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Quantitative computed tomography-based finite-element analysis (QCT/FEA) has become increasingly popular in an attempt to understand and possibly reduce vertebral fracture risk. It is known that scanning acquisition settings affect Hounsfield units (HU) of the CT voxels. Material properties assignments in QCT/FEA, relating HU to Young's modulus, are performed by applying empirical equations. The purpose of this study was to evaluate the effect of QCT scanning protocols on predicted stiffness values from finite-element models. One fresh frozen cadaveric torso and a QCT calibration phantom were scanned six times varying voltage and current and reconstructed to obtain a total of 12 sets of images. Five vertebrae from the torso were experimentally tested to obtain stiffness values. QCT/FEA models of the five vertebrae were developed for the 12 image data resulting in a total of 60 models. Predicted stiffness was compared to the experimental values. The highest percent difference in stiffness was approximately 480% (80 kVp, 110 mAs, U70), while the lowest outcome was ∼1% (80 kVp, 110 mAs, U30). There was a clear distinction between reconstruction kernels in predicted outcomes, whereas voltage did not present a clear influence on results. The potential of QCT/FEA as an improvement to conventional fracture risk prediction tools is well established. However, it is important to establish research protocols that can lead to results that can be translated to the clinical setting.

We report a new crosslinked polymersome with pH-responsive swelling properties through acidic hydrolysis of hydrophobic contents from the amphiphilic polymer chains. Its unique stability under physiological conditions and large swelling capability under low pH conditions give this polymersome promising potential for anticancer drug delivery.

We compared open stabilization of vertebral fractures to percutaneous spinal fixation techniques in patients with a diagnosis of either ankylosing spondylitis (AS) or diffuse idiopathic skeletal hyperostosis (DISH). A retrospective review of patients known to have AS or DISH treated for spinal column fracture at a single institution between 1995 and 2011 was performed. Patients were analyzed by the type of fixation, divided into either a percutaneous group (PG) or an open group (OG). There were 41 patients identified with a spinal column fracture and history of AS or DISH who received surgical intervention. There were 17 (42%) patients with AS and 24 (58%) with DISH. Patients in the PG and OG cohorts presented with similar mechanisms of injury, Injury Severity Scale, number of vertebral fractures, number of additional injuries, and Arbeitsgemeinschaft fur Osteosynthesefragen (AO) classification scores. Mean operative time (254.76minutes versus 334.67minutes, p=0.040), estimated blood loss (166.8 versus 1240.36mL, p

Surgical site infection (SSI) remains a significant risk for any clean orthopedic surgical procedure. Complications resulting from an SSI often require a second surgery and lengthen patient recovery time. The efficacy of antimicrobial agents delivered to combat SSI is diminished by systemic toxicity, bacterial resistance, and patient compliance to dosing schedules. We submit that development of localized, controlled release formulations for antimicrobial compounds would improve the effectiveness of prophylactic surgical wound antibiotic treatment while decreasing systemic side effects. Our research group developed and characterized oligo(poly(ethylene glycol)fumarate) / sodium methacrylate (OPF/SMA) charged copolymers as biocompatible hydrogel matrices. Here, we report the engineering of this copolymer for use as an antibiotic delivery vehicle in surgical applications. We demonstrate that these hydrogels can be efficiently loaded with vancomycin (over 500 μg drug per mg hydrogel) and this loading mechanism is both time- and charge-dependent. Vancomycin release kinetics are shown to be dependent on copolymer negative charge. In the first 6 hours, we achieved as low as 33.7% release. In the first 24 hours, under 80% of total loaded drug was released. Further, vancomycin release from this system can be extended past four days. Finally, we show that the antimicrobial activity of released vancomycin is equivalent to stock vancomycin in inhibiting the growth of colonies of a clinically derived strain of methicillin-resistant Staphylococcus aureus. In summary, our work demonstrates that OPF/SMA hydrogels are appropriate candidates to deliver local antibiotic therapy for prophylaxis of surgical site infection.

Research on biomaterial nerve scaffolds has been carried out for 50 years. Only three materials (collagen, polycaprolactone and polyglycollic acid) have progressed to clinical use. Pre-clinical animal models are critical for testing nerve scaffolds prior to implementation in clinical practice. We have conducted a systematic review of 416 reports in which animal models were used for evaluation of nerve regeneration into synthetic conduits. A valid animal model of nerve regeneration requires it to reproduce the specific processes that take place in regeneration after human peripheral nerve injury. No distinct animal species meets all the requirements for an ideal animal model. Certain models are well suited for understanding regenerative neurobiology while others are better for pre-clinical evaluation of efficacy. The review identified that more than 70 synthetic materials were tested in eight species using 17 different nerves. Nerve gaps ranged from 1 to 90 mm. More than 20 types of assessment methodology were used with no standardization of methods between any of the publications. The review emphasizes the urgent need for standardization or rationalization of animal models and evaluation methods for studying nerve repair.

In this work, we have investigated the development of a synthetic hydrogel that contains a negatively charged phosphate group for use as a substrate for bone cell attachment and differentiation in culture. The photoreactive, phosphate-containing molecule, bis(2-(methacryloyloxy)ethyl)phosphate (BP), was incorporated into oligo(polyethylene glycol) fumarate hydrogel and the mechanical, rheological and thermal properties of the resulting hydrogels were characterized. Our results showed changes in hydrogel compression and storage moduli with incorporation of BP. The modification also resulted in decreased crystallinity as recorded by differential scanning calorimetry. Our data revealed that incorporation of BP improved attachment and differentiation of human fetal osteoblast (hFOB) cells in a dose-dependent manner. A change in surface chemistry and mineralization of the phosphate-containing surfaces verified by scanning electron microscopy and energy dispersive X-ray analysis was found to be important for hFOB cell attachment and differentiation. We also demonstrated that phosphate-containing hydrogels support attachment and differentiation of primary bone marrow stromal cells. These findings suggest that BP-modified hydrogels are capable of sustaining attachment and differentiation of both bone marrow stromal cells and osteoblasts that are critical for bone regeneration.

Object Sacrectomy positioning must balance surgical exposure, localization, associated operative procedures, and patient safety. Poor positioning may increase hemorrhage, risk of blindness, and skin breakdown. Methods The authors prospectively identified positioning-related morbidity in 17 patients undergoing 19 prone sacral procedures from September 2008 to August 2009 following institution of a standardized positioning protocol. Key elements include skull traction/head suspension, an open radiolucent frame, and wide draping for associated closure and reconstructive procedures. Results Tumors included 5 chordomas, 4 high-grade sarcomas, 1 chondrosarcoma, 2 presacral extradural myxopapillary ependymomas, and 5 others. Mean patient age was 49.9 years (range 17–74 years); mean body mass index was 27.6 kg/m2 (range 19.3–43.9 kg/m2). Mean preoperative Braden skin integrity score was 21.1 (range 17–23). Average operative time was 501 minutes (range 158–1136 minutes). Prone surgery was a part of staged anterior/posterior resections in 8 patients. Localization was conducted using fluoroscopy in 13 patients and intraoperative CT in 4 patients. All imaging studies were successful. One patient developed a transient ulnar nerve palsy attributed to positioning. Three patients (two of whom were morbidly obese) developed Stage I pressure injuries to the chest and another developed Stage II pressure injury following a 1136-minute procedure. Morbidity was only observed in patients with morbid obesity or with procedures lasting in excess of 10 hours. Conclusions A positioning protocol using head suspension on an open radiolucent frame facilitates oncological sacral surgery with reasonable patient morbidity. Morbid obesity and procedure times in excess of 10 hours are risk factors for positioning-related complications. To the authors' knowledge, this is the first report of surgical positioning morbidity in this patient population.

