stem cells in vitro and in vivo.TelomeraseHighly expressed in ES cells and adult stem cells

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Regulation of Stem Cell Proliferation/Differentiation

P21 cip1/waf1 (cell cycle inhibitor)Hematopoietic stem cell quiescence maintained by p21;bone marrow transplant from p21-1- mice leads to hematopoietic failure.MAP kinase signalingThe growth of undifferentiated ES cells was enhanced by culture in MAP kinase (MEK) inhibitor (PD 098059).bcl-2Transgenic expression increased HSC in BM.

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Regulation of Stem Cell Proliferation/Differentiation

CDK2CDK2A CDK2 inhibitor prevented hair loss A CDK2 inhibitor prevented hair loss caused by chemotherapy (caused by chemotherapy (etopsideetopside) in mice.) in mice.Oxidative stressOxidative stressLowered oxygen and antioxidant (NLowered oxygen and antioxidant (N--acetylacetyl--LL--cysteinecysteine) promote growth of CNS and ) promote growth of CNS and glialglial precursors respectively.precursors respectively.

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High Susceptibility of a Human Breast Epithelial Cell Type with Stem Cell Characteristics to Telomerase Activation and Immortalization.

Sun, W., K.S. Kang, I. Morita, J.E. Trosko and C.C. Chang

Cancer Res. 59:6118-6123, 1999.

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Clinical Applications of Stem Cells

1. 1. HematopoiesisHematopoiesis as a model of stem and progenitor cell as a model of stem and progenitor cell transplantation.transplantation.

■■ Fanconi Fanconi Anemia Anemia —— an an autosomal autosomal recessive disorder recessive disorder characterized by progressive bone marrow failure leading characterized by progressive bone marrow failure leading to to aplastic aplastic anemia.anemia.

■■ Bone marrow ablative doses of chemotherapy and Bone marrow ablative doses of chemotherapy and radiotherapy to eliminate endogenous cancerradiotherapy to eliminate endogenous cancer

〔〔leukemia (CML), lymphoma (Hodgkinleukemia (CML), lymphoma (Hodgkin’’s), multiple s), multiple myelomamyeloma, breast cancer, breast cancer〕〕. .

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Clinical Applications of Stem Cells

2. 2. Transplantation therapy for acute and chronic degenerative diseaTransplantation therapy for acute and chronic degenerative diseaseseEpidermalEpidermal----skin graft for patients with severe burnsskin graft for patients with severe burns

NeuronalNeuronal----ParkinsonParkinson’’s disease, multiple sclerosis and other s disease, multiple sclerosis and other neurodegenerative disease (brain neurodegenerative disease (brain gangliosidosisgangliosidosis, , demyelinatingdemyelinatingdisease).disease).LiverLiver----For Liver damage by drugs, toxins or viral infection.For Liver damage by drugs, toxins or viral infection.IsletIslet--for insulin non for insulin non ----producing pancreasproducing pancreasSkeletal muscle satelliteSkeletal muscle satellite----muscular dystrophies or muscle loss;heart muscular dystrophies or muscle loss;heart diseasediseaseMesenchymalMesenchymal——tissue engineering in plastic reconstructive surgerytissue engineering in plastic reconstructive surgery

3. Gene therapy3. Gene therapy

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Stem Cell Therapy for Parkinson's Disease

•• Parkinson's disease: slow, rigid or uncontrollable Parkinson's disease: slow, rigid or uncontrollable movement caused by the death of a particular group of movement caused by the death of a particular group of brain neurons that produce dopamine.brain neurons that produce dopamine.

•• Logical candidate for cell replacement therapy Logical candidate for cell replacement therapy ----Conventional treatments have had limited success.Conventional treatments have had limited success.----Grafting immature neurons from aborted human Grafting immature neurons from aborted human fetuses have had up to 50% reduction in patients' fetuses have had up to 50% reduction in patients' symptoms and the effects appear to last.symptoms and the effects appear to last.

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Stem Cell Therapy for Parkinson's Disease

•• Promise of ES cellsPromise of ES cells----Potential to provide large quantity of Potential to provide large quantity of nigralnigral

(dopamine) neurons with better quality control.(dopamine) neurons with better quality control.•• Challenge of ES cell applicationChallenge of ES cell application

----EEthical concern (better with adult stem cells).thical concern (better with adult stem cells).----Developing techniques to grow and to Developing techniques to grow and to

differentiate ES cells into dopamine neuronsdifferentiate ES cells into dopamine neuronsand their supportive cells.and their supportive cells.

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Caplan , A.I . And Bruder,S.P. Trends in Mol.Medicine 7 : 259~264, 2001

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Bianco,P.& Robey, P.G.Nature 414:118~121, 2001

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Bianco,P.& Robey, P.G.Nature 414:118~121, 2001

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Stem Cell Research and Ethics

•• Millions of patients may benefit from the Millions of patients may benefit from the applications of stem cells.applications of stem cells.

•• Ethics of embryo destruction are not addressed. A Ethics of embryo destruction are not addressed. A time for restraint to develop a national policy and time for restraint to develop a national policy and ethical framework.ethical framework.

Stem Cell Research ApplicationsStem Cell Research Applications: A society leaving : A society leaving the original Garden of Eden, or A great promise the original Garden of Eden, or A great promise not only for unexpected insight into biology but not only for unexpected insight into biology but ultimately for the alleviation of human suffering.ultimately for the alleviation of human suffering.