stem cell types , their maintenance and homeostasis
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Manifestation of Novel Social Challenges of the European Union in the Teaching Material of Medical Biotechnology Master’s P rogrammes at the University of Pécs and at the University of Debrecen Identification number : TÁMOP-4.1.2-08/1/A-2009-0011. - PowerPoint PPT PresentationTRANSCRIPT
Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011
STEM CELL TYPES, THEIRMAINTENANCE AND HOMEOSTASIS
Dr. Péter Balogh and Dr. Péter EngelmannTransdifferentiation and regenerative medicine – Lecture 2
Manifestation of Novel Social Challenges of the European Unionin the Teaching Material ofMedical Biotechnology Master’s Programmesat the University of Pécs and at the University of DebrecenIdentification number: TÁMOP-4.1.2-08/1/A-2009-0011
TÁMOP-4.1.2-08/1/A-2009-0011Sources and types of stem cells: different origins and developmental spectraES: • Embryonic stem cells from the ICM (inner cell mass)• Primordial Germ Cells (PGCs) → Embryonic Germ (EG)
cells iPS: non-embryonic somatic cells developed by the introduction of specific key transcription factors: Oct4, Sox2, c-myc, Klf4 MSC: mesenchymal stem cells present in bone marrow, adipose tissue, umbilical cord blood, amniotic fluid, placenta, dental pulp, tendons, synovial membrane and skeletal muscle, capable of self-renewal and differentiation in vitro into a variety of cells of the mesenchymal lineages such as osteoblasts, chondrocytes, adipocytes and myoblasts
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Sources of embryonic stem cells (ESCs)
Morula Early blastocyst
Inner cell mass(ICM)
Late blastocyst
Epiblast
Egg cylinder stage
Primitiveectoderm Germ cell lineage
Trophectoderm
Blastocyst cavity
Primitiveendoderm
Parietalendoderm
Visceralendoderm
Extraembryonic ectoderm
Somatic cell lineagesEctodermMesodermEndoderm
Proamnioticcavity
Oct3/4
Cdx2
Gata6
Nanog
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Stem cell sources in the mouse embryo• Preimplantation embryo: inner cell mass
(ICM) of the blastocyst (early blastocyst stage).
• Late blastocyst stage: formation of epiblast • Postimplantation embryo: formation of
primitive ectoderm with restricted pluripotency → the germ cell lineage and somatic lineages of the embryo.
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Characteristics of ES cells• Derivation from the preimplantation or periimplantation embryo• Prolonged undifferentiated proliferation, • Stable developmental potential to form derivatives of all three
embryonic germ layers even after prolonged culture• EC cells: teratocarcinoma-derived pluripotent embryonal carcinoma
cells generating cells of all three germ layers
Cartilage(mesodermal)
Intestinal glands(endodermal)
Epidermis(ectodermal)
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Cell membrane markers for ESCs
Sia
Gal
Glc
Man
GlcNAc
GlcA
IdoA
Fuc
Xyl
GalNAc
Tra 1-60 (KSPG)
NG2 and 473HD (CSPG)
Lewis X
PSA-NCAM
CD34
SSEA-3SSEA-4
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Structure of glycoantigens characteristic for ES cells• SSEA-3 and SSEA-4: 5–6 monosaccharides
attached to a ceramide lipid tail, forming the globoseries glycosphingolipids GL-5 and GL-7, their expression is reduced upon differentiation.
• The TRA (tumor rejection antigens) TRA-1–60 and TRA-1–81 keratan sulfated proteoglycan (KSPG) epitopes , probably associated with podocalyxin, a heavily sialylated membrane protein structurally similar to CD34.