Well-differentiated liposarcoma (WDLS) is one of the most common malignant mesenchymal tumors and dedifferentiated liposarcoma (DDLS) is a malignant tumor consisting of both WDLS and a transformed nonlipogenic sarcomatous component. Cytogenetically, WDLS is characterized by the presence of ring or giant rod chromosomes containing several amplified genes, including MDM2, TSPAN31, CDK4, and others mainly derived from chromosome bands 12q13-15. However, the 12q13-15 amplicon is large and discontinuous. The focus of this study was to identify novel critical genes that are consistently amplified in primary (nonrecurrent) WDLS and with potential relevance for future targeted therapy. Using a high-resolution (5.0 kb) "single nucleotide polymorphism"/copy number variation microarray to screen the whole genome in a series of primary WDLS, two consistently amplified areas were found on chromosome 12: one region containing the MDM2 and CPM genes, and another region containing the FRS2 gene. Based on these findings, we further validated FRS2 amplification in both WDLS and DDLS. Fluorescence in situ hybridization confirmed FRS2 amplification in all WDLS and DDLS tested (n = 57). Real time PCR showed FRS2 mRNA transcriptional upregulation in WDLS (n = 19) and DDLS (n = 13) but not in lipoma (n = 5) and normal fat (n = 9). Immunoblotting revealed high expression levels of phospho-FRS2 at Y436 and slightly overexpression of total FRS2 protein in liposarcoma but not in normal fat or preadipocytes. Considering the critical role of FRS2 in mediating fibroblast growth factor receptor signaling, our findings indicate that FRS2 signaling should be further investigated as a potential therapeutic target for liposarcoma.

Osteogenesis imperfecta (OI) is most often caused by mutations in the type I procollagen genes (COL1A1/COL1A2). We identified two children with substitutions in the type I procollagen C-propeptide cleavage site, which disrupt a unique processing step in collagen maturation and define a novel phenotype within OI. The patients have mild OI caused by mutations in COL1A1 (Patient 1: p.Asp1219Asn) or COL1A2 (Patient 2: p.Ala1119Thr), respectively. Patient 1 L1-L4 DXA Z-score was +3.9 and pQCT vBMD was+3.1; Patient 2 had L1-L4 DXA Z-score of 0.0 and pQCT vBMD of -1.8. Patient BMD contrasts with radiographic osteopenia and histomorphometry without osteosclerosis. Mutant procollagen processing is impaired in pericellular and in vitro assays. Patient dermal collagen fibrils have irregular borders. Incorporation of pC-collagen into matrix leads to increased bone mineralization. FTIR imaging confirms elevated mineral/matrix ratios in both patients, along with increased collagen maturation in trabecular bone, compared to normal or OI controls. Bone mineralization density distribution revealed a marked shift toward increased mineralization density for both patients. Patient 1 has areas of higher and lower bone mineralization than controls; Patient 2's bone matrix has a mineral content exceeding even classical OI bone. These patients define a new phenotype of high BMD OI and demonstrate that procollagen C-propeptide cleavage is crucial to normal bone mineralization.

The effect of microsphere delivered Nerve Growth Factor (NGF) in a poly-lactic-co-glycolic-acid (PLGA) 85/15 nerve conduit bridging a 10mm rat sciatic nerve gap was assessed, comparing nine groups (n = 6): PLGA conduits filled with saline, saline and NGF, saline with blank microspheres; four different NGF microspheres (5, 20, 50, and 100 mg/ml); an autologous graft and sciatic nerve gap. Histomorphometry, retrograde tracing, electrophysiology, and functional outcomes were evaluated up to 16 weeks. The autologous graft showed the largest fascicular area (0.65 mm(2) ) and had a significantly greater number of myelinated fibers (P < 0.0001). Electrophysiology showed Compound Muscle Action Potential (CMAP) recordings for the autologous graft returning at 6 weeks after nerve transection, reaching their highest amplitude of 3.6 mV at endpoint. No significant differences were found in functional evaluation between groups or between conduits with microspheres and the saline filled conduit. A PLGA 85/15 nerve conduit is capable of sustaining nerve regeneration. The microsphere delivery system does not interfere with regeneration.

The mechanical and electrical properties of polycaprolactone fumarate-polypyrrole (PCLF-PPy) scaffolds were studied under physiological conditions to evaluate their ability to maintain the material properties necessary for application as conductive nerve conduits. PC12 cells cultured on PCLF-PPy scaffolds were stimulated with regimens of 10 μA of either a constant or a 20 Hz frequency current passed through the scaffolds for 1h per day. PC12 cellular morphologies were analyzed by fluorescence microscopy after 48 h. PCLF-PPy scaffolds exhibited excellent mechanical properties at 37 °C which would allow suturing and flexibility. The surface resistivity of the scaffolds was 2 kΩ and the scaffolds were electrically stable during the application of electrical stimulation (ES). In vitro studies showed significant increases in the percentage of neurite bearing cells, number of neurites per cell and neurite length in the presence of ES compared with no ES. Additionally, extending neurites were observed to align in the direction of the applied current. This study shows that electrically conductive PCLF-PPy scaffolds possess the material properties necessary for application as nerve conduits. Additionally, the capability to significantly enhance and direct neurite extension by passing an electrical current through PCLF-PPy scaffolds renders them even more promising as future therapeutic treatments for severe nerve injuries.

Biphenotypic sinonasal sarcoma (SNS) is a newly described tumor of the nasal and paranasal areas. Herein, we report the novel recurring chromosomal translocation t(2;4)(q35;q31.1) in SNS. The translocation results in the formation of the fusion protein PAX3-MAML3, which is a potent transcriptional activator of PAX3 response elements. The SNS phenotype is characterized by aberrant expression of genes involved in neuroectodermal and myogenic differentiation, which closely simulates the developmental roles of PAX3.

Regeneration of peripheral nerves after injury is suboptimal. We now report the long term delivery of nerve growth factor (NGF) by biodegradable poly-lactic-co-glycolic acid (PLGA) microspheres in vitro and in vivo. Lactic to glycolic acid ratios of 50:50 and 85:15 were fabricated using the double emulsion solvent, evaporation technique. Three different inherent viscosities (0.1 dL g(-1) : 1A, 0.4 dL g(-1) : 4A, 0.7 dL g(-1) : 7A) were analyzed. In vitro, release of NGF for 23 days was measured. Electron microscopy demonstrated intact spheres for at least 7 days (50:50 1A), 14 days (50:50 4A), or 35 days (50:50 7A and 85:15 7A). In vitro release kinetics was characterized by burst release, followed by release of NGF at a rate of 0.6-1.6% a day. Release curves for 50:50 1A and 85:15 7A differed significantly from other compositions (p0.01). In vivo, release was characterized by a novel radionuclide tracking assay. Release rates varied from 0.9 to 2.2% per day with linear kinetics. All but the 85:15 type of spheres showed different release profiles in vivo compared to in vitro conditions. On the basis of the surface morphology and release profiles, we found microspheres fabricated from 50:50 4A PLGA to be best suited for the use in a rat sciatic nerve injury model.

Comprehensive in vivo biodegradability and biocompatibility of unmodified and Arg-Gly-Asp (RGD) peptide-modified PEG/Sebacic acid based hydrogels were evaluated and compared to the control material poly(lactide-co-glycolide) (PLGA) using a cage implantation system, as well as direct subcutaneous implantation for up to 12 weeks. The total weight loss after 12 weeks of implantation for unmodified PEGSDA and RGD-modified PEGSDA in the cage was approximately 42% and 52%, respectively, with no statistical difference (p> 0.05). The exudate analysis showed that PEGSDA hydrogels induced minimal inflammatory response up to 21 days following implantation, similar to the controls (empty cage and the cage containing PLGA discs). Histology analysis from direct subcutaneous implantation of the hydrogels and PLGA scaffold showed statistically similar resolution of the acute and chronic inflammatory responses with development of the fibrous capsule between the PEGSDA hydrogels and the control (PLGA). The cage system, as well as the histology analysis, demonstrated that the degradation products of both hydrogels, with or without RGD peptide modification, are biocompatible without statistically significant differences in the inflammatory responses, as compared to PLGA.

