Investigation into diseases in free-ranging ring-necked pheasants (Phasianus colchicus) in northwestern Germany during population decline with special reference to infectious pathogens.

The population of ring-necked pheasants (Phasianus colchicus) is decreasing all over Germany since the years 2008/2009. Besides impacts of habitat changes caused by current rates of land conversion, climatic influences or predators, a contribution of infectious pathogens needs also to be considered. Infectious and non-infectious diseases in free-living populations of ring-necked pheasants have been scarcely investigated so far. In the present study, carcasses of 258 deceased free-ranging pheasants of different age groups, predominantly adult pheasants, collected over a period of 4 years in the states of Lower Saxony, North Rhine-Westphalia and Schleswig-Holstein, were examined pathomorphologically, parasitologically, virologically and bacteriologically, with a focus set on infectious pathogens. A periocular and perinasal dermatitis of unknown origin was present in 62.3% of the pheasants. Additional alterations included protozoal cysts in the skeletal musculature (19.0%), hepatitis (21.7%), enteritis (18.7%), gastritis (12.6%), and pneumonia (11.7%). In single cases, neoplasms (2.6%) and mycobacteriosis (1.7%) occurred. Further findings included identification of coronaviral DNA from trachea or caecal tonsils (16.8%), siadenoviral DNA (7.6%), avian metapneumoviral RNA (6.6%), and infectious bursal disease viral RNA (3.7%). Polymerase chain reaction (PCR) on herpesvirus, avian influenza virus (AIV), paramyxovirus type 1 (PMV-1), avian encephalomyelitis virus (AEV), and chlamydia were negative. Based on the present results, there is no indication of a specific pathogen as a sole cause for population decline in adult pheasants. However, an infectious disease can still not be completely excluded as it may only affect reproduction effectivity or a certain age group of pheasants (e.g., chicks) which were not presented in the study.

Effect of calcium phosphate coating and rhBMP-2 on bone regeneration in rabbit calvaria using poly(propylene fumarate) scaffolds.

Various calcium phosphate based coatings have been evaluated for better bony integration of metallic implants and are currently being investigated to improve the surface bioactivity of polymeric scaffolds. The aim of this study was to evaluate the role of calcium phosphate coating and simultaneous delivery of recombinant human bone morphogenetic protein-2 (rhBMP-2) on the in vivo bone regeneration capacity of biodegradable, porous poly(propylene fumarate) (PPF) scaffolds. PPF scaffolds were coated with three different calcium phosphate formulations: magnesium-substituted ß-tricalcium phosphate (ß-TCMP), carbonated hydroxyapatite (synthetic bone mineral, SBM) and biphasic calcium phosphate (BCP). In vivo bone regeneration was evaluated by implantation of scaffolds in a critical-sized rabbit calvarial defect loaded with different doses of rhBMP-2. Our data demonstrated that scaffolds with each of the calcium phosphate coatings were capable of sustaining rhBMP-2 release and retained an open porous structure. After 6weeks of implantation, micro-computed tomography revealed that the rhBMP-2 dose had a significant effect on bone formation within the scaffolds and that the SBM-coated scaffolds regenerated significantly greater bone than BCP-coated scaffolds. Mechanical testing of the defects also indicated restoration of strength in the SBM and ß-TCMP with rhBMP-2 delivery. Histology results demonstrated bone growth immediately adjacent to the scaffold surface, indicating good osteointegration and osteoconductivity for coated scaffolds. The results obtained in this study suggest that the coated scaffold platform demonstrated a synergistic effect between calcium phosphate coatings and rhBMP-2 delivery and may provide a promising platform for the functional restoration of large bone defects.

Gait cycle analysis: parameters sensitive for functional evaluation of peripheral nerve recovery in rat hind limbs.

