Capacitive Electrode-Based Electric Field Treatments on Redox-Toxic Iron Deposits in Transgenic AD Mouse Models: The Electroceutical Targeting of Alzheimer's Disease Feasibility Study.

Iron accumulation in the brain accelerates Alzheimer's disease progression. To cure iron toxicity, we assessed the therapeutic effects of noncontact transcranial electric field stimulation to the brain on toxic iron deposits in either the Aß fibril structure or the Aß plaque in a mouse model of Alzheimer's disease (AD) as a pilot study. A capacitive electrode-based alternating electric field (AEF) was applied to a suspension of magnetite (Fe3O4) to measure field-sensitized reactive oxygen species (ROS) generation. The increase in ROS generation compared to the untreated control was both exposure-time and AEF-frequency dependent. The frequency-specific exposure of AEF to 0.7-1.4 V/cm on a magnetite-bound Aß-fibril or a transgenic Alzheimer's disease (AD) mouse model revealed the degradation of the Aß fibril or the removal of the Aß-plaque burden and ferrous magnetite compared to the untreated control. The results of the behavioral tests show an improvement in impaired cognitive function following AEF treatment on the AD mouse model. Tissue clearing and 3D-imaging analysis revealed no induced damage to the neuronal structures of normal brain tissue following AEF treatment. In conclusion, our results suggest that the effective degradation of magnetite-bound amyloid fibrils or plaques in the AD brain by the electro-Fenton effect from electric field-sensitized magnetite offers a potential electroceutical treatment option for AD.

A Wireless Pressure Sensor Integrated with a Biodegradable Polymer Stent for Biomedical Applications.

This paper describes the fabrication and characterization of a wireless pressure sensor for smart stent applications. The micromachined pressure sensor has an area of 3.13 × 3.16 mm² and is fabricated with a photosensitive SU-8 polymer. The wireless pressure sensor comprises a resonant circuit and can be used without the use of an internal power source. The capacitance variations caused by changes in the intravascular pressure shift the resonance frequency of the sensor. This change can be detected using an external antenna, thus enabling the measurement of the pressure changes inside a tube with a simple external circuit. The wireless pressure sensor is capable of measuring pressure from 0 mmHg to 230 mmHg, with a sensitivity of 0.043 MHz/mmHg. The biocompatibility of the pressure sensor was evaluated using cardiac cells isolated from neonatal rat ventricular myocytes. After inserting a metal stent integrated with the pressure sensor into a cardiovascular vessel of an animal, medical systems such as X-ray were employed to consistently monitor the condition of the blood vessel. No abnormality was found in the animal blood vessel for approximately one month. Furthermore, a biodegradable polymer (polycaprolactone) stent was fabricated with a 3D printer. The polymer stent exhibits better sensitivity degradation of the pressure sensor compared to the metal stent.

Characterizing the effects of heparin gel stiffness on function of primary hepatocytes.

In the liver, hepatocytes are exposed to a large array of stimuli that shape hepatic phenotype. This in vivo microenvironment is lost when hepatocytes are cultured in standard cell cultureware, making it challenging to maintain hepatocyte function in vitro. Our article focused on one of the least studied inducers of the hepatic phenotype-the mechanical properties of the underlying substrate. Gel layers comprised of thiolated heparin (Hep-SH) and diacrylated poly(ethylene glycol) (PEG-DA) were formed on glass substrates via a radical mediated thiol-ene coupling reaction. The substrate stiffness varied from 10 to 110 kPa by changing the concentration of the precursor solution. ELISA analysis revealed that after 5 days, hepatocytes cultured on a softer heparin gel were synthesizing five times higher levels of albumin compared to those on a stiffer heparin gel. Immunofluorescent staining for hepatic markers, albumin and E-cadherin, confirmed that softer gels promoted better maintenance of the hepatic phenotype. Our findings point to the importance of substrate mechanical properties on hepatocyte function.

Factors predicting hamstring tendon autograft diameters and resulting failure rates after anterior cruciate ligament reconstruction.