Off-the-shelf availability in large quantities, drug delivery functionality, and modifiable chemistry and mechanical properties make synthetic polymers highly suitable candidates for bone grafting. However, most synthetic polymers lack the ability to support cell attachment, proliferation, migration, and differentiation, and ultimately tissue formation. Incorporating anionic peptides into the polymer that mimics acidic proteins, which contribute to biomineralization and cellular attachment, could enhance bone formation. Therefore, this study investigates the effect of a phosphate functional group on osteoconductivity and BMP-2-induced bone formation in an injectable and biodegradable oligo[(polyethylene glycol) fumarate] (OPF) hydrogel. Three types of OPF hydrogels were fabricated using 0%, 20%, or 40% Bis(2-(methacryloyloxy)ethyl) phosphate creating unmodified OPF-noBP and phosphate-modified OPF-BP20 and OPF-BP40, respectively. To account for the osteoinductive effect of various BMP-2 release profiles, two different release profiles (i.e., different ratios of burst and sustained release) were obtained by varying the BMP-2 loading method. To investigate the osteoconductive effect of phosphate modification, unloaded OPF composites were assessed for bone formation in a bone defect model after 3, 6, and 9 weeks. To determine the effect of the hydrogel phosphate modification on BMP-2-induced bone formation, BMP-2 loaded OPF composites with differential BMP-2 release were analyzed after 9 weeks of subcutaneous implantation in rats. The phosphate-modified OPF hydrogels (OPF-BP20 and OPF-BP40) generated significantly more bone in an orthotopic defect compared to the unmodified hydrogel (OPF-noBP). Furthermore, the phosphate functionalized surface-enhanced BMP-2-induced ectopic bone formation regardless of the BMP-2 release profile. In conclusion, this study clearly shows that phosphate functional groups improve the osteoconductive properties of OPF and enhanced BMP-2-induced bone formation. Therefore, functionalizing hydrogels with phosphate groups by crosslinking monomers into the hydrogel matrix could provide a valuable method for improving polymer characteristics and holds great promise for bone tissue engineering.

We have previously shown experimental transplantation of living allogeneic bone to be feasible without long-term immunosuppression by development of a recipient-derived neoangiogenic circulation within bone. In this study, we examine the role of angiogenic cytokine delivery with biodegradable microspheres to enhance this process. Microsurgical femoral allotransplantation was performed from Dark Agouti to Piebald Virol Glaxo rats. Poly(D,L-lactide-co-glycolide) microspheres loaded with buffer, basic fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), or both, were inserted intramedullarly along with a recipient-derived arteriovenous (a/v) bundle. FK-506 was administered daily for 14 days, then discontinued. At 28 days, bone blood flow was measured using hydrogen washout. Microangiography, histologic, and histomorphometric analyses were performed. Capillary density was greater in the FGF+VEGF group (35.1%) than control (13.9%) (p < 0.05), and a linear trend was found from control, FGF, VEGF, to FGF+VEGF (p < 0.005). Bone formation rates were greater with VEGF (p < 0.01) and FGF+VEGF (p < 0.05). VEGF or FGF alone increased blood flow more than when combined. Histology rejection grading was low in all grafts. Local administration of vascular and fibroblast growth factors augments angiogenesis, bone formation, and bone blood flow from implanted blood vessels of donor origin in vascularized bone allografts after removal of immunosuppression.

We present enhanced cell ingrowth and proliferation through cross-linked three-dimensional (3D) nanocomposite scaffolds fabricated using poly(propylene fumarate) (PPF) and hydroxyapatite (HA) nanoparticles. Scaffolds with controlled internal pore structures were produced from computer-aided design (CAD) models and solid freeform fabrication (SFF) technique, while those with random pore structures were fabricated by a NaCl leaching technique for comparison. The morphology and mechanical properties of scaffolds were characterized using scanning electron microscopy (SEM) and mechanical testing, respectively. Pore interconnectivity of scaffolds was assessed using X-ray microcomputed tomography (micro-CT) and 3D imaging analysis. In vitro cell studies have been performed using MC3T3-E1 mouse preosteoblasts and cultured scaffolds in a rotating-wall-vessel bioreactor for 4 and 7 days to assess cell attachment, viability, ingrowth depth, and proliferation. The mechanical properties of cross-linked nanocomposite scaffolds were not significantly different after adding HA or varying pore structures. However, pore interconnectivity of PPF/HA nanocomposite scaffolds with controlled pore structures has been significantly increased, resulting in enhanced cell ingrowth depth 7 days after cell seeding. Cell attachment and proliferation are also higher in PPF/HA nanocomposite scaffolds. These results suggest that cross-linked PPF/HA nanocomposite scaffolds with controlled pore structures may lead to promising bone tissue engineering scaffolds with excellent cell proliferation and ingrowth.

Oligo (polyethylene glycol) fumarate (OPF) hydrogel has been employed in musculoskeletal tissue engineering for photo-encapsulation of chondrocytes and as a matrix for marrow stromal cells differentiation. In this study, we have studied the application of OPF hydrogel for co-encapsulation of DNA and bone cells and examined whether co-encapsulation can enhance gene transfer by maintaining the DNA within the cellular microenvironment. Our results showed that plasmid DNA encoding green fluorescence protein (GFP), co-encapsulated with bone tumor cells, was capable of transfecting the cells and the transfected tumor cells continuously expressed GFP protein over the time course of study (21 days). Furthermore, we have examined the co-encapsulation of estrogen receptor (ER) encoding plasmid DNA and human fetal osteoblast cells (hFOB) that lack endogenous ER. Our results show that the transfected cells responded to estrogen as alkaline phosphatase (ALP), and estrogen response element (ERE)-directed luciferase enzyme activities increased with estrogen-treatment. Taken together, these studies show that OPF hydrogel could be further explored for targeted gene delivery in bone and other tissues encapsulated within the hydrogels.

We have developed a new fabrication technique to create three-dimensional (3D) porous poly(epsilon-caprolactone fumarate) (PCLF) scaffolds using hydrogel microparticle porogens, as an alternative to overcome certain limitations of traditional scaffold fabrication techniques such as a salt leaching method. Both natural hydrogel, gelatin, and synthetic hydrogel, poly(ethylene glycol) sebacic acid diacrylate, were used as porogens to fabricate 3D porous PCLF scaffolds. Hydrogel microparticles were prepared by a single emulsion technique with the particle size in the range of 100-500 microm after equilibrium in water. The pore size distribution, porosity, pore interconnectivity, and spatial pore heterogeneity of the 3D PCLF scaffolds were assessed using micro-computed tomography and imaging analysis. Scaffolds fabricated with the hydrogel porogens had higher porosity and pore interconnectivity as well as more homogeneous spatial pore distribution, compared to the scaffolds made from the salt leaching process. Compressive moduli of the scaffolds were also measured and showed that lower porosity yielded greater modulus of the scaffolds. Overall, the new fabrication technology using hydrogel porogens may be beneficial for certain tissue engineering applications.

Object Glial scar and cystic formation greatly contribute to the inhibition of axonal regeneration after spinal cord injury (SCI). Attempts to promote axonal regeneration are extremely challenging in this type of hostile environment. The objective of this study was to examine the surgical methods that may be used to assess the factors that influence the level of scar and cystic formation in SCI. Methods In the first part of this study, a complete transection was performed at vertebral level T9–10 in adult female Sprague-Dawley rats. The dura mater was either left open (control group) or was closed using sutures or hyaluronic acid. In the second part of the study, complete or subpial transection was performed, with the same dural closure technique applied to both groups. Histological analysis of longitudinal sections of the spinal cord was performed, and the percentage of scar and cyst formation was determined. Results Dural closure using sutures resulted in significantly less glial scar formation (p = 0.0248), while incorporation of the subpial transection surgical technique was then shown to significantly decrease cyst formation (p < 0.0001). Conclusions In this study, the authors demonstrated the importance of the vasculature in cyst formation after spinal cord trauma and confirmed the importance of dural closure in reducing glial scar formation.