BACKGROUND: Video-assisted gait kinetics analysis has been a sensitive method to assess rat sciatic nerve function after injury and repair. However, in conduit repair of sciatic nerve defects, previously reported kinematic measurements failed to be a sensitive indicator because of the inferior recovery and inevitable joint contracture. OBJECTIVE: This study aimed to explore the role of physiotherapy in mitigating joint contracture and to seek motion analysis indices that can sensitively reflect motor function. METHODS: Data were collected from 26 rats that underwent sciatic nerve transection and conduit repair. Regular postoperative physiotherapy was applied. Parameters regarding step length, phase duration, and ankle angle were acquired and analyzed from video recording of gait kinetics preoperatively and at regular postoperative intervals. RESULTS: Stride length ratio (step length of uninjured foot/step length of injured foot), percent swing of the normal paw (percentage of the total stride duration when the uninjured paw is in the air), propulsion angle (toe-off angle subtracted by midstance angle), and clearance angle (ankle angle change from toe off to midswing) decreased postoperatively comparing with baseline values. The gradual recovery of these measurements had a strong correlation with the post-nerve repair time course. CONCLUSIONS: Ankle joint contracture persisted despite rigorous physiotherapy. Parameters acquired from a 2-dimensional motion analysis system, that is, stride length ratio, percent swing of the normal paw, propulsion angle, and clearance angle, could sensitively reflect nerve function impairment and recovery in the rat sciatic nerve conduit repair model despite the existence of joint contractures.

LINAC-radiosurgery for nonsecreting pituitary adenomas. Long-term results.

BACKGROUND AND PURPOSE: Stereotactic linear accelerator-based radiosurgery (LINAC-RS) is increasingly used for microsurgically inaccessible or recurrent pituitary adenomas. This single-center study evaluates the long-term follow-up after LINAC-RS of nonsecreting pituitary adenomas (NSA). PATIENTS AND METHODS: Between 1992 and August 2008, 65 patients with NSA were treated. Patient treatment and follow-up were conducted according to a prospective protocol. Indications for LINAC-RS were (1) tumor recurrence or (2) residual tumor. Three patients were treated primarily. For analysis of prognostic factors, patients were grouped according to epidemiological or treatment-associated characteristics. RESULTS: A total of 61 patients with a follow-up ≥ 12 months (median 83 months, range 15-186 months, longest follow-up of published radiosurgery series) were evaluated with regard to their clinical, radiological, and endocrinological course. The median tumor volume was 3.5 ml (± 4.3 ml, range 0.3-17.3 ml) treated with a median surface and maximum dose of 13.0 Gy and 29.7 Gy, respectively. Local tumor control was achieved in 98%. One patient died of unrelated cause after 36 months and 1 patient developed a radiation-induced seizure disorder. Visual complications did not occur. In 37 of 41 patients (90.2%), pituitary function remained stable. Maximum dose to the pituitary ≤ 16 Gy and female gender were positive prognostic factors for the preservation of pituitary function. CONCLUSION: LINAC-RS is a minimally invasive, safe, and effective treatment for recurrent NSA or microsurgically inaccessible residual tumor. LINAC-RS yielded a high rate of local long-term tumor control with a small number of radiation-induced side effects.

Incorporation of phosphate group modulates bone cell attachment and differentiation on oligo(polyethylene glycol) fumarate hydrogel.

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.

Controlled release of vascular endothelial growth factor using poly-lactic-co-glycolic acid microspheres: in vitro characterization and application in polycaprolactone fumarate nerve conduits.

Vascular endothelial growth factor (VEGF) is a potent angiogenic stimulator. Controlled release of such stimulators may enhance and guide the vascularization process, and when applied in a nerve conduit may play a role in nerve regeneration. We report the fabrication and in vitro characterization of poly-lactic-co-glycolic acid (PLGA) microspheres encapsulating VEGF and the in vivo application of nerve conduits supplemented with VEGF-containing microspheres. PLGA microspheres containing VEGF were prepared by the double emulsion-solvent evaporation technique. This yielded 83.16% of microspheres with a diameter <53 µm. VEGF content measured by ELISA indicated 93.79±10.64% encapsulation efficiency. Release kinetics were characterized by an initial burst release of 67.6±8.25% within the first 24h, followed by consistent release of approximately 0.34% per day for 4 weeks. Bioactivity of the released VEGF was tested by human umbilical vein endothelial cell (HUVEC) proliferation assay. VEGF released at all time points enhanced HUVEC proliferation, confirming that VEGF retained its bioactivity throughout the 4 week time period. When the microsphere delivery system was placed in a biosynthetic nerve scaffold robust nerve regeneration was observed. This study established a novel system for controlled release of growth factors and enables in vivo studies of nerve conduits conditioned with this system.

Reformulating polycaprolactone fumarate to eliminate toxic diethylene glycol: effects of polymeric branching and autoclave sterilization on material properties.

