MUSCULOSKELETAL CENTER OFMUSCULOSKELETAL CENTER … · 2015-09-26 · • UMass Memorial...
Transcript of MUSCULOSKELETAL CENTER OFMUSCULOSKELETAL CENTER … · 2015-09-26 · • UMass Memorial...
MUSCULOSKELETAL CENTER OFMUSCULOSKELETAL CENTER OF EXCELLENCE
STEM CELLS: OPPORTUNITIES AND STEM CELLS: OPPORTUNITIES AND CHALLENGESCHALLENGES
• Center for Stem Cell Biology & Regenerative Medicine• UMass Memorial Musculoskeletal Center of ExcellenceUMass Memorial Musculoskeletal Center of Excellence
STEM CELLS: OPPORTUNITIES AND CHALLENGES
• Biology of stem cells and applications for treatingBiology of stem cells and applications for treatinghuman disease
• UMASS initiatives in developing stem ll b d thcell-based therapy
STEM CELLS: OPPORTUNITIES AND CHALLENGES (Cont’d)
• Biology of stem cells and applications for treatingBiology of stem cells and applications for treatinghuman disease
• UMASS initiatives in developing stem cell-based ththerapy
What is a Stem Cell?What is a Stem Cell?
• What is a Stem Cell?
• What are different types of stem cells?• What are different types of stem cells?
• What is the promise of stem cell research?
S C ll Wh h ?Stem Cells: What are they?
D l f H E bDevelopment of a Human Embryo
Totipotent Pluripotent Lineage Commitment
Single-cellembryo
3-dayembryo
5-7dayembryo
4-weekembryo
6-weekembryoembryo embryo embryo embryo embryo
Isolation of Embryonic Stem Cells
Properties of Embryonic Stem CellsProperties of Embryonic Stem Cells
Isolated from an early stage embryo
Exhibit normal karyotypean early stage embryo
Express Markers Pluripotent
Th P i f S C ll R hThe Promise of Stem Cell Research
Why Stem CellsWhy Stem Cells…
Ti li f S C ll R hTimeline of Stem Cell Research
1956First successful bone marrow transplant
1981Embryonic
1998Human embryonic stem cells isolated at Univ. Wisconsin
Embryonicstem cells are isolated from
blmouse blastocysts
2007iPS cells first developed
T f S C llTypes of Stem Cells
• Adult Stem Cells
• Embryonic Stem Cells (hESC)
• Somatic Cell Nuclear Transfer (SCNT) Stem Cells
• Induced Pluripotent Stem (iPS) Cells
Ad l S C llAdult Stem Cells
Where have adult stem cells been found?
Ad l S C ll ( ’d)Adult Stem Cells (cont’d)
Derivation Method Isolation from adult tissues.
AdvantagesSuccessful treatments demonstrated.
Stem cells can be matched to patient
Limitations
Cells not found in all tissues.
Produce a limited number of cell types.
Difficult to identify, isolate and grow.
Current UsesResearch.
Current UsesSome in clinical use (e.g. bone marrow)
Ethical Concerns None to date.
Lineage Committed Stem Cells S B F iSupport Bone Formation
(Development and Tissue Regeneration)
OCHistoneFibronectinCollagenc-Fos/c-JunTGFβ1
APBSPCollagenFra2/JunB
OCOPCollagenase
BAXp53
ProliferationMatrix
Maturation Mineralization
Osteopontin p53c-FosMsx2
Self RenewalLineage
Commitment
BMPTGFβ
BMPTGFβ
Vitamin DTGFβ
GlucocorticoidVitamin D
BMP
Stem Cell Osteoproginator Pre-Osteoblast Osteocyte
E b i S C llEmbryonic Stem Cells
Human Embryonic Stem Cells (hES Cells)
H ES ll l
Human Embryonic Stem Cells (hES Cells)
Human ES cell colony on mouse fibroblast
feedersHuman Blastocyst
feeders
Inner Cell Mass (ICM)
hES C ll ( t’d)hES Cells (cont’d)
Derivation MethodRemoval of cells from Inner Cell Mass (ICM) of blastocyst embryo from IVF.
Differentiate into all somatic cell typesAdvantages
Differentiate into all somatic cell types.
Excess of IVF embryos exist.
Limited number of lines available for federally funded
Limitationsresearch.
Risk of tumors (teratomas) from transplanting undifferentiated cells
Current Uses
Research.
Currently no clinical trials for therapeutic use.
In development for drug screening and toxicity testing
Ethical ConcernsDestruction of embryo.
Donor consent requiredDonor consent required.
D i ti f i d d Pl i t t St C llDerivation of induced Pluripotent Stem Cells
Induced Pluripotent Stem CellsInduced Pluripotent Stem Cells
Reprogramming & Correcting Genetic Defect
mm
ing
rogr
amRe
pr
iPS CellsiPS Cells
Derivation MethodReprogramming of somatic cells by introduction of specific regulatory factor genes.p g y g
Advantages“ES cell – like” characteristics.
