Kacey G. Marra

In vitro analysis of biodegradable polymer blend/hydroxyapatite composites for bone tissue engineering

Blends of biodegradable polymers, poly(caprolactone) and poly(D, L-lactic-co-glycolic acid), have been examined as scaffolds for applications in bone tissue engineering. Hydroxyapatite granules have been incorporated into the blends and porous discs were prepared. Mechanical properties and degradation rates in vitro of the composites were determined. The discs were seeded with rabbit bone marrow or cultured bone marrow stromal cells and incubated under physiological conditions. Polymer/ceramic scaffolds supported cell growth throughout the scaffold for 8 weeks. Scanning and transmission electron microscopy, and histological analyses were used to characterize the seeded composites. This study suggests the feasibility of using novel polymer/ceramic composites as scaffold in bone tissue engineering applications.

Composition options for tissue-engineered bone.

The logical assembly of tissue-engineered bone is ultimately directed by the clinical status of the patient. The basic elements for tissue-engineered bone should include signaling molecules, cells, and extracellular matrix. The assembly of these basic elements may need to be modified by tissue engineers to account for patient variables of age, gender, health, systemic conditions, habits, and anatomical implant. Moreover, different regions of the body will have different functional loads and vascularity. This review discusses several basic options that may be necessary to engineer bone, including spatial and temporal assembly of signaling factors, cells, and biomimetic extracellular matrices. Moreover, the importance of the health care status of the patient who may be receiving the tissue-engineered composition is emphasized.

Characterization of osteoblast-like behavior of cultured bone marrow stromal cells on various polymer surfaces

The creation of novel bone substitutes requires a detailed understanding of the interaction between cells and materials. This study was designed to test certain polymers, specifically poly(caprolactone) (PCL), poly(D,L-lactic-CO-glycolic acid) (PLGA), and combinations of these polymers for their ability to support bone marrow stromal cell proliferation and differentiation. Bone marrow stromal cells were cultured from New Zealand White rabbits and were seeded onto glass slides coated with a thin layer of PCL, PLGA, and combinations of these two polymers in both a 40:60 and a 10:90 ratio. Growth curves were compared. At the end of 2 weeks, the cells were stained for both matrix mineralization and alkaline phosphatase activity. There was no statistically significant difference in growth rate of the cells on any polymer or polymer combination. However, there was a striking difference in Von Kossa staining and alkaline phosphatase staining. Cells on PCL did not show Von Kossa staining or alkaline phosphatase staining. However, in the 40:60 and 10:90 blends, there was both positive Von Kossa and alkaline phosphatase staining. These data indicate that PCL alone may not be a satisfactory material for the creation of a bone substitute. However, it may be used in combination with PLGA for the creation of a bone substitute material.

The influence of polymer blend composition on the degradation of polymer/hydroxyapatite biomaterials

The in vitro degradation of biodegradable polymer/ceramic composites was assessed in two different environments under both static and pseudodynamic conditions. The blends, consisting of polycaprolactone, poly(lactic-co-glycolic acid), and hydroxyapatite, have potential use in bone tissue engineering applications, thus it is essential to establish a standardized method of characterizing the degradation of new biomaterials. In this study, the variation in polymer blend ratio was examined to observe a change in degradation rate. The porous blends were degraded in water and serum-containing media. A previous study examined in vitro degradation in serum-free buffer. Molecular weight loss, gravimetric weight loss, pH changes and morphological changes were evaluated. The changes in porosity were observed with scanning electron microscopy and quantitatively assessed using image analysis. There was a significant difference in molecular weight loss and gravimetric weight loss between the blends after 10 weeks in vitro. Blends containing the greatest amount of poly(lactic-co-glycolic acid) degraded most rapidly.

Rabbit Calvarial Wound Healing by Means of Seeded Caprotite® Scaffolds:

Autologous bone is the most successful bone-grafting material; however, limited supply and donor site morbidity are problematic. Synthetic bone substitutes are effective, but healing is slow and unpredictable. Osseous wound healing may be enhanced if bone substitutes are combined with autologous bone marrow cells. To test this hypothesis, we created 40 calvarial defects in 20 12-week-old New Zealand White rabbits, divided into four groups: (1) unrepaired controls, (2) autologous bone grafts, (3) unseeded Caprotite® (a polymer-ceramic composite) grafts, and (4) Caprotite® grafts seeded with autologous bone marrow stromal cells. CT scans were obtained at 0, 6, and 12 weeks post-operatively, and defects were harvested for histology. Defects repaired with autologous bone had significantly (p < 0.05) more bone than the other three groups, although seeded Caprotite® defects showed different wound-healing sequelae. Results suggest that seeded Caprotite® scaffolds did not significantly enhance osseous defect healing compared with controls.

Using PC12 cells to evaluate poly(caprolactone) and collagenous microcarriers for applications in nerve guide fabrication.

Tissue‐engineered nerve guides can provide mechanical support as well as chemical stimulation for nerve regeneration. PC12 cells were used to test the novel combination of poly(caprolactone) (PCL) and macroporous collagen‐based microcarriers (CultiSphers) as an initial phase in the fabrication of multichanneled nerve guides. Laminin‐coated PCL was an effective matrix for the attachment, proliferation, and viability of PC12 cells, relative to uncoated PCL. PC12 cells attached to laminin‐coated PCL and extended neurites when cultured in the presence of nerve growth factor (NGF). PC12 cells attached and proliferated on CultiSphers and extended neurites in response to NGF. A novel PCL/CultiSpher composite material also supported PC12 attachment and proliferation and provides a potentially useful material for the fabrication of synthetic nerve guides.

Novel three Dimensional Biodegradable Scaffolds for Bone Tissue Engineering

We have constructed osteogenic scaffolds using solid freeform fabrication techniques. Blends of biodegradable polymers, polycaprolactone and poly(D,L-lactic-co-glycolic acid), have been examined as scaffolds for applications in bone tissue engineering. Hydroxyapatite granules were incorporated into the blends and porous discs were prepared. Mechanical properties and degradation rates of the composites were determined. The discs were seeded with rabbit bone marrow or cultured bone marrow stromal cells and in vitro studies were conducted. Electron microscopy and histological analysis revealed an osteogenic composite that supports bone cell growth not only on the surface but throughout the 1 mm thick scaffold as well. Seeded and unseeded discs were mechanically assembled in layers and implanted in a rabbit rectus abdominis muscle. Bone growth was evident after eight weeks in vivo . Electron microscopy and histological analyses indicate vascularization and primitive bone formation throughout the seeded composite, and also a “fusion” of the layers to form a single, solid construct. Finally, we have begun to incorporate the growth factor IGF-I into the scaffold to enhance osteogenicity and/or as an alternative to cell seeding.