PURPOSE: The purposes of this study are to confirm factors that affect the diameter of hamstring tendon autograft and to compare failure rates between the factors after anterior cruciate ligament (ACL) reconstruction. METHODS: A total of 296 patients that underwent reconstruction using hamstring tendon autograft at our clinics for ACL injury between September 2005 and June 2008 were enrolled for this study. The diameters of gracilis and semitendinosus tendons (harvested from the affected knee) and four-strand graft tendon made by folding the gracilis and semitendinosus tendons in two layers were measured. Before operating, we recorded the age, height, weight, Body Mass Index (BMI), gender and athlete versus non-athlete identity of the subjects and checked their correlations with graft diameters. Patients that recorded a grade C or D on the International Knee Documentation Committee Knee Examination Form, as well as patients that underwent revision, were defined as failures and analysed by related factors. RESULTS: The mean diameter was 1.5 mm ± 0.2 for gracilis tendon, 2.2 mm ± 0.3 for semitendinosus tendon and 7.2 mm ± 0.7 for graft tendon. Except for age, factors including height, weight, BMI, gender and athlete versus non-athlete identity were found to be significantly related to graft diameter. Correlation was strongest with height (p < 0.001). With respect to failure rates after ACL reconstruction, patients with a graft diameter of 8.0 mm or more demonstrated statistically better results than patients with a diameter of below 8.0 mm (p = 0.043). However, failure rates did not differ significantly with respect to other factors. CONCLUSIONS: The diameter of hamstring tendon autograft may be different depending on height, weight, BMI and gender of the patient, as well as whether or not the patient is an athlete. Although we did not find statistically significant differences in failure rates after ACL reconstruction, this study demonstrated relatively better results in patients with a graft diameter of 8.0 mm or more. LEVEL OF EVIDENCE: Case series, Level IV.

Combination therapy with BMP-2 and BMSCs enhances bone healing efficacy of PCL scaffold fabricated using the 3D plotting system in a large segmental defect model.

The three-dimensional (3D) plotting system is a rapidly-developing scaffold fabrication method for bone tissue engineering. It yields a highly porous and inter-connective structure without the use of cytotoxic solvents. However, the therapeutic effects of a scaffold fabricated using the 3D plotting system in a large segmental defect model have not yet been demonstrated. We have tested two hypotheses: whether the bone healing efficacy of scaffold fabricated using the 3D plotting system would be enhanced by bone marrow-derived mesenchymal stem cell (BMSC) transplantation; and whether the combination of bone morphogenetic protein-2 (BMP-2) administration and BMSC transplantation onto the scaffold would act synergistically to enhance bone regeneration in a large segmental defect model. The use of the combined therapy did increase bone regeneration further as compared to that with monotherapy in large segmental bone defects.

Effect of Animal Condition and Fluvoxamine on the Result of [(18)F]N-3-Fluoropropyl-2ß-carbomethoxy-3ß-(4-iodophenyl) Nortropane ([(18)F]FP-CIT) PET Study in Mice.

PURPOSE: PET (positron emission tomography) is a noninvasive imaging technique, visualizing biological aspects in vivo. In animal models, the result of PET study can be affected more prominently than in humans by the animal conditions or drug pretreatment. We assessed the effects of anesthesia, body temperature, and pretreatment with selective serotonin reuptake inhibitor on the results of [(18)F]N-3-fluoropropyl-2ß-carbomethoxy-3ß-(4-iodophenyl) nortropane ([(18)F]FP-CIT) PET in mice. METHODS: [(18)F]FP-CIT PET of C57BL/6 mice was performed in three different conditions: (1) anesthesia (isoflurane) with active warming (38°C) as a reference; (2) no anesthesia or warming; (3) anesthesia without warming at room temperature. Additional groups of mice pretreated with escalating doses of fluvoxamine (5, 20, 40, 80 mg/kg) were imaged in condition (1). The time activity curve and standardized uptake value of the striatum, cerebral cortex, and bone were compared among these conditions. RESULTS: In all conditions, radioactivities of the striatum and cortex tended to form a plateau after rapid uptake and washout, but that of bone tended to increase gradually. When anesthetized without any warming, all the mice developed hypothermia and showed reduced bone uptake with slightly increased striatal and cortical uptakes compared to the reference condition. In conditions without anesthesia, striatal and cortical uptakes were reduced, whereas the bone uptake showed no change. Pretreatment with fluvoxamine increased the striatal uptake and striatal specific to cortical non-specific uptake ratio, whereas the bone uptake was reduced. CONCLUSION: Anesthesia, body temperature, and fluvoxamine affect the result of [(18)F]FP-CIT PET in mice by altering striatal and bone uptakes.