Stem cells can be matched to patientStem cells can be matched to patient.
Unknown if cells can differentiate into all cell types.
Risk of tumors (teratomas) from transplantingLimitations Risk of tumors (teratomas) from transplanting undifferentiated cells and from expression of introduced genes.
Current Uses Research.
Ethical Concerns None to date.
Comparative Properties of Stem CellsComparative Properties of Stem Cells
• Embryonic Stem Cells: Indefinite growth; Potential to differentiate into any cell typeto differentiate into any cell type
• Lineage-Specific Stem Cells: Limited growth;• Lineage-Specific Stem Cells: Limited growth; Potential to differentiate into defined cell types
• Cancer Stem Cells: Indefinite growth; Cannot differentiate into a defined cell typedifferentiate into a defined cell type
Why so many types of stem cells?
hi l C C ll/ i il bili
Because none are perfect.
Ethical Concerns
Immune Rejection Concerns
CancerConcerns
Cell/Tissue Types
Availability
hESC Yes Yes Yes All Many lines
SCNT ES cells Yes Yes Yes All? No
Adult Stem Cells No No No Limited
Many(limited types)
iPS Cells No No Yes All? Many
UMASS Initiatives in Developing Stem Cell-Based Therapy
A Collaboration of:
• UMASS Center for Stem Cell Biology & Regenerative MedicineRegenerative Medicine
• UMASS Memorial Musculoskeletal• UMASS Memorial Musculoskeletal Center for Excellence
UMASS Contributions to two Rate-LimitinggParameters of Stem Cell Control
• Mechanisms that regulate rapid and infinite• Mechanisms that regulate rapid and infinite proliferation of stem cells
• Mechanisms that establish and maintain lineage commitment following differentiationcommitment following differentiation
Genetic and Epigenetic RegulationGenetic and Epigenetic Regulation
• Genetic Control: DNA-encoded Regulatory Information.
Epigenetic Control: Regulatory Information for cell structure, function and specificity NOT encoded in DNA that IS transmitted to progeny cells during cell division.
Genetic and Epigenetic RegulationGenetic and Epigenetic Regulation
• Genetic Control
Genetic and Epigenetic RegulationGenetic and Epigenetic Regulation
• Epigenetic Control Histone CodeHistone Code
Genetic and Epigenetic RegulationGenetic and Epigenetic Regulation
• Epigenetic Control Regulatory Factor “Architectural Signature” Regulatory Factor Architectural Signature
Transcriptional Suppression
HDACHDAC YAPYAP
Basal Basal –– Suppressed (Differentiated)Suppressed (Differentiated)
HLHHLH** RUNXRUNX C/EBPC/EBP
APAP 11 DLXDLXAcAcAcAcRUNXRUNXAPAP--11
HDACHDAC YAPYAPYAPYAPRUNXRUNX
mRNAmRNASite ASite A Site BSite B Site CSite C
OCOC--BoxBoxTATATATA
C/EBPC/EBPEE--BoxBox
VDREVDRE
RUNXRUNXAPAP--11 DLXDLXRUNXRUNXAPAP 11RUNXRUNX
Maximal Expression
H ATH AT H LHH LH TFIIBTFIIB TAF’STAF’SV DR/RX RV DR/RX R HATHATAcAcA cA cAA
V itam in D V itam in D -- Enhanced (D ifferentiated)Enhanced (D ifferentiated)
A cA cAcAc
p
H ATH AT H LHH LH**
TFIIBTFIIBR N A R N A PolPol IIII
TAF S
TFIIDTFIIDRU NXRU NXm R N Am R N A
S ite AS ite A S ite BS ite B S ite CS ite CO CO C --Bo xBo x
TATATATAC /EBPC /EBP
EE --BoxBoxVD REVD RE
RU NXRU NX C /EBPC /EBPA PA P --11 D LXD LXRU NXRU NX
A PA P --11AcAcAcAcA cA cA cA c A cA c A cA c
M h i th t t l id d i fi it• Mechanisms that control rapid and infinite proliferation of stem cells
• Mechanisms that establish and maintain lineage it t f ll i diff ti ticommitment following differentiation
The Mammalian Cell CycleThe Mammalian Cell Cycle
An Abbreviated Cell Cycle in Human yEmbryonic Stem Cells
Dynamics of Combinatorial Organizationy gand Assembly of Regulatory Machinery in
Nuclear Microenvironments
RNA P l II M di t d R l t M h i f C ll C l (Hi t ) GRNA Pol-II Mediated Regulatory Mechanisms for Cell Cycle (Histone) Gene
Expression:
G1 G1/S S G2
• Mechanisms that control rapid and infinite proliferation of stem cells
• Mechanisms that establish and maintain lineage commitment following differentiation
Lineage Specific Retention of RegulatoryLineage-Specific Retention of RegulatoryMachinery for Cell Fate Determination
During MitosisDuring Mitosis
• Post-mitotic gene expression requires restoration of nuclear organization and assembly of regulatorynuclear organization and assembly of regulatory complexes.