Human mesenchymal stem cells offer a potential alternative to embryonic stem cells in clinical applications. The ability of these cells to self-renew and differentiate into multiple tissues, including bone, cartilage, fat, and other tissues of mesenchymal origin, makes them an attractive candidate for clinical applications. Patients who experience fracture nonunion and metabolic bone diseases, such as osteogenesis imperfecta and hypophosphatasia, have benefited from human mesenchymal stem cell therapy. Because of their ability to modulate immune responses, allogeneic transplant of these cells may be feasible without a substantial risk of immune rejection. The field of regenerative medicine is still facing considerable challenges; however, with the progress achieved thus far, the promise of stem cell therapy as a viable option for fracture nonunion and metabolic bone diseases is closer to reality. In this review, we update the biology and clinical applicability of human mesenchymal stem cells for bone repair and metabolic bone diseases.

Novel biodegradable injectable poly(ethylene glycol)-(PEG) based macromers were synthesized by reacting low-molecular weight PEG (MW: 200) and dicarboxylic acids such as sebacic acid or terephthalic acid. Chemical structures of the resulting polymers were confirmed by Fourier transform infrared (FTIR) and nuclear magnetic resonance (NMR) spectroscopy characterizations. Differential scanning calorimetry (DSC) showed that these polymers were completely amorphous above room temperature. After photopolymerization, dynamic elastic shear modulus of the crosslinked polymers was up to 1.5 MPa and compressive modulus was up to 2.2 MPa depending on the polymer composition. The in vitro degradation study showed that mass losses of these polymers were gradually decreased over 23 weeks of period in simulated body fluid. By incorporating up to 30 wt % of 2-hydroxyethyl methylmethacrylate (HEMA) into the crosslinking network, the dynamic elastic modulus and compressive modulus was significantly increased up to 7.2 and 3.2 MPa, respectively. HEMA incorporation also accelerated the degradation as indicated by substantially higher mass loss of up to 27% after 20 weeks of incubation. Cytocompatability studies using osteoblasts and neural cells revealed that cell metabolic activity on these polymers with or without HEMA was close to the control tissue culture polystyrene. The PEG-based macromers developed in this study may be useful as scaffolds or cell carriers for tissue engineering applications.

Regeneration of endogenous axons through a Schwann cell (SC)-seeded scaffold implant has been demonstrated in the transected rat spinal cord. The formation of a cellular lining in the scaffold channel may limit the degree of axonal regeneration. Spinal cords of adult rats were transected and implanted with the SC-loaded polylactic co-glycollic acid (PLGA) scaffold implants containing seven parallel-aligned channels, either 450 μm (n = 19) or 660 μm in diameter (n = 14). Animals were sacrificed after 1, 2 and 3 months. Immunohistochemistry for neurofilament expression was performed. The cross-sectional area of fibrous tissue and regenerative core was calculated. We found that the 450 μm scaffolds had significantly greater axon fibers per channel at the 1 month (186 ± 37) and 3 month (78 ± 11) endpoints than the 660 μm scaffolds (90 ± 19 and 40 ± 6, respectively) (p = 0.0164 and 0.0149, respectively). The difference in the area of fibrous rim between the 450 and 660 μm channels was most pronounced at the 1 month endpoint, at 28,046 ± 6551 and 58,633 ± 7063 μm2, respectively (p = 0.0105). Our study suggests that fabricating scaffolds with smaller diameter channels promotes greater regeneration over larger diameter channels. Axonal regeneration was reduced in the larger channels due to the generation of a large fibrous rim. Optimization of this scaffold environment establishes a platform for future studies of the effects of cell types, trophic factors or pharmacological agents on the regenerative capacity of the injured spinal cord.

Autologous nerve grafts are currently the best option for the treatment of segmental peripheral nerve defects. However, autografts have several drawbacks including size mismatch and loss of sensation in the donor nerve's sensory distribution. In this work, we have investigated the development of a synthetic hydrogel that contains positive charge for use as a substrate for nerve cell attachment and neurite outgrowth in culture. We have demonstrated that modification of oligo-(polyethylene glycol) fumarate (OPF) with a positively charged monomer improves primary sensory rat neuron attachment and differentiation in a dose-dependent manner. Positively charged hydrogels also supported attachment of dorsal root ganglion (DRG) explants that contain sensory neurons, Schwann cells and neuronal support cells. Furthermore, charged hydrogels were analyzed for the appearance of myelinated structures in a co-culture containing DRG neurons and Schwann cells. DRGs and Schwann cells remained viable on charged hydrogels for a time period of three weeks and neurites extended from the DRGs. Sudan black staining revealed that neurites emerging from DRGs were accompanied by migrating Schwann cells. These findings suggest that charged OPF hydrogels are capable of sustaining both primary nerve cells and the neural support cells that are critical for regeneration.

Two poly(ε-caprolactone fumarate)s (PCLFs) with distinct physical properties have been employed to prepare nanocomposites with hydroxyapatite (HA) nanoparticles via photo-crosslinking. The two PCLFs are PCLF530 and PCLF2000, named after their precursor PCL diol molecular weight of 530 and 2000 g.mol-1, respectively. Crosslinked PCLF530 is amorphous while crosslinked PCLF2000 is semi-crystalline with a melting temperature (Tm) of ∼40 °C and a crystallinity of 40%. Consequently, the rheological and mechanical properties of crosslinked PCLF2000 are significantly greater than those of crosslinked PCLF530. Structural characterizations and physical properties of both series of crosslinked PCLF/HA nanocomposites with HA compositions of 0%, 5%, 10%, 20%, and 30% have been investigated. By adding HA nanoparticles, crosslinked PCLF530/HA nanocomposites demonstrate enhanced rheological and mechanical properties while the enhancement in compressive modulus is less prominent in crosslinked PCLF2000/HA nanocomposites. In vitro cell attachment and proliferation have been performed using rat bone marrow stromal cells (BMSCs) and correlated with the material properties. Cell attachment and proliferation on crosslinked PCLF530/HA nanocomposite disks have been enhanced strongly with increasing the HA composition. However, surface morphology and surface chemistry such as composition, hydrophilicity, and the capability of adsorbing protein cannot be used to interpret the cell responses on different samples. Instead, the role of surface stiffness in regulating cell responses can be supported by the correlation between the change in compressive modulus and BMSC proliferation on these two series of crosslinked PCLFs and PCLF/HA nanocomposites.

In an effort to achieve suitable biomaterials for peripheral nerve regeneration, we present a material design strategy of combining a crystallite-based physical network and a crosslink-based chemical network. Biodegradable polymer disks and conduits have been fabricated by photo-crosslinking three poly(epsilon-caprolactone fumarate)s (PCLF530, PCLF1250, and PCLF2000), which were synthesized from the precursor poly(epsilon-caprolactone) (PCL) diols with nominal molecular weights of 530, 1250, and 2000 g mol(-1), respectively. Thermal properties such as glass transition temperature (T(g)), melting temperature (T(m)), and crystallinity of photo-crosslinked PCLFs were examined and correlated with their rheological and mechanical properties. Furthermore, in vitro degradation of uncrosslinked and crosslinked PCLFs in PBS crosslinked PCLFs in 1 N NaOH aqueous solution at 37 degrees C was studied. In vitro cytocompatibility, attachment, and proliferation of Schwann cell precursor line SPL201 cells on three PCLF networks were investigated. Crosslinked PCLF2000 with the highest crystallinity and mechanical properties was found to best support cell attachment and proliferation. Using a new photo-crosslinking method, single-lumen crosslinked PCLF nerve conduits without defects were fabricated in a glass mold. Crosslinked PCLF2000 nerve conduits were selected for evaluation in a 1cm gap rat sciatic nerve model. Histological evaluation demonstrated that the material was biocompatible with sufficient strength to hold sutures in place after 6 and 17 weeks of implantation. Nerve cable with myelinated axons was found in the crosslinked PCLF2000 nerve conduit.

