C. M. Agrawal

Orthopaedic applications for PLA-PGA biodegradable polymers.

Biodegradable polymers, especially those belonging to the family of polylactic acid (PLA) and polyglycolic acid (PGA), play an increasingly important role in orthopaedics. These polymers degrade by hydrolysis and enzymatic activity and have a range of mechanical and physical properties that can be engineered appropriately to suit a particular application. Their degradation characteristics depend on several parameters including their molecular structure, crystallinity, and copolymer ratio. These biomaterials are also rapidly gaining recognition in the fledging field of tissue engineering because they can be fashioned into porous scaffolds or carriers of cells, extracellular matrix components, and bioactive agents. Although their future appears to be bright, several questions regarding the biocompatibility of these materials linger and should be addressed before their wide-scale use. In the context of musculoskeletal tissue, this report provides a comprehensive review of properties and applications of biodegradable PLA/PGA polymers and their copolymers. Of special interest are orthopaedic applications, biocompatibility studies, and issues of sterilization and storage of these versatile biomaterials. Also discussed is the fact that terms such as PLA, PGA, or PLA-PGA do not denote one material, but rather a large family of materials that have a wide range of differing bioengineering properties and concomitant biological responses. An analysis of some misconceptions, problems, and potential solutions is also provided.

Fundamentals of Biomechanics in Tissue Engineering of Bone

The objective of this review is to provide basic information pertaining to biomechanical aspects of bone as they relate to tissue engineering. The review is written for the general tissue engineering reader, who may not have a biomechanical engineering background. To this end, biomechanical characteristics and properties of normal and repair cortical and cancellous bone are presented. Also, this chapter intends to describe basic structure-function relationships of these two types of bone. Special emphasis is placed on salient classical and modern testing methods, with both material and structural properties described.

Effects of fluid flow on the in vitro degradation kinetics of biodegradable scaffolds for tissue engineering

Scaffolds fabricated from biodegradable polymers are used extensively in the field of tissue engineering. Many of these scaffolds are subjected to fluid flow, either in vivo or in bioreactors ex vivo. The goal of this study was to examine the effects of fluid flow on the degradation characteristics and kinetics of scaffolds in vitro. Scaffolds with different porosity and permeability values were fabricated using a copolymer of polylactic acid and polyglycolic acid. These scaffolds were subjected to degradation in phosphate buffered saline at 37 degrees C for up to 6 weeks under two test conditions: static and flow (250 microl/min). The porosity of the scaffolds decreased up to 2 weeks and then increased, while the elastic modulus first increased and then decreased over the course of the study. The mass and molecular weight of the scaffolds exhibited a steady decrease up to 6 weeks. The results further indicated that lower the porosity and permeability of the scaffolds, the faster their rate of degradation. Additionally, fluid flow decreased the degradation rate significantly. It is possible that the high rates of degradation observed here were due to autocatalysis of the degradation reaction by the acidic degradation products.

Microstructural heterogeneity and the fracture toughness of bone.

Age-related changes in the skeleton often lead to an increase in the susceptibility of bone to fracture. The purpose of this study was to determine whether differences in material properties between the osteonal and interstitial regions of bone have an effect on bone fracture properties. Parameters such as longitudinal fracture toughness, transverse fracture toughness, porosity, interstitial microhardness, osteonal microhardness, bone density, and weight fractions of the mineral and organic phases of bone were examined as a function of age using female baboon femurs. With increasing age, the longitudinal fracture toughness decreased significantly as did transverse fracture toughness, whereas the interstitial microhardness increased. However, no significant differences in the other parameters were observed as a function of age. Using the ratio of interstitial microhardness to osteonal microhardness as a measure of the differences in the material properties in these two regions, correlation analysis revealed that the longitudinal fracture toughness of bone has a significant correlation with the microhardness ratio. Localized differences in material properties between osteonal and interstitial regions of bone increase with age; such differences may result in high stress concentrations at cement lines and facilitate longitudinal crack propagation.

Protein release kinetics of a biodegradable implant for fracture non-unions.