• Two models: Degradation re-synthesis andDegradation, re synthesis, and
assembly/organization Retention and partitioningRetention and partitioning
Runx2: A Transcription FactorInvolved in Lineage Commitment and
Required for Osteogenesis in vivo
PluripotentPluripotentMesenchymalMesenchymal CellCell
Tendon CellsTendon CellsFibroblastsFibroblasts
??
PPARPPARγγ
MyoDMyoDMyotubesMyotubes
Runx2Runx2?
Runx2Runx2
Mature OsteoblastsMature OsteoblastsHypertrophicHypertrophic
ImmatureImmatureChondrocytesChondrocytes
AdipocytesAdipocytes
PPARPPARγγ
Runx2Runx2
Mature OsteoblastsMature OsteoblastsHypertrophicHypertrophicChondrocytesChondrocytes
Matrix ProteinsMatrix ProteinsMatrix ProteinsMatrix Proteins
Contributions of Runx Factors to Genetic and Epigenetic Control: Scaffolds for Spatial Organization of Promotor Regulatory Complexes (P/DNA and P/P)
Basal Basal –– Suppressed (Differentiated)Suppressed (Differentiated)Transcriptional Suppression
XHDACHDAC YAPYAP
pp ( )pp ( )
HLHHLH** RUNXRUNX C/EBPC/EBP
AP 1 DLXDLXAcAcAcAcRUNXRUNXAPAP 11
HDACHDAC YAPYAPYAPYAPRUNXRUNX
Transcriptional Suppression
y, 3
3: 3
84:7
491
282-
9445
X mRNAmRNASite ASite A Site BSite B Site CSite C
OCOC--BoxBoxTATATATA
C/EBPC/EBPEE--BoxBox
VDREVDRE
RUNXRUNXAPAP--11 DLXDLXRUNXRUNXAPAP--11RUNXRUNX
Bioc
hem
istr
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ol, 1
9:Bi
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hem
, 2
HLHHLH TFIIBTFIIB TAF’STAF’SVDR/RXRVDR/RXR HATHATAcAc
Vitamin D Vitamin D -- Enhanced (Differentiated)Enhanced (Differentiated)
AcAcAcAc
Maximal Expression
al (1
994)
B99
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007)
J B
HATHAT HLHHLH**
TFIIBTFIIBRNA RNA PolPol IIII
TAF’STAF S
TFIIDTFIIDRUNXRUNXmRNAmRNA
Site ASite A Site BSite B Site CSite C TATATATAEE--BoxBox
RUNXRUNX C/EBPC/EBPAPAP--11 DLXDLXRUNXRUNX
APAP--11
HATHATAcAcAcAcAcAcAcAc AcAc AcAc
AcAcAc
onte
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aved
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Site ASite A Site BSite B Site CSite COCOC--BoxBox
TATATATAC/EBPC/EBP
EE BoxBoxVDREVDREM J a G
u
Re organization of Regulatory MachineryRe-organization of Regulatory MachineryDuring Mitosis
INTERPHASE MITOSIS
Young et al (2007) Nature, 445: 442
Runx2 Remains Stably Associated withRunx2 Remains Stably Associated withMitotic Chromosomes
Young et al (2007) PNAS, 104: 3189
Chromosomal Association of Runx2 is AllelicChromosomal Association of Runx2 is Allelic
Runx FociRunx Foci
Young et al (2007) Nature, 445: 442
A Multi Dimensional Approach to UnderstandA Multi-Dimensional Approach to Understand Stem Cell Biology and Therapeutic Potential
• Stem cells exhibit a shorter cell cycle: sustainedStem cells exhibit a shorter cell cycle: sustained growth and unrestricted availability.
• Cell fate, lineage commitment and maintenance require the combined contributions of genetic andrequire the combined contributions of genetic and epigenetic mechanisms in nuclear “microenvironments” during andmicroenvironments during and following cell division.
Challenges and Opportunities of g ppStem Cell Research
• Moral and ethical considerations
• Immuno compatibility
• Procedure-related compromises (e.g., insertionalmutagenesis, tumorigenesis)
• Retention of genetic basis for diseases
Success Stories with Stem CellsSuccess Stories with Stem Cells
• Bone Marrow Transplantation- Cancer
Hemotological Diseases- Hemotological Diseases
• Cardio-vascular Stem Cells
• Neurological Stem Cellsl- Spinal Cord Injury
- Adult Macular Degeneration- ALS- ALS
• Skeletal Reconstruction
Acknowledgements
Members of the UMASS Center for Stem Cell Biology &
Acknowledgements
Members of the UMASS Center for Stem Cell Biology & Regenerative Medicine
Members of the UMASS Memorial MusculoskeletalCenter of Excellence
Janet L. SteinJane B. LianJane B. Lian
Andre van WijnenKlaus A. Becker
Prachi GhuleMatthew Mandellive
Daniel YoungDaniel YoungSayyed Kaleem Zaidi