Patients with malignant lumbosacral pelvic lesions present a difficult surgical challenge. Because of the insidious onset of symptoms, lesions are often diagnosed late in their course, and by that time they have attained a large size. Surgical resection is made more difficult by the complex surrounding anatomy and involvement of the sacral nerves responsible for bowel, bladder, and sexual function. Spinopelvic reconstruction is often required after resection. This article presents techniques for sacral resection and subsequent spinopelvic reconstruction. Biomechanical studies are summarized on construct stability, and recommendations are made as to when reconstruction is required. The expected bowel and bladder functional outcomes are summarized, based on the level of sacral resection.

Bone regeneration is a coordinated cascade of events regulated by several cytokines and growth factors. Angiogenic growth factors are predominantly expressed during the early phases for re-establishment of the vascularity, whereas osteogenic growth factors are continuously expressed during bone formation and remodeling. Since vascular endothelial growth factor (VEGF) and bone morphogenetic proteins (BMPs) are key regulators of angiogenesis and osteogenesis during bone regeneration, the aim of this study was to investigate if their sequential release could enhance BMP-2-induced bone formation. A composite consisting of poly(lactic-co-glycolic acid) microspheres loaded with BMP-2 embedded in a poly(propylene) scaffold surrounded by a gelatin hydrogel loaded with VEGF was used for the sequential release of the growth factors. Empty composites or composites loaded with VEGF and/or BMP-2 were implanted ectopically and orthotopically in Sprague-Dawley rats (n=9). Following implantation, the local release profiles were determined by measuring the activity of (125)I-labeled growth factors using scintillation probes. After 8 weeks blood vessel and bone formation were analyzed using microangiography, microCT and histology. The scaffolds exhibited a large initial burst release of VEGF within the first 3 days and a sustained release of BMP-2 over the full 56-day implantation period. Although VEGF did not induce bone formation, it did increase the formation of the supportive vascular network (p=0.03) in ectopic implants. In combination with local sustained BMP-2 release, VEGF significantly enhanced ectopic bone formation compared to BMP-2 alone (p=0.008). In the orthotopic defects, no effect of VEGF on vascularisation was found, nor was bone formation higher by the combination of growth factors, compared to BMP-2 alone. This study demonstrates that a sequential angiogenic and osteogenic growth factor release may be beneficial for the enhancement of bone regeneration.

Non-invasive imaging can provide essential information for the optimization of new drug delivery-based bone regeneration strategies to repair damaged or impaired bone tissue. This study investigates the applicability of nuclear medicine and radiological techniques to monitor growth factor retention profiles and subsequent effects on bone formation. Recombinant human bone morphogenetic protein-2 (BMP-2, 6.5 mu g/scaffold) was incorporated into a sustained release vehicle consisting of poly(lactic-co-glycolic acid) microspheres embedded in a poly(propylene fumarate) scaffold surrounded by a gelatin hydrogel and implanted subcutaneously and in 5-mm segmental femoral defects in 9 rats for a period of 56 days. To determine the pharmacokinetic profile, BMP-2 was radiolabeled with (125)I and the local retention of (125)I-BMP-2 was measured by single photon emission computed tomography (SPECT), scintillation probes and ex vivo scintillation analysis. Bone formation was monitored by micro-computed tomography (mu CT). The scaffolds released BMP-2 in a sustained fashion over the 56-day implantation period. A good correlation between the SPECT and scintillation probe measurements was found and there were no significant differences between the non-invasive and ex-vivo counting method after 8 weeks of follow up. SPECT analysis of the total body and thyroid counts showed a limited accumulation of 1251 within the body. Ectopic bone formation was induced in the scaffolds and the femur defects healed completely. In vivo mu CT imaging detected the first signs of bone formation at days 14 and 28 for the orthotopic and ectopic implants, respectively, and provided a detailed profile of the bone formation rate. Overall, this study clearly demonstrates the benefit of applying non-invasive techniques in drug delivery-based bone regeneration strategies by providing detailed and reliable profiles of the growth factor retention and bone formation at different implantation sites in a limited number of animals. (C) 2008 Elsevier B.V. All rights reserved.

Few data exist regarding the outcomes in patients undergoing surgery for pelvic tumors of neurogenic origin. Our aim was to characterize the clinical and pathologic features of pelvic neurogenic tumors and assess surgical outcomes. All patients who underwent operations for pelvic neurogenic tumors at our institution between 1956 and 2004 were identified. Data analyzed included demographics, clinical features, histopathology, local recurrence, and survival. Eighty-nine patients were identified, of whom 44 were male. Median age was 38 years. The most common presenting symptom was low back or pelvic pain (56%). Malignant lesions were found in 43 patients (48%). Schwannomas were the most common benign tumor (61%) and malignant peripheral nerve sheath tumors the most common malignant lesion (81%). Median tumor size was 9.5 cm (range 0.8–32 cm). Malignant tumors had histopathologic evidence of infiltration of surrounding structures in 49% of cases. Intralesional resection was the most common surgical technique for both benign and malignant tumors. Thirty-day mortality was nil; major morbidity was seen in 13%. Adjuvant therapy was given to 91% of the patients with malignant disease. Five-year local recurrence rates for benign and malignant lesions were 35.9% and 35.0%, respectively. Distant recurrence for malignant lesions was 65.1% at 5 years. Five-year disease-free survival for malignant tumors was 25.9%. Pelvic neurogenic tumors occurring in young patients may be large when detected and present with nonspecific symptoms. Benign and malignant tumors had a high local recurrence rate and survival for malignant tumors was poor. Early detection and aggressive surgical intervention should improve outcome.

We present a material design strategy of combining crystallinity and crosslinking to control the mechanical properties of polymeric biomaterials. Three polycaprolactone fumarates (PCLF530, PCLF1250, and PCLF2000) synthesized from the precursor polycaprolactone (PCL) diols with nominal molecular weights of 530, 1250, and 2000 g mol−1, respectively, were employed to fabricate polymer networks via photo-crosslinking process. Five different amounts of photo-crosslinking initiator were applied during fabrication in order to understand the role of photoinitiator in modulating the crosslinking characteristics and physical properties of PCLF networks. Thermal properties such as glass transition temperature (Tg), melting temperature (Tm), and degradation temperature (Td) of photo-crosslinked PCLFs were examined and correlated with their rheological and mechanical properties.

Complete spinal cord transection provides a valuable model for determining the cellular and molecular factors necessary for promoting axonal regeneration. Although contusion and compression models mimic the types of injury that occur most frequently in patients,1 interpretation of results using these models can be complicated by the occurrence of axonal regeneration and sprouting from distal segments of surviving axons. The use of biodegradable scaffolds in transection injuries allows separate manipulation of the 3-dimensional structural, cellular, and molecular environment of the regenerating cord. An 85:15 polylactic acid: poly-lactic-co-glycolic acid (PLGA) scaffold has been shown to maintain its structural integrity for at least 8 weeks and starts to degrade significantly by 24 weeks, thereby providing an adequate time-frame for axonal regeneration.2 This type of scaffold has been used in nerve and bone repair and in brain with few adverse effects.3–6 Biodegradable polymer inserts have also been extensively used in animal models to promote axonal regeneration in the peripheral nervous system,7–10 which has subsequently led to the development of suitable models for the central nervous system.2,11,12 Axonal regeneration was demonstrated through channels created in PLGA scaffolds that were loaded with Schwann cells and inserted into the spinal cord.2 However, the resulting weakness that occurs at and below the level of transection can lead to the development of kypho-scoliosis in a significant number of animals. This, in turn, can compromise the alignment and stability of the implanted scaffold, which can hinder interpretation of the resulting axonal regeneration and functional recovery. Spinal stabilization offers a solution to this problem. Fixation of the vertebral column has been shown to improve neurologic function in clinical spinal cord surgery.13 However, many animal models of SCI currently do not incorporate a fixation procedure, despite its reported benefits.14 This study was undertaken to assess the effect of a novel method of spinal fixation on scoliosis, kyphosis, and scaffold alignment in an animal transection model of SCI. Radiographic and 3-dimensional-magnetic resonance microscopy (MRM) analysis were used to quantify the degree of spinal curvature and malalignment of the scaffold within the cord. Radiographic visualization of the scaffold within the cord was achieved by radio-opaque barium sulfate impregnation. The barium sulfate did not interfere with MRM imaging.