Polycaprolactone fumarate (PCLF) is a cross-linkable derivative of polycaprolactone diol that has been shown to be an effective nerve conduit material that supports regeneration across segmental nerve defects and has warranted future clinical trials. Degradation of PCLF (PCLF(DEG)) releases toxic small molecules of diethylene glycol used as the initiator for the synthesis of polycaprolactone diol. In an effort to eliminate this toxic degradation product we present a strategy for the synthesis of PCLF from either propylene glycol (PCLF(PPD)) or glycerol (PCLF(GLY)). PCLF(PPD) is linear and resembles the previously studied PCLF(DEG), while PCLF(GLY) is branched and exhibits dramatically different material properties. The synthesis and characterization of their thermal, rheological, and mechanical properties are reported. The results show that the linear PCLF(PPD) has material properties similar to the previously studied PCLF(DEG). The branched PCLF(GLY) exhibits dramatically lower crystalline properties resulting in lower rheological and mechanical moduli, and is therefore a more compliant material. In addition, the question of an appropriate Food and Drug Administration approvable sterilization method is addressed. This study shows that autoclave sterilization of PCLF materials is an acceptable sterilization method for cross-linked PCLF and has minimal effect on the PCLF thermal and mechanical properties.

Electrically conductive surface modifications of three-dimensional polypropylene fumarate scaffolds.

Polypropylene fumarate (PPF) scaffolds fabricated by rapid prototyping were surface modified by solution deposition of electrically conductive polypyrrole coatings with or without hydroxyapatite. Scaffolds were electrically conductive with resistivity as low as 2Ω. Scaffold characterization by Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy and thermo gravimetric analysis shows both polypyrrole and hydroxyapatite are present. Cell viability, attachment, proliferation, and differentiation were analyzed using human fetal osteoblast cells. These studies show that surface modification using hydroxyapatite improved cell attachment and proliferation of osteoblasts onto the PPF scaffolds. Alkaline phosphatase activity as a marker for osteogenic differentiation of cell to mature osteoblasts was analyzed. Our data reveal that osteoblasts maintained their phenotype on PPF scaffolds with and without coatings. Thus, these scaffolds could be appropriate candidates for our future in vivo studies.

Material properties and electrical stimulation regimens of polycaprolactone fumarate-polypyrrole scaffolds as potential conductive nerve conduits.

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.

Cross-linking characteristics and mechanical properties of an injectable biomaterial composed of polypropylene fumarate and polycaprolactone co-polymer.

In this work, a series of co-polymers of polypropylene fumarate-co-polycaprolactone (PPF-co-PCL) were synthesized via a three-step polycondensation reaction of oligomeric polypropylene fumarate (PPF) with polycaprolactone (PCL). The effects of PPF precursor molecular weight, PCL precursor molecular weight and PCL fraction in the co-polymer (PCL feed ratio) on the maximum cross-linking temperature, gelation time and mechanical properties of the cross-linked co-polymers were investigated. The maximum cross-linking temperature fell between 38.2 ± 0.3 and 47.2 ± 0.4°C, which increased with increasing PCL precursor molecular weight. The gelation time was between 4.2 ± 0.2 and 8.5 ± 0.7 min, and decreased with increasing PCL precursor molecular weight. The compressive moduli ranged from 44 ± 1.8 to 142 ± 7.4 MPa, with enhanced moduli at higher PPF precursor molecular weight and lower PCL feed ratio. The compressive toughness was in the range of 4.1 ± 0.3 and 17.1 ± 1.3 kJ/m(3). Our data suggest that the cross-linking and mechanical properties of PPF-co-PCL can be modulated by varying the composition. Therefore, the PPF-co-PCL co-polymers may offer increased versatility as an injectable, in situ polymerizable biomaterial than the individual polymers of PPF and PCL.

Development of electrically conductive oligo(polyethylene glycol) fumarate-polypyrrole hydrogels for nerve regeneration.

