frank baaijens et al. laboratory for tissue biomechanics and tissue engineering, department of...
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Frank Baaijens et al.Laboratory for Tissue Biomechanics and Tissue Engineering, Department of Biomedical Engineering, TU/e
Simon Hoerstrup et al.Laboratory for Tissue Engineering and Cell Transplantation, Clinic for Cardiovascular Surgery, University Hospital Zurich
Bert Meijer et al.Laboratory for Macro-Molecular and Organic Chemistry, Department of Biomedical Engineering, TU/e
Jan Feijen et al.Polymer Chemistry and Biomaterials, Department of Chemical Engineering, UT
Polymers for health carePolymers for functional tissue engineering
of cardiovascular substitutes
• All procedures that restore missing tissue in patients require some type of replacement structure.
• Traditionally: totally artificial substitutes, nonliving processed tissue, or transplantation.
• New alternative, tissue engineering: the replacement of living tissue with living tissue, designed and constructed for each individual patient.
• Cardiovascular substitutes market estimated at 80 B€.
Tissue Engineering (The Lancet)
Small diameter vascular graft
Tunica media
• Cardiovascular disease leading cause of adult death
• No synthetic vascular graft available for diameters < 6mm
Thrombogenicity
Neo-intima hyperplasia (excessive proliferation of SMCs)
external elastic lamina
smooth muscle cells
internal elastic lamina
endothelium
Aortic heart valve
Valve replacements
Artificial durability remarks
mechanical life-long trombogenic, noise
synthetic ? mechanical and hemodynamical behaviour ok
Biological
xenograft 7-10 yr Chemical fixation
allograft 7-10 yr Donor dependent
autograft > 15 yr Pulmonary valve transplant
No growth, no repair and adaptation to functional demands
• 300,000 heart valve replacements each year
• Open and close 100,000 times each day, 3 billion in a lifetime
TE valves: Chain-of-Knowledge
Implantation
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
Isolation of cells from vessels
Seeding in scaffold Culture, conditioning
Tissue formation
vsmc endothelial cells
Challenges
Create functional, living cardiovascular tissues: strong: collagen structure elastic: elastin network non-thrombogenic: endothelial lining three dimensional tissue architecture
external elastic lamina
smooth muscle cells
internal elastic lamina
endothelium
Role of scaffold
• Initial attachment of cells (shape)
• Supply the tissue with sufficient strength
• Bioactivity to control 3D architecture modulate proliferation and differentiation modulate ECM synthesis and degradation stimulate angiogenesis (vasculature)
time
scaffold degradation
ECM remodeling
load
bea
ring
prop
.
implantation ?
Tissue engineering of heart valves
• Successfully implanted at pulmonary site in juvenile sheep
• Not suitable for implantation at aortic site
• In-vivo tissue maturation takes 20 weeksHoerstrup et al., Circulation (2000)
Tissue engineered heart valve 6 weeks 16 weeks 20 weeks
• Optimal cell source?
• Design requirements of scaffold?
• What is optimal loading protocol in bioreactor for optimal tissue (collagen) architecture?
• What is mechanical load on tissue?
• How to test functionality of tissue-engineered valves?
How to improve strength?