3D polycaprolactone scaffolds with controlled pore structure using a rapid prototyping system.

Designing a three-dimensional (3-D) ideal scaffold has been one of the main goals in biomaterials and tissue engineering, and various mechanical techniques have been applied to fabricate biomedical scaffolds used for soft and hard tissue regeneration. Scaffolds should be biodegradable and biocompatible, provide temporary support for cell growth to allow cell adhesion, and consist of a defined structure that can be formed into customized shapes by a computer-aided design system. This versatility in preparing scaffolds gives us the opportunity to use rapid prototyping devices to fabricate polymeric scaffolds. In this study, we fabricated polycaprolactone scaffolds with interconnecting pores using a 3-D melt plotting system and compared the plotted scaffolds to those made by salt leaching. Scanning electron microscopy, a laser scanning microscope, micro-computed tomography, and dynamic mechanical analysis were used to characterize the geometry and mechanical properties of the resulting scaffolds and morphology of attached cells. The plotted scaffolds had the obvious advantage that their mechanical properties could be easily manipulated by adjusting the scaffold geometry. In addition, the plotted scaffolds provided more opportunity for cells to expand between the strands of the scaffold compared to the salt-leached scaffold.

In vitro and animal study of novel nano-hydroxyapatite/poly(epsilon-caprolactone) composite scaffolds fabricated by layer manufacturing process.

The purpose of this study was to propose a computer-controllable scaffold structure made by a layer manufacturing process (LMP) with addition of nano- or micro-sized particles and to investigate the effects of particle size in vitro. In addition, the superiority of this LMP method over the conventional scaffolds made by salt leaching and gas forming process was investigated through animal study. Using the LMP, we have created a new nano-sized hydroxyapatite/poly(epsilon-caprolactone) composite (n-HPC) scaffold and a micro-sized hydroxyapatite/poly(epsilon-caprolactone) composite (m-HPC) scaffold for bone tissue engineering applications. The scaffold macropores were well interconnected, with a porosity of 73% and a pore size of 500 microm. The compressive modulus of the n-HPC and m-HPC scaffolds was 6.76 and 3.18 MPa, respectively. We compared the cellular responses to the two kinds of scaffolds. Both n-HPC and m-HPC exhibited good in vitro biocompatibility. Attachment and proliferation of mesenchymal stem cells were better on the n-HPC than on the m-HPC scaffold. Moreover, significantly higher alkaline phosphatase activity and calcium content were observed on the n-HPC than on the m-HPC scaffold. In an animal study, the LMP scaffolds enhanced bone formation, owing to their well-interconnected pores. Radiological and histological examinations confirmed that the new bony tissue had grown easily into the entire n-HPC scaffold fabricated by LMP. We suggest that the well-interconnected pores in the LMP scaffolds might encourage cell attachment, proliferation, and migration to stimulate cell functions, thus enhancing bone formation in the LMP scaffolds. This study shows that bioactive and biocompatible n-HPC composite scaffolds prepared using an LMP have potential applications in bone tissue engineering.

Effects of intermittent hydrostatic pressure on cell adhesive forces and other related parameters under various resting periods.

The purpose of this study is to investigate the effects of intermittent hydrostatic pressure with various resting periods on the cell adhesive forces and other parameters related to spreading in early stage of cell adhesion. For this, bovine pulmonary arterial endothelium (CPAE, cell line), porcine articular chondrocytes, and human endothelial cells (HECs) were used. The cells were divided into six different experimental groups. Control group was cultured without stimulation, while the constant pressure was applied to group 1 for 2 h. Groups 2-5 were intermittently pressurized for 2 min at a time over a 2-h period with 5, 10, 15, and 20-min resting periods, respectively. Each group was then split into two subgroups, depending whether it experienced extra 60 min stabilization period after stimulation. The average adhesive force and the number and area of focal contacts were significantly higher in the group 4 subgroup, which received an extra 60 min of culture than in the other groups. Similarly, other parameters in this subgroup were significantly different from those in the other groups. The focal contact area and adhesive force were closely related (r = 0.990). We concluded that the mechanical stimuli affect cell adhesion and that the length of the resting period influences the adhesive forces generated at the early stages of adhesion.