Electrically conductive hydrogel composites consisting of oligo(polyethylene glycol) fumarate (OPF) and polypyrrole (PPy) were developed for applications in nerve regeneration. OPF-PPy scaffolds were synthesized using three different anions: naphthalene-2-sulfonic acid sodium salt (NSA), dodecylbenzenesulfonic acid sodium salt (DBSA), and dioctyl sulfosuccinate sodium salt (DOSS). Scaffolds were characterized by ATR-FTIR, XPS, AFM, dynamic mechanical analysis, electrical resistivity measurements, and swelling experiments. OPF-PPy scaffolds were shown to consist of up to 25 mol % polypyrrole with a compressive modulus ranging from 265 to 323 kPa and a sheet resistance ranging from 6 to 30 × 10(3) Ohms/square. In vitro studies using PC12 cells showed OPF-PPy materials had no cytotoxicity and PC12 cells showed distinctly better cell attachment and an increase in the percent of neurite bearing cells on OPF-PPy materials compared to OPF. The neurite lengths of PC12 cells were significantly higher on OPF-PPyNSA and OPF-PPyDBSA. These results show that electrically conductive OPF-PPy hydrogels are promising candidates for future applications in nerve regeneration.

The development of electrically conductive polycaprolactone fumarate-polypyrrole composite materials for nerve regeneration.

Electrically conductive polymer composites composed of polycaprolactone fumarate and polypyrrole (PCLF-PPy) have been developed for nerve regeneration applications. Here we report the synthesis and characterization of PCLF-PPy and in vitro studies showing PCLF-PPy materials support both PC12 cell and dorsal root ganglia (DRG) neurite extension. PCLF-PPy composite materials were synthesized by polymerizing pyrrole in preformed PCLF scaffolds (M(n) 7,000 or 18,000 g mol(-1)) resulting in interpenetrating networks of PCLF-PPy. Chemical compositions and thermal properties were characterized by ATR-FTIR, XPS, DSC, and TGA. PCLF-PPy materials were synthesized with five different anions (naphthalene-2-sulfonic acid sodium salt (NSA), dodecylbenzenesulfonic acid sodium salt (DBSA), dioctyl sulfosuccinate sodium salt (DOSS), potassium iodide (I), and lysine) to investigate effects on electrical conductivity and to optimize chemical composition for cellular compatibility. PCLF-PPy materials have variable electrical conductivity up to 6 mS cm(-1) with bulk compositions ranging from 5 to 13.5 percent polypyrrole. AFM and SEM characterization show microstructures with a root mean squared (RMS) roughness of 1195 nm and nanostructures with RMS roughness of 8 nm. In vitro studies using PC12 cells and DRG show PCLF-PPy materials synthesized with NSA or DBSA support cell attachment, proliferation, neurite extension, and are promising materials for future studies involving electrical stimulation.

[Balloon assisted kyphoplasty: new technique for treatment of vertebral compression fractures]. / La kyphoplastie par ballonnets: nouvelle technique dans le traitement percutané des tassements vertébraux.

Balloon assisted kyphoplasty and vertebroplasty are minimally invasive procedures with established value in the management of osteoporotic and neoplastic vertebral compression fractures. Kyphoplasty, a variation of vertebroplasty, is a technique with indications and therapeutic particularities that require validation.

Radiosurgery for cerebral arteriovenous malformations in hereditary hemorrhagic telangiectasia.

The authors evaluated the efficacy of radiosurgery (RS) for cerebral arteriovenous malformations in hereditary hemorrhagic telangiectasia (HHT AVMs). Two patients with seven HHT AVMs were treated by linear accelerator-RS. Complete obliteration was achieved 18 to 24 months post-treatment without side effects. Because HHT AVMs are small and multiple, RS is superior to microsurgery because it is noninvasive and all AVMs can be treated in one session regardless of their location.

[Contribution of quantitative radio-scintigraphy to diagnosis of wrist injuries undetected on plain films: a prospective study of 154 cases]. / Apport de la radioscintigraphie quantitative dans les traumatismes du poignet avec radiographies initiales normales: étude prospective de 154 cas.