Objective The accuracy of motor axon regeneration becomes an important issue in the development of a nerve tube for motor nerve repair. Dispersion of regeneration across the nerve tube may lead to misdirection and polyinnervation. In this study, we present a series of methods to investigate the accuracy of regeneration, which we used to compare regeneration across autografts and single-lumen poly(lactic-co-glycolic acid) (PLGA) nerve tubes. We also present the concept of the multichannel nerve tube that may limit dispersion by separately guiding groups of regenerating axons. Methods The simultaneous tracing of the tibial and peroneal nerves with fast blue and diamidino yellow was performed 8 weeks after the repair of a 1-cm nerve gap in the rat sciatic nerve to determine the percentage of double-projecting motoneurons. Sequential tracing of the peroneal nerve with diamidino yellow 1 week before repair and fast blue 8 weeks after repair was performed to determine the percentage of correctly directed peroneal motoneurons. Results In the cases in which there was successful regeneration across single-lumen nerve tubes, more motoneurons had double projections to both the tibial and peroneal nerve branches after single-lumen nerve tube repair (21.4%) than after autograft repair (5.9%). After multichannel nerve tube repair, this percentage was slightly reduced (16.9%), although not significantly. The direction of regeneration was nonspecific after all types of repair. Conclusion Retrograde tracing techniques provide new insights into the process of regeneration across nerve tubes. The methods and data presented in this study can be used as a basis for the development of a nerve tube for motor nerve repair.

This study describes investigation of porous photocrosslinked oligo[(polyethylene glycol) fumarate] (OPF) hydrogels as potential matrix for osteoblastic differentiation of marrow stromal cells (MSCs). The porosity and interconnectivity of porous hydrogels were assessed using magnetic resonance microscopy (MRM) as a noninvasive investigative tool that could image the water construct inside the hydrogels at a high spatial resolution. MSCs were cultured onto the porous hydrogels and cell number was assessed using PicoGreen DNA assay. Our results showed 10% of cells initially attached to the surface of scaffolds. However, cells did not show significant proliferation over a time period of 14 days. MSCs cultured on porous hydrogels had increased alkaline phosphatase activity as well as deposition of calcium, suggesting successful differentiation and maturation to the osteoblastic phenotype. Moreover, continued expression of type I collagen and osteonectin over 14 days confirmed osteoblastic differentiation of MSCs. MRM was also applied to monitor osteogenesis of MSCs on porous hydrogels. MRM images showed porous scaffolds became consolidated with osteogenic progression of cell differentiation. These findings indicate that porous OPF scaffolds enhanced MSC differentiation leading to development of bone-like mineralized tissue.

Study Design. Anatomic study of the internal carotid artery (ICA) location with respect to C1 based on computed tomography (CT) scans with contrast medium. Objective. To measure the location of the ICA relative to the anterior aspect of C1 to assess the risk of placing C1–C2 transarticular or C1 lateral mass screws. Summary of Background Data. Vertebral artery injury is a known risk from placement of screws in C1. A previous case report revealed an ideally placed C1–C2 transarticular screw abutting and narrowing the ICA. The risk of ICA injury from C1 screws is unknown. Methods. Fifty random head and neck CT scans with contrast medium were retrospectively analyzed. Measurements were taken bilaterally including the closest distance from the ICA lumen to C1 and the distance from the medial edge of the ICA to a line drawn along the medial border of the foramen transversarium. The risk of inserting bicortical C1–C2 transarticular and C1 lateral mass screws was estimated based on these measurements. Results. The mean distance from the ICA to C1 was 2.88 mm on the left and 2.89 mm on the right. The ICA lumen was medial to the foramen transversarium in 42 (84%) of 50 cases (mean: 2.78 mm on the left and 3.00 mm on the right). The proximity of the ICA to C1 posed moderate risk in 46% of cases and high risk in 12% (on at least one side). Conclusion. Because of the risk of ICA injury from a drill bit or the tip of a bicortical screw, we recommend preoperative CT scan with contrast medium in all cases in which a screw is to be placed into C1. If the ICA is in close proximity to the anterior border of C1, unicortical fixation or a different fusion technique should be considered.

Aiming to achieve suitable polymeric biomaterials with controlled physical properties for hard and soft tissue replacements, we have developed a series of blends consisting of two photo-cross-linkable polymers: polypropylene fumarate (PPF) and polycaprolactone fumarate (PCLF). Physical properties of both un-cross-linked and UV cross-linked PPF/PCLF blends with PPF composition ranging from 0% to 100% have been investigated extensively. It has been found that the physical properties such as thermal, rheological, and mechanical properties could be modulated efficiently by varying the PPF composition in the blends. Thermal properties including glass transition temperature (T g) and melting temperature (T m) have been correlated with their rheological and mechanical properties. Surface characteristics such as surface morphology, hydrophilicity, and the capability of adsorbing serum protein from culture medium have also been examined for the cross-linked polymer and blend disks. For potential applications in bone and nerve tissue engineering, in vitro cell studies including cytotoxicity, cell adhesion, and proliferation on cross-linked disks with controlled physical properties have been performed using rat bone marrow stromal cells and SPL201 cells, respectively. In addition, the role of mechanical properties such as surface stiffness in modulating cell responses has been emphasized using this model blend system.

When the normal physiologic reaction to fracture does not occur, such as in fracture nonunions or large-scale traumatic bone injury, surgical intervention is warranted. Autografts and allografts represent current strategies for surgical intervention and subsequent bone repair, but each possesses limitations, such as donor-site morbidity with the use of autograft and the risk of disease transmission with the use of allograft. Synthetic bone-graft substitutes, developed in an effort to overcome the inherent limitations of autograft and allograft, represent an alternative strategy. These synthetic graft substitutes, or matrices, are formed from a variety of materials, including natural and synthetic polymers, ceramics, and composites, that are designed to mimic the three-dimensional characteristics of autograft tissue while maintaining viable cell populations. Matrices also act as delivery vehicles for factors, antibiotics, and chemotherapeutic agents, depending on the nature of the injury to be repaired. This intersection of matrices, cells, and therapeutic molecules has collectively been termed tissue engineering. Depending on the specific application of the matrix, certain materials may be more or less well suited to the final structure; these include polymers, ceramics, and composites of the two. Each category is represented by matrices that can form either solid preformed structures or injectable forms that harden in situ. This article discusses the myriad design considerations that are relevant to successful bone repair with tissue-engineered matrices and provides an overview of several manufacturing techniques that allow for the actualization of critical design parameters.

Multichannel conduits have been developed for experimental peripheral nerve and spinal cord repair. We present a series of methods to characterize multichannel nerve tubes for properties of bending, deformation, swelling, and degradation and introduce a new method to test the permeability of multichannel nerve tubes from the rate of diffusion of different-sized fluorescent dextran molecules (10, 40, and 70 kDa). First, single-lumen nerve tubes made with different poly(lactic-co-glycolic acid) (PLGA) ratios (50:50, 75:25, and 85:15) were compared. One ratio (75:25 PLGA) was subsequently used to compare single-lumen and multichannel nerve tubes. Nerve tubes made with lower PLGA ratios were found to be more flexible than nerve tubes made with a higher PLGA ratio. For low ratios, however, swelling was also greater as a result of a faster rate of degradation. Multichannel structure did not interfere with the permeability of the tube; the rate of diffusion into multichannel 75:25 PLGA nerve tubes appeared to be even higher than that into single-lumen ones, but this was only significant for 70-kDa molecules. Also, multichannel 75:25 PLGA nerve tubes were more flexible and, at the same time, more resistant to deformation. However, swelling significantly decreased the total cross-sectional lumen area, especially in multichannel 75:25 PLGA nerve tubes. Permeability, bending, deformation, swelling, and degradation are important properties to characterize in the development of multichannel nerve tubes. The methods presented in this study can be used as a basis for optimizing these properties for future, possibly clinical, application.

