State of the art and future directions of scaffold-based bone engineering from a biomaterials perspective.

Scaffold-based bone tissue engineering aims to repair/regenerate bone defects. Such a treatment concept involves seeding autologous osteogenic cells throughout a biodegradable scaffold to create a scaffold-cell hybrid that may be called a tissue-engineered construct (TEC). A variety of materials and scaffolding fabrication techniques for bone tissue engineering have been investigated over the past two decades. This review aims to discuss the advances in bone engineering from a scaffold material point of view. In the first part the reader is introduced to the basic principles of bone engineering. The important properties of the biomaterials and the scaffold design in the making of tissue engineered bone constructs are discussed in detail, with special emphasis placed on the new material developments, namely composites made of synthetic polymers and calcium phosphates. Advantages and limitations of these materials are analysed along with various architectural parameters of scaffolds important for bone tissue engineering, e.g. porosity, pore size, interconnectivity and pore-wall microstructures.

Repair of calvarial defects with customised tissue-engineered bone grafts II. Evaluation of cellular efficiency and efficacy in vivo.

We have demonstrated in Part I of this study [see Schantz, J.-T., et al., Tissue Eng. 2003;9(Suppl. 1): S-113-S-126; this issue] that bone marrow-derived progenitor cells and calvarial osteoblasts could be successfully directed into the osteogenic lineage and cultured in three-dimensional (3-D) polycaprolactone (PCL) scaffolds. The objective of the second part of the study was to evaluate and to compare tissue engineered cell-polymer constructs using calvarial osteoblasts (group I) and mesenchymal progenitor cells (MPCs; group II) for the reconstruction of critical-size and three-dimensionally complex cranial defects. In 30 New Zealand White rabbits, bilateral parietal critical-size defects were created. On the basis of computed tomography scans, customized PCL scaffolds with precisely controlled microarchitecture were fabricated, using a rapid prototyping technology. Bone marrow-derived progenitor cells and osteoblasts were isolated and expanded in culture. Osteoblasts (group I) and mesenchymal progenitor cells (group II) were seeded in combination with a fibrin glue suspension into 40 PCL scaffolds. After incubating for 3 days in static culture, the PCL scaffold-cell constructs as well as nonseeded PCL scaffolds (control group) were implanted into 15-mm-diameter calvarial defects. Reconstruction of the cranium and bone formation were evaluated after 3 months. In vivo results indicated osseous tissue integration within the implant and functionally stable restoration of the calvarium. Islands of early bone formation could be observed in X-ray radiographs and in histological sections. Implants showed a high cell:ECM ratio and a dense vascular network. Mechanical testing of the reconstructed area revealed partial integration with the surrounding corticocancellous calvarial bone. The amount (area) of calcification, measured by clinical computed tomography, indicated that cell-seeded constructs measured about 60% more than unrepaired or unseeded scaffolds. Mechanical investigations revealed that stiffness reached 52 +/- 29 and 44 +/- 16 MPa for MPC- and osteoblast-seeded scaffolds, respectively. The yield strength for the push-out tests was 180 +/- 36 N for normal calvarial bone, 90 +/- 1 N for unrepaired site, and 106 +/- 10 N for unseeded constructs, which is about 60% of normal bone strength. MPC- and osteoblast-seeded scaffolds indicated a yield strength of 149 +/- 15 and 164 +/- 42 N, respectively, which is about 85-90% of normal bone. This study demonstrated that customized biodegradable polymeric implants may be used to deliver osteogenic cells and enhance bone formation within critically-sized defects in vivo. The use of rapid prototyping technology to produce scaffolds with controlled external geometry and microarchitecture offers new possibilities in the functional and aesthetic reconstruction of complex craniofacial defects.

Repair of calvarial defects with customized tissue-engineered bone grafts I. Evaluation of osteogenesis in a three-dimensional culture system.

Bone generation by autogenous cell transplantation in combination with a biodegradable scaffold is one of the most promising techniques being developed in craniofacial surgery. The objective of this combined in vitro and in vivo study was to evaluate the morphology and osteogenic differentiation of bone marrow derived mesenchymal progenitor cells and calvarial osteoblasts in a two-dimensional (2-D) and three-dimensional (3-D) culture environment (Part I of this study) and their potential in combination with a biodegradable scaffold to reconstruct critical-size calvarial defects in an autologous animal model [Part II of this study; see Schantz, J.T., et al. Tissue Eng. 2003;9(Suppl. 1):S-127-S-139; this issue]. New Zealand White rabbits were used to isolate osteoblasts from calvarial bone chips and bone marrow stromal cells from iliac crest bone marrow aspirates. Multilineage differentiation potential was evaluated in a 2-D culture setting. After amplification, the cells were seeded within a fibrin matrix into a 3-D polycaprolactone (PCL) scaffold system. The constructs were cultured for up to 3 weeks in vitro and assayed for cell attachment and proliferation using phase-contrast light, confocal laser, and scanning electron microscopy and the MTS cell metabolic assay. Osteogenic differentiation was analyzed by determining the expression of alkaline phosphatase (ALP) and osteocalcin. The bone marrow-derived progenitor cells demonstrated the potential to be induced to the osteogenic, adipogenic, and chondrogenic pathways. In a 3-D environment, cell-seeded PCL scaffolds evaluated by confocal laser microscopy revealed continuous cell proliferation and homogeneous cell distribution within the PCL scaffolds. On osteogenic induction mesenchymal progenitor cells (12 U/L) produce significantly higher (p < 0.05) ALP activity than do osteoblasts (2 U/L); however, no significant differences were found in osteocalcin expression. In conclusion, this study showed that the combination of a mechanically stable synthetic framework (PCL scaffolds) and a biomimetic hydrogel (fibrin glue) provides a potential matrix for bone tissue-engineering applications. Comparison of osteogenic differentiation between the two mesenchymal cell sources revealed a similar pattern.