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 Yes No 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 No REJECT Optimize scaffold Yes Further screening until clinical approvement Macrostructural shortcoming No perfusion perfusion possible No Yes Clinical approved scaffold Yes 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  µm (PorousAnalyser) –Permeability  As high as possible (PoreNet) > m 2 (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  GPa (cortical bone) MPa (trabecular bone) –Stretch on surface  (500-) µstrain

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 Violet Pink Orange Red  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 Histological image Binarized histological Section 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

Acknowledgement Special thanks goes to: Jan Schrooten Tim van Cleynenbreugel Barbara Neirinck Silvia Truscello Greet Kerckhofs -Thanks-

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