The use of bone grafts for orthopedic applications have increased steadily over the past decade. With improvements in surgical technique, combined with an increasing aged population requiring orthopedic treatment, the need for bone grafts substitutes have also increased. To be useful clinically, the bone graft substitute must be biocompatible, bioabsorbable, and have convenient handling properties. In addition, it must possess a microarchitecture that allows cellular ingrowth and remodeling while simultaneously providing mechanical strength. Poly(propylene fumarate) (PPF) has been investigated as an injectable, biodegradable scaffold for orthopedic applications. Various methods to create a porous, interconnected polymer scaffold are available. The foaming technique is a convenient method to accomplish this task. Reactions between bicarbonate salts and weak acids generate CO2 gas, causing a bubbling reaction during the polymerization process. This technique allows the porosity of the scaffold to be modulated. Image analysis and mechanical testing of porous PPF fabricated using the foaming technique shows that a highly porous, interconnected scaffold can be produced. At ∼50% porosity, the scaffold has excellent handling properties, contains pore sizes ranging from 50 to 500 μm with an elastic modulus ranging from 20 to 40 MPa. The foaming technique provides an additional method by which clinically useful polymers can be fabricated for use in various bone tissue engineering applications. © 2007 Wiley Periodicals, Inc. J Biomed Mater Res, 2008

Ankle motion analysis may provide a better method to assess function in the rat sciatic nerve model than the standard method, the sciatic functional index (SFI), but it is not widely used in experiments on nerve regeneration possibly because of complicated analysis. In this study, we investigated the practical use of a two-dimensional (2D) digital video motion analysis system. Reproducibility was investigated in normal rats. Recovery of ankle motion was analyzed after sciatic, tibial, and peroneal nerve crush injury. Results were compared with scores for the SFI. Results were not significantly different from animal-to-animal and day-to-day. Interobserver variability also was small. In the analysis of recovery after separate nerve crush injuries, subtle differences in ankle plantar flexion and dorsiflexion could be detected. The method was also more sensitive than the SFI: whereas scores for the SFI had returned to normal 4 weeks after sciatic nerve crush injury, the ankle angle at mid-stance was still significantly different from that in sham-operated animals 6 weeks after the injury. 2D digital video ankle motion analysis is a practical and sensitive method to assess function in the rat sciatic nerve model.

This article reports a simple one-step method of attaching folic acid (FA) to gold nanoparticles (AuNPs) and its fine tuning using different polyethylene glycol (PEG) backbones. PEG backbones used in this study are PEG-diamine with molecular weights 2000 (PAM2-2K) and 10,000 (PAM2-10K), PEG-tetramine with molecular weight 20,000 (PAM4-20K), and PEG-dithiol with molecular weight 2000 (PSH2-2K). The nanoconjugates were characterized with ultraviolet-visible spectroscopy, transmission electron microscopy (TEM), thermogravimetric analysis, Fourier transform infrared spectroscopy, inductively coupled plasma analysis, and radioactivity measurement with a scintillation counter. Attachment and release profiles of FAs from the nanoconjugates are done using 3 H-labelled FAs ( 3 FA). The binding of 3 FA follows the order Au-PAM4-20K > Au-PAM2-10K > Au-PAM2-2K > Au > Au-PSH2-2K, whereas its release profile follows the reverse order. Au-PAM4-20K-FA has been used for folate receptor (FR)–mediated targeting of AuNPs to cancer cells. Seven different cancer cell lines (SKOV-3, OVCAR-5, OV-202, OV-167, OPM-1, RPMI, and U266) were screened for expression of FRs. Among ovarian cancer cells, the expression pattern of FRs follows the order OV-167 > SKOV-3 > OV-202 > OVCAR-5, and multiple myeloma cell lines follow the order OPM-1 > U266 > RPMI. Intracellular uptakes of the nanoconjugates containing FA or no FA were monitored with digital optical photography and TEM. Quantitation of the internalization of nanoconjugates in different cell lines was determined by gold analysis with inductively coupled plasma. It is found that the uptake of the nanoconjugates correlates with FR expression. Maximum uptake is observed for OV-167, whereas it is minimum for OVCAR-5. TEM images of the cells treated with Au-PAM4-20K-FA confirm the endocytosis of the nanoconjugates. This study is an important step for targeted delivery of anticancer drugs as well as metal nanoparticles for targeted therapy, tumor imaging, and ablation exploiting the overexpression of FRs on cancer cells.

BACKGROUND: Others have reported significant changes in red blood cell (RBC) transfusion practice during the past two decades during knee, hip, prostate, and carotid surgery. Similar data for patients undergoing major spine surgery, however, are not available. STUDY DESIGN AND METHODS: After institutional review board approval, adult patients undergoing elective major spine surgery were stratified into one of two transfusion-related groups: 1) 1980 to 1985 (i.e., before human immunodeficiency virus screening, early practice group; n = 699) or 2) 1995 to 2000 (i.e., late practice group; n = 610). RESULTS: Patients in the late practice group were older, had greater numbers of preoperative coexisting diseases (e.g., hypertension, cardiac dysrhythmias, coronary artery disease, prior myocardial infarction, diabetes mellitus, renal disease, cerebrovascular disease, and asthma), and were exposed to longer operations (p

Photo-cross-linkable oligo[poly(ethylene glycol) fumarate] (OPF) hydrogels have been developed for use in tissue engineering applications. We demonstrated that compressive modulus of these hydrogels increased with increasing polymer concentration, and hydrogels with different mechanical properties were formed by altering the ratio of cross-linker/polymer in precursor solution. Conversely, swelling of hydrogels decreased with increasing polymer concentration and cross-linker/polymer ratio. These hydrogels are degradable and degradation rates vary with the change in cross-linking level. Chondrocyte attachment was quantified as a method for evaluating adhesion of cells to the hydrogels. These data revealed that cross-linking density affects cell behavior on the hydrogel surfaces. Cell attachment was greater on the samples with increased cross-linking density. Chondrocytes on these samples exhibited spread morphology with distinct actin stress fibers, whereas they maintained their rounded morphology on the samples with lower cross-linking density. Moreover, chondrocytes were photoencapsulated within various hydrogel networks. Our results revealed that cells encapsulated within 2-mm thick OPF hydrogel disks remained viable throughout the 3-week culture period, with no difference in viability across the thickness of hydrogels. Photoencapsulated chondrocytes expressed the mRNA of type II collagen and produced cartilaginous matrix within the hydrogel constructs after three weeks. These findings suggest that photo-cross-linkable OPF hydrogels may be useful for cartilage tissue engineering and cell delivery applications.