Chemical, structural properties, and osteoconductive effectiveness of bone block derived from porcine cancellous bone.

The purpose of this study is to determine the efficacy of bioactive calcium phosphate obtained from porcine cancellous bone for the treatment of bone defects and nonunion. Porcine cancellous bone blocks were heat treated at 1300 degrees C for 2 h. The chemical composition, calcium-to-phosphate ratio, and microstructure of the porcine bone blocks were examined. For in vivo implantation, bone defects were created on the anteromedial aspect of the proximal tibia in seven beagle dogs and the xenograft bone blocks were placed into these defects. Plain radiographs were taken at 2-week intervals for roentgenographic evaluation. At 12 weeks, the specimens were stained with hematoxylin and eosin (H&E). The composition and morphology of heat-treated porcine cancellous bone were found to be similar to heat-treated human cancellous bone. Radiographs showed union between the host bone/bone-block interfaces. At 12 weeks, uniform and substantial new bone formation was observed. It is concluded that heat-treated porcine cancellous bone demonstrated effective osteoconductivity. This high-temperature heat-treatment technique has several advantages, including decreased risk of disease transmission and immunoreactivity, while also offering excellent biocompatibility.

In vitro study of osteogenic differentiation of bone marrow stromal cells on heat-treated porcine trabecular bone blocks.

This in vitro study investigated the potential of the heat-treated porcine trabecular bone block as a bone substitute for the treatment of bone defects or related diseases. Chemical, mechanical, and morphological studies of bone blocks were performed. The resultant properties were compared with the properties of currently available commercial products from bovine trabecular bones. The major component of the bone block was hydroxyapatite, and the ratio of Ca/P was 1.65-1.66. The average values of the compressive Young's modulus and the ultimate strength were 346.33 +/- 83.15 and 6.66 +/- 1.62 MPa, respectively. The pore size of the heat-treated bone blocks was approximately 300-500 microm. For the biological investigations, expanded bone marrow stromal cells (BMSCs) were isolated from the femurs of New Zealand White rabbits and were dynamically seeded into the heat-treated porcine bone block (10x10x5 mm3). Before the cells were seeded, the heat-treated porcine bone blocks were divided into two groups: collagen coated blocks (n=16) and uncoated blocks (n=16). Within each group, the blocks were again divided into two groups, depending on the culture method, i.e., static or rotating culture. Cells were cultured in the blocks for up to 6 weeks. Scanning electron microscopic examination after 4 weeks showed that the cell layers attached to the porcine bone block. Proliferation and osteogenic differentiation were analyzed by cell counting, an MTT assay, alkaline phosphatase activity, and total protein content. The deposition of extracellular substances and osteoid formation surrounded by osteoblast-like cuboidal cells were confirmed through histochemical staining and transmission electron microscopy. Based on the results of this study, we conclude that heat-treated porcine trabecular bone has great potential as a bone substitute and may even be superior to currently available commercial products.

A study of compatibility between cells and biopolymeric surfaces through quantitative measurements of adhesive forces.

The mechanism of cell adhesion to biomaterials or components of the extracellular matrix is an important topic in the field of tissue engineering and related biotechnological processes. Many factors affect cell adhesion, and many biochemical and biological studies have attempted to identify their roles in the adhesion mechanism. Systematic studies of this nature require quantification of the adhesive force of a cell to identify the effect of a specific factor. However, most studies of cell adhesive force have used qualitative approaches. We propose a new technique for quantifying the force by which cells adhere to various biomaterial surfaces, which utilizes the relationship between the deflection of a cantilever beam and the required force. A micropipette was used as the cantilever beam. This technique was used to measure the attachment forces of chondrocytes seeded on three different biodegradable polymers commonly used in tissue engineering and medicine: poly epsilon-carprolactone (PCL), poly(L-lactide) (PLLA) and poly(lactic-co-glycolic acid) (PGLA, L/G = 75:25). The bond between the cells and the three polymers was evaluated using the quantified adhesive forces. The adhesive forces were also measured 8, 12, 24 h and 5 days after seeding the chondrocytes on the polymer surfaces. Results of statistical analysis showed that the cells attached to the PLLA had the strongest average attachment force for up to 24 h after seeding (P < 0.05).