Characterization of porous scaffold materials for bone tissue

engineering - Saartje Impens -

Micro-CT symposium 31/05/07

GBE project

• 2 different aims:

1. Setting up a protocol for the healing of large and complex, but critical bone defects

2. High throughput screening of different scaffolds (= porous structure)

With the aid of micro-CT evaluation

1. Healing of critical bone defects

Bone defect

Operation Room

Patient own cells + growth

factors

Scaffold

Haeled bone defect

Bioreactor

Scaffold seeding and culturing with cells

In vitro

Cells + medium

Macrostructural & Mechanical parameters

Fluid Flow

input = cells

3D cell seeding

time point analysis

2D plates

3D cell culture

Optimization possible?

If 2D plates are possible

2D cell seeding

2D cell culture

µ-CT screening

3D scaffold

output = proliferation

differentiation

in vivo screening nude mice

Not Ok

Ok

Not Ok

Ok

input = material + coating +

growth factors

Toxicity testing

NoREJECT

Optimize scaffold

Yes

Further screening until clinical approvement

Macrostructural shortcoming

No perfusion perfusion possible

No

Yes

Clinical approved scaffold

Yes

Yes

No

No

2. High throughput screening

GBE strategy

• Multidisciplinary approach

Micro-CT use

1. Micro-CT based characterization of scaffolds– Calculate structural parameters– Calculate mechanical parameters with the aid of a

FE-model– Calculate fluid flow

2. Evaluation of bone formation in explanted scaffold

– Replacement of histology?

1. Scaffold characterization• Important parameters for bone formation in Matlab

– Porosity As high as possible (100%)– Specific surface area As high as possible

(Mentat) >3,95mm-1 (Ding et al. based on bone)

– Pore size 100-800µm (PorousAnalyser)

– Permeability As high as possible(PoreNet) > 10-8m2 (Kohles et al. based on

bone)– Interconnectivity As high as possible (100%)

• Mechanical parameters with FE-modeling (Mesh creation in Matlab) Expected load during walking is 1,2 x body weight – Strength 100% under yield strength– Stiffness 17-20 GPa (cortical bone)

10-1500MPa (trabecular bone)– Stretch on surface (500-)1500-4000µstrain

1. Scaffold characterization

Scaffolds Reconstructed micro-CT FE-mesh Image

1. Scaffold characterization

• Structural and biomechanical parameters

Scaffold characterization

• Extra important parameter for the GBE project– Fluid flow

• Nutrient & Oxygen transport

– Wall shear stress• May stimulate proliferation and differentiation

i.e. May stimulate bone formation

Ideally Computing Fluid Flow of micro-

CT based models

1. Scaffold characterization

• 2D Fluid flow on µCT based model

Scaffold: Ø 6 mm, L 8 mm

Inflow: 1 ml/min

Figures: Tim van Cleynenbreugel

• 3D Fluid flow on CAD-based model

1. Scaffold characterization

Figures: Silvia Truscello

1. Scaffold characterization

• Problems occur when meshing regular scaffolds produced by rapid prototyping

Blue Best

VioletPinkOrangeRed Worst

Manually remeshing

2. Substitute for Histology

• Evaluation different scaffold materials– Time consuming

• Embedding 2 weeks• Sectioning

– 1 scaffold/day– Labor intensive

• Staining – 1 day

• Analysis– 1 scaffold/day– Labor intensive

*

1. Scaffold characterization

2. Substitute for Histology

• Polymer scaffolds

Histologicalimage

Binarized histologicalSection

Interpolated micro-CT image

After registration Green: Overlap Blue: only histology Red: only micro-CT

2. Substitute for Histology• Distinguish between scaffold and bone by

thresholding?

Difficult, depends on scaffold material

Bone

Scaffold

Zone of bone ingrowth

2. Substitute for Histology

• Micro-CT analysis– Micro-CT Scanning

– Micro-CT scanning explant

– Positioning and subtrac- ting in Mimics to determine the amount of bone ingrowth

2. Substitute for Histology

Conclusion

• Micro-CT is a very useful tool for this type of research– Scaffold parameters can be calculated

• Prior to implantation• Non destructive

– Time consuming histology• Can be replaced• If necessary, histology can be performed after scanning

• If FE models and meshing problems are solved– Fluid flow – Wall shear stresses

can be calculated

AcknowledgementSpecial thanks goes to:

• Jan Schrooten• Tim van Cleynenbreugel• Barbara Neirinck• Silvia Truscello• Greet Kerckhofs

-Thanks-

-Thanks-