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Characteristics of CD-defined antigens for ES cells• CD34: HSC/endothelial shared antigen
expressed hemopoietic stem cells/progenitors
• CD133: Five transmembrane domain cell-surface glycoprotein, expressed by neural stem cells
TÁMOP-4.1.2-08/1/A-2009-0011Main regulatory mechanisms of stem cells – external and internal effectsExternalInteractions with the matrix proteins, soluble factors and other cell types in stem cell niches, direct interactions with ECM proteins, complex signaling feedback from adjacent ESC niche cells (stromal/differentiated). InternalTF network regulating pluripotency or differentiation
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Stem cell niches in various organs
Germarium of the ovary
Terminalfilament
Cap cell
Cystoblast
BL
GSC
Inner sheathcell SSC
16-cellcyst
Folliclecells
Eggchamber
The apex of the testis
Hub cells
BLSSCGSC
Spermatogonia
The subventricular zone (SVZ) of the brain
NeuroblastAstrocyte
Lateral ventricle
BV
BL
Transit-amplifying
Ependymal cells
The bone marrow
Bone marrow
Osteoblast
Stromal cell
Multipotent SCHSC
LymphoidMyeloid
The crypt of an intestinal villus
Enteroendocrine cells Villus
Goblet cells
Crypt
BL
Transit amplifyingStem cells
Paneth cells
The bulge of the hair follicleHair shaft
BL
Hairbulb
MatrixDermal papilla
MuscleSebaceou
sglandBulge SC
MeiosisSpermatocytes
GonialblastCyst cell
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Stem cell environment – examples for stem cell niche• Germanium region of the ovary and the apex
of the testis (germ-line stem cell and somatic stem cell)
• Subventricular zone in the brain (neural stem cell)
• Bulge of hair follicle (epithelial stem cell)• Crypt of intestinal villi (endodermal stem cell)• Bone marrow (hemopoietic stem cell)
TÁMOP-4.1.2-08/1/A-2009-0011Multiple interactions involved in stem cell homeostasis
ESC regulators
Oct4 Nanog Tbx3
Wnt signaling
Tcf3 Tle1 Fzd5
Epigenetic regulators
Jarid2 Phc1 N-myc
RNA bindingprotein
Dppa5
Telomereassociated
Rif1
Tumorsuppressor
Trp53bp1
Oct4Nanog Tcf3
Pluripotency Differentation
Oct4
Nanog
Tcf3
Oct4
Oct4
Sox2Sall4
GCNFLRH1
TÁMOP-4.1.2-08/1/A-2009-0011Antagonistic regulatory circuits between differentiation and pluripotency• ESC/iPS regulation – hierarchic transcription factors• Wnt signaling• Epigenetic regulators• RNA binding• Telomere associated effectors• Tumor suppression• Cell cycle regulation
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mRNA regulation of stem cell gene expression
Other factors
Oct4Sox2 Nanog
mRNAs
AAAAAAAAAA
Alternatively spliced mRNAs
AAAAAAAAAA
Intergenic spliced mRNAs
AAAAAAAAAA siRNAs?
Other RNAs?
miRNAs
Intergenic transcripts
Antisense transcripts
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TF regulation forself-renewal/differentiation• Oct3/4, Nanog, Sox2, Stat3: maintenance of
proliferation• Cdx2: Inhibitory cross-interaction with Oct3/4
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Reprogramming: Induction of pluripotency in iPS cells
Target genes
Epigenetic modifiers
Transcription factorsSox2Oct3/4 Klf4
c-Myc
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Reprogramming: Lineage shift in differentiated cells• Reprogramming of B-cell lineage into macrophages –
role of C/EBPa• Induction of neuronal commitment from fibroblasts –
Ascl1, Brn2 and Mytl1
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Sequential maturation and regeneration of pluripotency
EctodermMesodermEndodermPluripotent cell
Pluripotent cell
Ectodermprogenitor
Neuronalprogenitor
Mature neuron
Pluripotent cell
TÁMOP-4.1.2-08/1/A-2009-0011Differentiation-associated commitment and reversibilityDifferentiation is coupled with• commitment and loss of
pluripotency/transdifferentiation capacity BETWEEN lineages
• Requirement for continuous stimulation for promoting specification WITHIN a lineage.
Reversal: Introduction of iPS-associated multilineage differentiation is associated with LOWERING of pleiotropic induction requirement and ELEVATION of differentiation signal threshold
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Summary
• Depending on their origin and developmental spectra, stem cells are quite heterogeneous, where their homeostasis is determined by their (a) endogenous programming with various levels of regulating gene expression and (b) external factors, including cytokines and adhesion proteins binding to extacellular matrix an other cell comprising the stem cell niche.
• Stem cell commitment and differentiation are not irreversible, as differentiated cell can be modulated to regain multipotency.