Non-union of long bone fractures is often a serious complication of fracture healing. It is estimated that 100 000 non-unions occur in the united States annually and result in the loss of function of the involved limb. The present study was performed to develop a microporous polylactic acid-polyglycolic acid (PLA-PGA) implant for the delivery of bone morphogenetic protein (BMP) to sites of fracture non-unions, and to characterize the protein release kinetics of such an implant in vitro. A 50:50 copolymer of PLA-PGA was used to fabricate the implants using a gel formation technique. The implants were subjected to hydrolytic degradation in phosphate-buffered saline at 37 degrees C for up to 72 d. The protein release and the polymer degradation were monitored during this time period. The release kinetics of these implants were studied using a model protein, soybean trypsin inhibitor (TI), as well as BMP. The results indicate that there is a burst release of the proteins in the initial 48 h followed by a lower elution rate. The release of both the proteins followed similar trends. The molecular weight of the polymer decreased at a faster rate compared to its mass.

Changes in the fracture toughness of bone may not be reflected in its mineral density, porosity, and tensile properties

Age-related changes in the skeleton often lead to an increase in the susceptibility of bone to fracture. Such changes most likely occur in the constituents of bone, namely, the mineral and organic phases, and in their spatial arrangement manifested as orientation and microstructure. In the past, however, bone loss or decline in bone mineral density has been considered to be the major contributing factor for the increased risk of bone fractures, and elastic modulus and ultimate strength have been commonly used to assess bone quality and strength. However, whether these properties provide sufficient information regarding the likelihood of bone to fracture remains debatable. Using a novel fracture toughness test, which measures the energy or stress intensity required to propagate a crack within a material, the objective of this study was to investigate if the mineral density and mechanical properties of bone can accurately predict bone fragility as measured by fracture toughness. Changes in fracture toughness (K(IC)), bone mineral density (BMD), elastic modulus (E), yield and ultimate strength (sigma y and sigma s), porosity (P0), and microhardness (Hv) of bone were examined as a function of age in a baboon model. With increasing age, the fracture toughness of bone decreased, and its microhardness increased. However, no significant changes were found in BMD, E, P0, sigma y, and sigma s as a function of age. In addition, simple regression analyses revealed no significant correlation between bone fracture toughness and the other parameters, except for microhardness of bone. The results of this study indicate that changes in bone fracture toughness may not be necessarily reflected in its mineral density, porosity, elastic modulus, yield strength, and ultimate strength.

Effect of collagen denaturation on the toughness of bone.

The purpose of this study was to explore the relationship between the integrity of collagen and biomechanical properties of bone. In this study, age (range, 5-26 years old) and gender related changes in cortical bone samples from 33 baboon femurs (15 males and 18 females) were examined. The percentage of denatured collagen was determined using a selective digestion technique. The fracture toughness, elastic modulus, yield and ultimate strength, and energy to fracture of bone were determined in three-point bending configurations. The porosity and weight fractions of the mineral and organic phase also were measured. A two-way analysis of variance showed that age dependent changes were reflected primarily in the amount of denatured collagen, fracture toughness, energy to fracture, and elastic modulus, whereas gender had effects on the fracture toughness, elastic modulus, and porosity of bone. In addition, regression analyses indicated that the percentage of denatured collagen had an inverse correlation with the toughness of bone and a positive correlation with its elastic modulus, whereas mineral content had positive correlation with the strength and elastic modulus of bone. The results of this study suggest collagen influences the toughness of bone, whereas mineral content predominantly contributes to bone stiffness and strength.

The effect of ultra-high molecular weight polyethylene wear debris on MG63 osteosarcoma cells in vitro.

