stem cell therapy for vascular disease cells research.pdf · 2013-02-20 · stem cell therapy for...
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Stem Cell Therapy for Vascular Disease
Qingbo Xu, MD, PhDCardiovascular DivisionKing’s College London
China-UK Forum in Biomedical Innovation 17-21 March 2008
*Background
*Stem cells
*Cardiovascular Disease
*Stem cell repairing/therapy
ProteomicsManuel Mayr
Stem Cells and Vascular Diseases
Cell biologyLingfang Zeng
Animal ModelYanhua Hu
Clinic/humanStefan Kiechl
Proteomics workflowSample Prep
Image analysis
Sample labelling
2-D Separation
Automated spot picking
Automated spot digestion
Automated MALDI spotting
Mass spectrometry
Image acquisition
Endothelial cells
Stem cells
SMCs
Adult and embryonic stem cell culture and differentiation into vascular cells
Mouse models established in our laboratory
• Spontaneous atherosclerosis
• Endothelial injury of artery
• Vein graft atherosclerosis
• Heart infarction
• Heart transplantation
With MoviesFor SurgicalProcedures
Human/clinic studies, e.g.Bruneck Study
-Brief introduction about our work
-Background about stem/progenitor cells
-Human studies
-Mechanistic studies on stem cell differentiation
Derivation of the ES cell lines. The ES cell lines were generated from the blastocysts. These cells were isolated by immunosurgeryand plated on the MEF feeder layer. ES cells can be propagated in the undifferentiated state while being cultured on top of the MEF feeder layer. When they are cultivated in suspension as 3D cell aggregates (EBs), the ES cells differentiate into specialized cells.
Egg Sperm
Day 7
Progenitor cell mobilisation and homing
Stem cellpool
in bloodAorta
Adipose
BM Spleen
Liver
Intestine
SMP EPC
AS
Xu Q. Circ. Res. 2008
*Background
*Stem cells
*Cardiovascular Disease
*Stem cell therapy
Endothelial cells
Smooth muscle cells
Stem cells
Atherosclerosis - a vascular blocking diseaseTreated with a balloon catheter, i.e. angioplasty
CoronaryArtery
Treated with a balloon catheter, i.e. angioplasty
Endothelial cells
Repairing with stem cells
Endothelial cell lost
*Background
*Stem cells
*Cardiovascular Disease
*Stem cell therapy: Mechanism of cell differentiation
Attached “progenitors”A B C
D EWild-type 2 months ApoE-/- 2 months ApoE-/- 18 months
ApoE-/- 18 months
Endothelial cells
Stem cells
SMCs
What is the mechanism for stem cell differentiation into vascular cells?
?
?
Flow Shear Induces Stem Cell Differentiation:Model & Hyppothesis
Xu Q. Curr. Opin. Lipidol. 2007.
Shear stress
VEGF receptor
Endothelial markers
Akt
eNOSHDAC3
p53
p21
NO
HoxA
Smooth muscle markers
HDAC7
PI3K FAK
HDAC7 splicing
MEF2C
PDGF-BB Collagen IV
Receptor-ββ
Flow Shear Induces Stem Cell Differentiation toward Endothelial Cells
Static/H.E
Shear/H.E
Shear/X-gal
Zeng et al. J. Cell Biol. 2006
In vitro In vivo matrigel
Flow Shear Induces Stem Cell Differentiation toward Endothelial Cells
Flk-1
eNOS
α−tubulin
ES ST SH ST SH
12h 24h
CD31
CD133
CD144
Flk-1
Flt-1
β-actin
ESC Sca1+
C ST SH C ST SH
Zeng et al. J. Cell Biol. 2006
Flow Shear Induces Stem Cell Differentiation:Impact of HDAC3
Flt-1
p53K317
α-tubulin
p53
HDAC1
p21
st sh st sh st sh st sh
CTL CTLsi HDAC1si
st sh st sh st sh st sh
CTL CTLsi HDAC3si
Flt-1
p53K370
α-tubulin
p53
HDAC3
p21
Flow Shear Induces Stem Cell Differentiation:Model & Hyppothesis
Xu Q. Curr. Opin. Lipidol. 2007.
Shear stress
VEGF receptor
Endothelial markers
Akt
eNOSHDAC3
p53
p21
NO
HoxA
Smooth muscle markers
HDAC7
PI3K FAK
HDAC7 splicing
MEF2C
PDGF-BB Collagen IV
Receptor-ββ
HDAC7 and SMC differentiation
HDAC7
SMA
Calponin
α-tubulin
105KD
42KD
34KD
50KD
0 1 3 5 7 9 mSMC
Days of ES/Col IV
HDAC7 SMA Merge
HDAC7 Calponin Merge
HDAC7 and SMC differentiation
1.0
2.0
3.0
4.0
SMA-Luc SM22-Luc
R L
U
2.0
4.0
6.0
8.0
SMA-Luc SM22-Luc
R L
U
Mock
HDAC7
HDAC7-1
HDAC7-2
Mock
HDAC7
HDAC7-1
HDAC7-2
**
** *
*
HDAC7 and SMC differentiation
Adenovirus-HDAC7-1
0 2 5 10 20 MOI
HDAC7
SMA
α-tubulin
Calponin2.0
4.0
LucSM22
PBSPDGF
ctlsi HDAC7siR
L U
siRNA-HDAC7-1
Endothelial cells
Stem cells
SMCs
ConclusionsHDAC3 is crucial for stem cell differentiation into endothelial cells, while HDAC7 is for SMCs
HDAC3
HDAC7
Can progenitor cells differentiate to endothelial cells in vivo in response to flow shear?
Shearing stem cells in vivo
Isolation of sca-1+ progenitors
Apply the cells to denuded artery
Endothelial phenotype
Flow shear Induces Stem Cell Differentiation:Endothelial cell therapy
β-gal CD31X-gal
Zeng et al. J. Cell Biol. 2006
Flow Shear Induces Stem Cell Differentiation:Sca-1+ cell therapy
Xiao et al. ATVB 2006
Replace blocked vessels with a normal vessel, e.g. saphenous vein
blocked vessels
Normal vessel Vein graft atheroma
Endothelial cells
Stem cells
SMCs
Stem cell differentiation
Mouse model of vein bypass graft atherosclerosis
Carotid artery
Vessel segment
QuickTime™ and aYUV420 codec decompressor
are needed to see this picture.
stem cell-derived vessel
Ongoing work: replacing diseased artery with stem cell-derived vessel
Summary -Two types of stem cells: embryonic and adult cells
-VEGF plays a crucial role of stem/progenitor cells homing to the damaged vessels
-Signal pathways for endothelial differentiation involving in HDAC3-p53/p21
-Stem cell-derived endothelial cells can repairing damaged vessels in vivo
Innovation & Collaboration
New drug discovery to direct stem cell differentiation into endothelial cells, e.g. HDAC7 inhibitor or HDAC3 enhancer
Biomaterials for vascular production
Stem cell therapy in patients
Prof Q Xu Dr M Mayr Mr Z Zhang Dr G Pula
Dr A Zampetaki Dr X YinMs X Bai Ms F Zhang Dr Q Xiao
Dr L Zeng
Dr A Margriti
Mr D MartinDr U Mayr Dr Y Hu
Mrs S King Ms A E Pepe
Ms A Sidibe
Ms Z Luo
Dr E De Falco
Dr J P KirtonDr H LiMs M Prokopi Mr S Alam
Dr T N Tsai