Elke Berneel

Synergistic effect of surface modification and scaffold design of bioplotted 3-D poly-ε-caprolactone scaffolds in osteogenic tissue engineering.

The hydrophobic nature and the regular scaffold architecture of bioplotted poly(ε-caprolactone) (PCL) scaffolds present some hurdles for homogeneous tissue formation and differentiation. The current hypothesis is that a synergistic effect of applied surface modification and scaffold design enhances colonization and osteogenic differentiation. First, PCL scaffolds with a 0/90° lay-down pattern (0/90) were plotted and subjected to an oxygen plasma (O2) or multistep surface modification, including post-argon 2-amino-ethylmethacrylate grafting (AEMA), followed by immobilization of gelatin type B (gelB) and physisorption of fibronectin (gelB Fn). Secondly, scaffolds of different designs were plotted (0/90° shift (0/90 S), 0/45° and 0/90° with narrow pores (0/90 NP)) and subjected to the double protein coating. Preosteoblasts were cultured on the scaffolds and the seeding efficiency, colonization and differentiation were studied. The data revealed that a biomimetic surface modification improved colonization (gelB Fn>gelB>AEMA>O2). Compact scaffold architectures (0/90 NP, 0/45, 0/90 S>0/90) positively influenced the seeding efficiency and differentiation. Interestingly, the applied surface modification had a greater impact on colonization than the scaffold design. In conclusion, the combination of a double protein coating with a compact design enhances tissue formation in the plotted PCL scaffolds.

Double protein-coated poly-ε-caprolactone scaffolds: successful 2D to 3D transfer.

In the past decade, tissue engineering has evolved from a promising technology to an established scientific field. Large attention has focussed on developing scaffolds from both biodegradable and nondegradable polymers to be cultivated with cells, to replace human body defects. The major drawback of most polymers is however their limited cell-interactive properties. An additional complication when developing a surface modification protocol for those materials is the transferability of protocols from 2D substrates to 3D scaffolds. In the present work, we therefore report on possible biological effects originating from the transfer of a double protein coating protocol, involving gelatin type B and fibronectin, from 2D poly-ε-caprolactone (PCL) films to 3D PCL scaffolds produced by rapid prototyping. A variety of techniques including scanning electron microscopy, X-ray photoelectron spectroscopy and confocal fluorescence microscopy confirmed a successful and homogeneous protein-coating on both 2D and 3D substrates. Interestingly, the biological performance of the double protein-coated PCL substrates, reflected by the initial cell adhesion, proliferation, and colonization was superior compared to the other surface modification steps, independent of the material dimension.

Redifferentiation of High-Throughput Generated Fibrochondrocyte Micro-Aggregates: Impact of Low Oxygen Tension

In meniscus tissue engineering strategies, enhancing the matrix quality of the neomeniscal tissue is important. When the differentiated phenotype of fibrochondrocytes is lost, the quality of the matrix becomes compromised. The objective of this study was to produce uniform fibrochondrocyte micro-aggregates with desirable phenotype and tissue homogeneity in large quantities using a simple and reproducible method. Furthermore, we investigated if hypoxia could enhance the matrix quality. Porcine fibrochondrocytes were expanded at 21% oxygen until passage 3 (P3) and a gene expression profile was determined. P3 fibrochondrocytes were cultivated in chondrogenic medium at 5 and 21% oxygen in high-throughput agarose chips containing 2,865 microwells 200 µm in diameter. Evaluation included live/dead staining, histological examination, immunohistochemistry, dimethylmethylene blue assay and real-time reverse transcriptase quantitative polymerase chain reaction of the micro-aggregates. Gene expression analysis showed a drastic decline in collagen II and high expression of collagen I during monolayer culture. After 4 days, uniform and stable micro-aggregates could be produced. The redifferentiation and matrix quality of the hypoxic cultured micro-aggregates were enhanced relative to the normoxic cultures. Sulfated glycosaminoglycan synthesis was significantly higher, and collagen II expression and the collagen II/collagen I ratio were significantly upregulated in the hypoxic cultures. High-throughput production of uniform microtissues holds promise for the generation of larger-scale tissue engineering constructs or optimization of redifferentiation mechanisms for clinical applications.