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
Mechanical load on valve: systole
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
De Hart et al, J. Biomechanics (2003)
In-vitro testing of life tissue
Bioreactor: Physiological flow and pressure
Bio-prosthetic valve
MRI: velocity profiles in bioprosthesis
Rutten et al (2003)
MRI: velocity profiles in bioprosthesis
Rutten et al (2003)
Heart valve collagen orientation prediction
Driessen et al (2003) Diastole
Computational study of collagen synthesis, alignment and distribution in response to mechanical loading:
• Loading in closed configuration is optimal
• 10 % straining needed
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
Impact of cyclic straining on ECM
StaticScaffold Cyclic straining
600 10 20 30 40 500.00
0.05
0.10
0.15
0.20
Str
ess
(MP
a)
Strain (%)
Static
10 % straining (optimal)
A. Mol et al, Thorac. Cardiovasc. Surg., (2003)
Bioreactor design
Diastole is critical to obtain proper collagen structure
• Change of paradigm for in-vitro mechanical conditioning protocol: new bioreactor design
Mol et al, van Lieshout et al (2003)
Design requirements of scaffold
• ‘Trivial’: biocompatible, cell attachment, biodegradable, etc
• Elasticity: accommodate cyclic strains of order 10 %
• Strength: stresses of order 1 MPa
• Bioactive to control tissue architecture
• Degradation: both fast (~ 2 weeks) and slow (~ 20 weeks)
• Bio-mimicking: appropriate micro-environment
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
Collagen structure in arterial wall
Bioactive scaffolds
Building blocks: PGA, PCL, PTMC, etc
• PGA (‘golden’ standard)
fast degradation (~ 2 weeks)
brittle
• PCL
slow degradation (> 20 weeks)
elastic, ductile, strong
• PTMC
enzymatic in-vivo degradation
elastic, strong
surface erosion: controlled drug release
Meijer et al, Feijen et al (2003)
Bioactive scaffolds
Building blocks: PGA, PCL, PTMC, etc
Bioactive supramolecular polymer
ureido-pyrimidinone (UPy) polymers
UPy-GRGDS & UPy-PHSRNUPy-GRGDS UPy-PHSRN
UPy-GRGDS
UPy-PHSRN
Synergistic effect on cell-attachment
Dankers et al (2003)
Electro spinning of bio-mimicking scaffolds
PCL scaffold 1 week culture 2 weeks, confluent
Vaz et al (2003)
Multiple layers for site specific bioactivity
ECM organization: 6-12 months!
6 weeks
20 weeks
time
scaffold degradation
ECM remodelling
load
bea
ring
prop
.
implantation
PGA
Hybrid scaffold
PGA+PCL
• First, successful, trial with bone marrow derived mesenchymal stem cells
• Electrospinning of strong, elastic and bioactive scaffolds
• New bioreactor design and loading protocol, extensive in-vitro studies in Zurich and Eindhoven
• In-vitro testing capabilities
• Animal studies in Zurich in progress (pulmonary)
• First human implantation, upon successful completion of animal and in-vitro tests, in pediatric age group
Summary & Outlook
Cells Scaffold(Mechanical)preconditioning
Tissue formation,matrix remodelling
Implantation/Model system
Acknowledgements
• Core DPI program BioPolymers R-0d
• TU/e ‘Bio-Initiative’ grant
Hybrid scaffold for vascular graft
• Slow formation of elastin > aneurysm
Porous, elastic support
• Neo-intima hyperplasia
Compliance matching
• Thrombogenicity
Confluent endothelial lining
Fast degradation
Slow degradation
Elastic support
‘Golden standard’: Coated PGA scaffold
Deformation PGA/P4HB
0
10
20
30
40
50
60
0 2 4 6 8 10 12 14 16
Applied strain (%)
Def
orm
atio
n (%
of a
pplie
d st
rain
)
Biocompatible +
Cell attachment +
Highly porous (98 %) +
Complex shapes -
Mechanical strength -
Elasticity -
Bioactivity -
Elastic biopolymer: TMC
TMC
Low Tg
in-vivo degradable
cross-linked: no-creep
Example: Scaffold for vascular graft
Inner layer P(TMC)
Particulate leaching
Pore size: 1-10 m
Outer layer P(TMC-CL) (10:90)
Fiber winding
Pore size: 20-60 m Feijen, Grijpma
DPI Biopolymers for TE program
Hybrid Scaffolds
Baaijens et al. TU/e
Supramolecular Bioactive Polymers
Meijer et al. TU/e
Elastic TMC
Feijen et al. UT
DPI
Biopolymers for Medicine
Effect of mechanical conditioning
0 10 20 30 40 50 600.00
0.05
0.10
0.15
0.20
Str
ess
(MP
a)
Strain (%)
Control Stretched
Cyclic straining results in :
more pronounced and organized tissue formation
increased load-bearing properties
trend towards cell orientation parallel to the applied strain
tissue strength/stiffness proportional to strain magnitude
0
50
100
150
200
250
DNA GAG HP
%
*** *
StaticMax. 7% strain
Max. 9% strain
Max. 10% strain
A. Mol et al, Thorac. Cardiovasc. Surg., (2003)