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Micro-CT symposium 31/05/07 Characterization of porous scaffold materials for bone tissue engineering - Saartje Impens - 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 Scaffold Scaffold seeding and culturing with cells Cells + medium Bioreactor Bone Patient own + growth defect cells factors Operation Haeled bone Room defect In vitro 2. High throughput screening input = material + coating + growth factors Optimize scaffold Toxicity testing Yes Not Ok If 2D plates are possible REJECT Clinical approved scaffold No Ok 2D plates Optimization possible? 3D scaffold Yes No Macrostructural & µ-CT screening Mechanical parameters Fluid Flow Macrostructural shortcoming Further screening until clinical approvement Not Ok Yes input = cells Ok 2D cell seeding 3D cell seeding 2D cell culture 3D cell culture time point analysis No No perfusion perfusion possible in vivo screening nude mice output = proliferation differentiation No Yes 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 – Specific surface area (Mentat) – Pore size As high as possible (100%) As high as possible >3,95mm-1 (Ding et al. based on bone) 100-800µm (PorousAnalyser) – Permeability (PoreNet) – Interconnectivity As high as possible > 10-8m2 (Kohles et al. based on bone) 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 – Stiffness – Stretch on surface 100% under yield strength 17-20 GPa (cortical bone) 10-1500MPa (trabecular bone) (500-)1500-4000µstrain 1. Scaffold characterization Scaffolds Reconstructed micro-CT Image FE-mesh 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 microCT based models 1. Scaffold characterization • 2D Fluid flow on µCT based model Inflow: 1 ml/min Scaffold: Ø 6 mm, L 8 mm Figures: Tim van Cleynenbreugel 1. Scaffold characterization • 3D Fluid flow on CAD-based model 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 Binarized histological Section Histological image 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? Bone Scaffold Zone of bone ingrowth Difficult, depends on scaffold material 2. Substitute for Histology • Micro-CT analysis – Micro-CT Scanning – Micro-CT scanning explant – Positioning and subtracting 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- -Thanks-