Our approach to cartilage tissue-engineering scaffolds combines image-based design and solid free-form (SFF) fabrication to create load-bearing constructs with user-defined parameters. In this study, 3-dimensional scaffolds with cubic and ellipsoidal pore architecture were fabricated using poly(propylene fumarate) (PPF). To increase seeding efficiency and cellular retention, hydrogels were used to deliver cells into the scaffolds. The first objective of this study was to evaluate the concentrations of composite hyaluronic acid (HyA) and collagen I hydrogels best able to stimulate proteoglycan synthesis in porcine chondrocytes in vitro and in vivo. The second objective was to evaluate the differences in extracellular matrix production due to pore geometry and scaffold design. For the in vitro assessment, chondrocytes were encapsulated in collagen I hydrogels with varying concentrations of HyA. Hydrogels were cultured for 1 and 2 weeks, and then the sulfated glycosaminoglycan (sGAG) content was quantified using a dimethyl-methylene blue assay. The concentration of HyA best able to increase ECM synthesis was 5% HyA/collagen I, or 0.23 mg/mL HyA. The results from the in vitro experiment were used as culture parameters for the in vivo analysis. Composite 5% HyA/collagen I or collagen I-only hydrogels were used to seed chondrocytes into SFF-fabricated scaffolds made of PPF with designed cubic or ellipsoidal pore geometry. The scaffolds were implanted subcutaneously in immunocompromised mice for 4 weeks. Histomorphometric analyses of sections stained with Safranin O were used to quantify the amount of ECM deposited by cells in the scaffolds. Scaffolds seeded with 5% HyA/collagen hydrogels had significantly greater areas of positive Safranin O staining (approximately 60%, compared with 30% for scaffolds with collagen I hydrogels only), indicating that greater numbers of chondrocytes retained their metabolic activity in the ectopic environment. These scaffolds also had greater stain intensities (corresponding to greater amounts of sGAG in the ECM) than their counterparts seeded with collagen I hydrogels alone. Significant differences in matrix production were not found between the scaffold pore designs. Overall, these results indicate that a combination of composite HyA hydrogels and designed SFF scaffolds could provide a functional tissue-engineered construct for cartilage repair with enhanced tissue regeneration in a load-bearing scaffold.

Stereolithography using photo-cross-linkable polymeric biomaterials is an effective technique for fabricating highly complex three-dimensional (3D) scaffolds with controlled microstructures for tissue engineering applications. In this study, we have optimized the UV curable polymer solution composition and laser parameters for the stereolithography machine. Poly(propylene fumarate) (PPF) was used as the biomaterial, diethyl fumarate (DEF) was used as the solvent, and bisacrylphosphrine oxide (BAPO) was used as the photoinitiator. Three different weight ratios of PPF/DEF and BAPO contents were characterized by measuring the viscosities and thermal properties of the un-cross-linked solutions and the mechanical properties of the formed scaffolds. After optimizing the resin composition by satisfying both the viscosity limitation and the mechanical requirement, laser parameters such as critical exposure (Ec) and penetration depth (Dp) were determined from the working curve and the relationship between laser speed and energy by measuring the thickness of predesigned windows fabricated in stereolithography with different ranges of Ec and Dp. Three-dimensional scaffolds with various pore sizes, pore shapes, and porosities were designed in computer-aided design (CAD) software and were fabricated in stereolithography. The fabricated scaffolds were characterized by measuring external dimensions, porosities, mean pore sizes, and compressive moduli and were compared to the CAD models. Feature accuracy in the xy-plane was achieved and overcuring of the resin in z-axis was minimized. The stereolithographically fabricated scaffolds with controlled microstructures can be useful in diverse tissue engineering applications.

Chondrosarcoma is a cartilage tumor and is the second most common malignant bone cancer. Unlike many tumors, chondrosarcomas are resistant to conventional chemotherapy and radiotherapy. Autophagy is a homeostatic mechanism through which cellular proteins and organelles are subjected to lysosomal degradation and recycling. Autophagy could play a dual role in cancer by facilitating either cell death or cell survival. To determine whether autophagy plays a role in cell death in chondrosarcoma, we have studied the effect of the anti-tumor compound 2-methoxyestradiol (2-ME) in chondrosarcoma cells in culture. Transmission electron microscopy imaging indicates that 2-ME treatment leads to the accumulation of autophagosomes in human chondrosarcoma (SW1353 and Hs819T) cells. Also, 2-ME induces the conversion of microtubule-associated protein LC3-I to LC3-II, a protein marker that is correlated with the formation of autophagosomes. Our results show that siRNAs directed against ATG3 blocks 2-ME-induced autophagosome formation in chondrosarcoma cells. In addition, treatment with Bafilomycin A1 (Baf) and 3-methyladenine (3-MA), the inhibitors of autophagy, further increased the cell death in 2-ME-treated chondrosarcoma cells. Taken together, our studies demonstrate that autophagy causes resistance to cytotoxicity in chondrosarcoma cells, and the efficacy and anti-tumor effects of drugs in chondrosarcoma could be enhanced by modulating autophagy.

There is a need to improve the prediction of fracture risk for patients with metastatic bone disease. CT-based rigidity analysis (CTRA) is a sensitive and specific method, yet its influence on clinical decision-making has never been quantified. What is the influence of CTRA on providers’ perceived risk of fracture? (2) What is the influence of CTRA on providers’ treatment recommendations in simulated clinical scenarios of metastatic bone disease of the femur? (3) Does CTRA improve interobserver agreement regarding treatment recommendations? We conducted a survey among 80 academic physicians (orthopaedic oncologists, musculoskeletal radiologists, and radiation oncologists) using simulated vignettes of femoral lesions presented as three separate scenarios: (1) no CTRA input (baseline); (2) CTRA input suggesting increased risk of fracture (CTRA+); and (3) CTRA input suggesting decreased risk of fracture (CTRA−). Participants were asked to rate the patient’s risk of fracture on a scale of 0% to 100% and to provide a treatment recommendation. Overall response rate was 62.5% (50 of 80). When CTRA suggested an increased risk of fracture, physicians perceived the fracture risk to be slightly greater (37% ± 3% versus 42% ± 3%, p < 0.001; mean difference [95% confidence interval {CI}] = 5% [4.7%–5.2%]) and were more prone to recommend surgical stabilization (46% ± 9% versus 54% ± 9%, p < 0.001; mean difference [95% CI] = 9% [7.9–10.1]). When CTRA suggested a decreased risk of fracture, physicians perceived the risk to be slightly decreased (37% ± 25% versus 35% ± 25%, p = 0.04; mean difference [95% CI] = 2% [2.74%–2.26%]) and were less prone to recommend surgical stabilization (46% ± 9% versus 42% ± 9%, p < 0.03; mean difference [95% CI] = 4% [3.9–5.1]). The effect size of the influence of CTRA on physicians’ perception of fracture risk and treatment planning varied with lesion severity and specialty of the responders. CTRA did not increase interobserver agreement regarding treatment recommendations when compared with the baseline scenario (κ = 0.41 versus κ = 0.43, respectively). Based on this survey study, CTRA had a small influence on perceived fracture risk and treatment recommendations and did not increase interobserver agreement. Further work is required to properly introduce this technique to physicians involved in the care of patients with metastatic lesions. Given the number of preclinical and clinical studies outlining the efficacy of this technique, better education through presentations at seminars/webinars and symposia will be the first step. This should be followed by clinical trials to establish CTRA-based clinical guidelines based on evidence-based medicine. Increased exposure of clinicians to CTRA, including its underlying methodology to study bone structural characteristics, may establish CTRA as a uniform guideline to assess fracture risk. Level III, economic and decision analyses.

In this article, a "bedside to bench and back" approach for developing tissue engineered medical products (TEMPs) for clinical applications is reviewed. The driving force behind this approach is unmet clinical needs. Preclinical research, both in vitro and in vivo using small and large animal models, will help find solutions to key research questions. In clinical research, ethical issues regarding the use of cells and tissues, their sources, donor consent, as well as clinical trials are important considerations. Regulatory issues, at both institutional and government levels, must be addressed prior to the translation of TEMPs to clinical practice. TEMPs are regulated as drugs, biologics, devices, or combination products by the U.S. Food and Drug Administration (FDA). Depending on the mode of regulation, applications for TEMP introduction must be filed with the FDA to demonstrate safety and effectiveness in premarket clinical studies, followed by 510(k) premarket clearance or premarket approval (for medical devices), biologics license application approval (for biologics), or new drug application approval (for drugs). A case study on nerve cuffs is presented to illustrate the regulatory process. Finally, perspectives on commercialization such as finding a company partner and funding issues, as well as physician culture change, are presented.