Structural Assessment of a Tissue Engineered Scaffold for Bone Repair

The limitations of current grafting materials have driven the search for synthetic alternatives to cancellous bone. A variety of biodegradable polymer foams composed of poly(lactide-coglycolide) [PLAGA] have been evaluated for such uses. However, structural limitations may restrict the clinical use of these scaffolds. We have developed a sintered microsphere scaffold composed of 85:15 poly(lactide-co-glycolide) with a biomimetic pore system equivalent to the structure of cancellous bone. Analysis of the structural data, indicated that the microsphere matrix sintered at a temperature of 160°C with a microsphere diameter of 355-425 μm resulted in a optimal, biomimetic structure with an approximate pore diameter of 75 to 275 μm, 35% porosity, and a compressive modulus of 272 MPa. The in vitro evaluation of human osteoblasts on the sintered matrix indicated that the structure was capable of supporting the attachment and proliferation of the cells throughout its pore system. Immunofluorescent staining of actin showed that the cells were proliferating 3-dimensionally through the pore system. The stain for osteocalcin showed that the cells had maintained the phenotypic expression for this bone specific protein. Through this work, it was shown that an osteoconductive PLAGA scaffold with a pore system equivalent to the structure of cancellous bone could be fabricated through the sintered microsphere method. INTRODUCTION Bone grafting is a surgical technique used to repair large skeletal defects. Currently, the most common graft material is cancellous bone taken from the patient’s iliac crest. The other types of commercially available grafting materials are allografts such as demineralized bone matrix particles and deproteinized cancellous chips, and synthetic alloplasts such as calcium sulfate pellets and porous calcium phosphate materials. Although autograft is the gold standard, synthetic bone graft materials are increasing in popularity. This is due to the intrinsic problems associated with using autogenous bone grafts. Limitations such as supply, donor site pain and infection, and unpredictable healing have stimulated the development of synthetic matrices engineered specifically for bone replacement applications. The majority of the tissue engineering scaffolds currently being researched are based on the biodegradable copolymer known as poly(lactide-co-glycolide) [PLAGA]. Our laboratory has conducted several studies evaluating the ability of PLAGA to promote osteoblast attachment, spread, and proliferation over its surface [1,2]. With a 30 year FDA approval, it has been extensively tested in vitro, in vivo, and in clinical settings. It is a degradable copolymer whose properties can be tailored based on the ratio of lactic acid to glycolic acid, and the molecular weight. Using a variety of techniques, PLAGA has been fabricated into 3-dimensional, porous scaffolds that are often combined with growth factors and/or osteoblasts for bone repair applications. The structure of many of these PLAGA matrices is based on foam techniques. A foam structure is characterized by a high % porosity (80-95% porous) and low mechanical properties (compressive modulus < 50 MPa). Although these structures have been shown to possess high porosity, the clinical use of these implants is restricted due to certain characteristic properties. The primary factor is that the mechanical properties of the implant may preclude its use in surgery. In our lab, we have taken a different approach to matrix fabrication. Using PLAGA microspheres, we have developed a sintered microsphere matrix with pore diameters ranging from 100-300 μm and mechanical properties in the mid range of cancellous bone [3]. In this technique, PLAGA microspheres of a narrow size range were thermally fused together in a 3-dimensional porous structure. Due to the nature of sphere packing, the resulting pores system had an interconnectivity of 100% and a % porosity of 30-40%. The goal of this study was to optimize the structure of the sintered microsphere matrix and examine the osteoconductivity of the optimal structure with respect to primary culture human osteoblasts. MATERIALS AND METHODS Microsphere Preparation (Solvent Evaporation Technique)