BACKGROUND Focal osteolysis due to ultra-high molecular weight polyethylene wear debris involves effects on both bone resorption and bone formation. METHODS The response of MG63 osteoblast-like osteosarcoma cells to ultra-high molecular weight polyethylene wear debris isolated by enzymatic digestion of granulomatous tissue obtained from the sites of failed total hip arthroplasties was examined. Scanning electron microscopy, particle-size analysis, and Fourier transform infrared spectroscopy were used to characterize the number, morphology, size distribution, and chemical composition of the particles. Cell response was assessed by adding particles at varying dilutions to confluent cultures and measuring changes in cell proliferation (number of cells and [3H]-thymidine incorporation), osteoblast function (alkaline-phosphatase-specific activity and osteocalcin production), matrix production (collagen production and proteoglycan sulfation), and local cytokine production (prostaglandin-E2 production). RESULTS The mean size of the particles was 0.60 micrometer, and 95 percent of the particles had a size of less than 1.5 micrometers. The number of particles per gram of tissue ranged from 1.39 to 3.38x10(9). Three of the four batches of particles were endotoxin-free. Exposure of the cells to particles of wear debris significantly increased the number of cells (p<0.05) and the [3H]-thymidine incorporation (p<0.05) in a dose-dependent manner. In contrast, the addition of particles decreased alkaline-phosphatase-specific activity and osteocalcin production. Collagen production and proteoglycan sulfation were also decreased, while prostaglandin-E2 synthesis was increased by the addition of particles. CONCLUSIONS Ultra-high molecular weight polyethylene particles isolated from human tissue stimulated osteoblast proliferation and prostaglandin-E2 production and inhibited cell differentiation and matrix production. These results indicate that particles of wear debris inhibit cell functions associated with bone formation and that osteoblasts may produce factors in response to wear debris that influence neighboring cells, such as osteoclasts and macrophages. CLINICAL RELEVANCE Particles of wear debris, especially ultra-high molecular weight polyethylene, have been implicated in the loosening of implants and the development of osteolysis. The present study shows that particles of ultra-high molecular weight polyethylene isolated from human tissue inhibit osteoblast functions associated with bone formation. In addition, particles of wear debris induced osteoblasts to secrete factors capable of influencing neighboring cells, such as osteoclasts and macrophages. These results suggest that osteoblasts may play a role in the cascade of events leading to granuloma formation, osteolysis, and failure of orthopaedic implants.

Effects of Collagen Unwinding and Cleavage on the Mechanical Integrity of the Collagen Network in Bone

The objective of this study was to investigate how molecular level changes in the collagen network affect its mechanical integrity. Our hypothesis is that the cleavage and unwinding of triple helices of collagen molecules significantly reduce the mechanical integrity of the collagen network in bone, whereas collagen crosslinks play a major role in sustaining the structural integrity of the collagen network. To test this hypothesis, the collagen molecular structure was altered in demineralized human cadaveric bone samples in the following two ways: heat induced unwinding and pancreas elastase induced cleavage of collagen molecules. Along with control specimens, the treated specimens were mechanically tested in tension to determine their strength, elastic modulus, toughness, and strain to failure. Also, the percentage of denatured collagen molecules and amounts of two major collagen crosslinks (hydroxylysylpyridinoline and lysylpyridinoline) were determined using high-performance liquid chromatography techniques. It was found that unwinding of collagen molecules may cause more reduction in stiffness (E) but less strain to failure (ef) than cleavage. Both collagen denaturation types cause similar changes in the strength (ss) and work to fracture (Wf) of the collagen network with no significant changes in hydroxylysylpyridinoline and lysylpyridinoline crosslinks. The results of this study indicate that the integrity of collagen molecules significantly affect the mechanical properties of the collagen network in bone, and that collagen crosslinks may play an important role in maintaining the mechanical integrity of the collagen network after collagen denaturation occurs.

Salient Degradation Features of a 50:50 PLA/PGA Scaffold for Tissue Engineering

An implant system that undergoes a gradual, time-dependent, nontoxic degradation process may offer an efficacious, safe, and desirable alternative to metallic materials used in the treatment of various musculoskeletal conditions. Such a scaffold may also be a suitable vehicle for growing cells and tissue in the laboratory for tissue engineering applications. We have used a scaffold of this type previously in animal studies for biological resurfacing of large articular cartilage defects.(1) This study examined important in vitro degradation characteristics of a 50:50 polylactic acid/polyglycolic acid (PLG) implant during an 8-week period. It was determined that this particular implant degraded in a biphasic fashion. The initial phase occurred during the first 2 weeks with a decrease in molecular weight and surface axial strain, coupled with an increase in percent porosity. The second phase demonstrated a decline in surface axial strain by 4 weeks and an ongoing decline in molecular weight. Loss of gross structural properties was not evident until the start of the second phase and was complete at 8 weeks. This study demonstrated the potential uses for this implant as a means of providing structural support for cells and tissue ingrowth for up to 8 weeks. Further studies need to be conducted in order to determine the biological effects of the degrading polymer byproducts on host tissues.