PURPOSE OF THE STUDY: Fractures of the scaphoid must be diagnosed quickly to avoid persistent nonunion and the risk of osteoarthritis. Despite meticulous physical examination and adequate x-ray detection, numerous occult fractures still go unrecognized. The aim of this prospective study was to analyze the pertinence of quantitative radio-scintigraphy (QRS) presently used for the diagnosis of occult wrist fractures. MATERIAL AND METHODS: Quantitative radio-scintigraphy (QRS) is a new imaging technique associating quantitative bone scan and numerical fusion between bone scan images and x-ray images. We conducted a prospective study between November 1994 and March 1999 to evaluate the pertinence of this examination technique for the diagnosis of occult wrist fractures in patients presenting clinical symptoms suggestive of wrist fracture but whose plain x-rays were initially considered normal. Further some patients had several series of plain x-rays performed at several week intervals in order to search for fractures becoming progressively visible on plain x-rays. After the QRS data was acquired, these patients' x-rays were reviewed again. We also compared the cost of QRS, repeated x-rays, bone scan and MRI at the Besançon University Hospital. RESULTS: QRS was performed in all 154 patients and revealed 61 fractures (56 single-line and 5 multiple-line fractures). Thus 43.5% of these patients had occult wrist fractures (41% of which involved the carpal scaphoid). DISCUSSION: Occult fracture of the wrist, particularly the carpal scaphoid, is frequent. Repeated x-ray examination does not increase the rate of detection of these fractures. Bone scans may also fail to reveal occult fractures. MRI is a key examination in the assessment of wrist fracture symptoms, but is presently not available in all institutions. Bone scan is classically insufficiently precise. QRS is a rapidly available low-cost examination which we have found to be indispensable for the diagnosis of occult wrist fractures. With early QRS diagnosis, the risk of neglected carpal scaphoid fracture and subsequent nonunion and osteoarthritis together with the personal, social, and medicolegal consequences can be avoided.

4-hydroxynonenal induces apoptosis, NF-kappaB-activation and formation of 8-isoprostane in vascular smooth muscle cells.

Oxidation of lipids is considered a key feature of atherogenesis. Lipid peroxidation products such as oxidized LDL or the bioactive aldehyde 4-hydroxynonenal (HNE) exert mitogenic effects on vascular smooth muscle cells (VSMC). These effects appear to be concentration-dependent since in addition to our previous reports on growth promotion at lower concentrations we here indicate induction of apoptosis in VSMC by 4-hydroxynonenal (HNE) at higher concentrations (100 micromol/L). In a line with HNE's previously documented effects on key mitogenic signaling elements, we also report on activation by this aldehyde of the redox-sensitive transcription factor NF-kappaB, a key regulator of apoptosis: HNE (1.0 micromol/L) induced DNA-binding of NF-kappaB in VSMC. The effect was inhibited by antioxidants, N-acetylcysteine and pyrrolidine dithio-carbamate. HNE caused phosphorylation but not degradation of the inhibitory subunit IkappaB-alpha. HNE itself acts as an oxidant as was investigated with measurements of 8-isoprostane which ranks among the most valuable available biomarkers of lipid peroxidation: HNE (1.0 micromol/L) increased 8-isoprostane levels in VSMC by 4.5-fold (p < 0.05). Compared to the controls, plasma samples from apoEnull mice exhibited elevated levels of 8-isoprostane (40 pg/mL, 3.2-fold increase) and the combined aldehydes HNE and malonaldehyde (1.5 micromol/L, 2.5-fold increase), (p < 0.05, resp). In addition, immunohistochemistry indicated the presence of HNE-protein adducts in atheroscerlotic lesions of apoEnull mice. Thus HNE is present in atherosclerotic tissue at concentrations that are bioactive in vitro. The data further indicate the involvement of the lipid peroxidation product HNE in atherogenesis.

Aging, oxidative responses, and proliferative capacity in cultured mouse aortic smooth muscle cells.

The cellular mechanisms that contribute to the acceleration of atherosclerosis in aging populations are poorly understood, although it is hypothesized that changes in the proliferative capacity of vascular smooth muscle cells is contributory. We addressed the relationship among aging, generation of reactive oxygen species (ROS), and proliferation in primary culture smooth muscle cells (SMC) derived from the aortas of young (4 mo old) and aged (16 mo old) mice to understand the phenotypic modulation of these cells as aging occurs. SMC from aged mice had decreased proliferative capacity in response to alpha-thrombin stimulation, yet generated higher levels of ROS and had constitutively increased mitogen-activated protein kinase activity, in comparison with cells from younger mice. These effects may be explained by dysregulation of cell cycle-associated proteins such as cyclin D1 and p27Kip1 in SMC from aged mice. Increased ROS generation was associated with decreased endogenous antioxidant activity, increased lipid peroxidation, and mitochondrial DNA damage. Accrual of oxidant-induced damage and decreased proliferative capacity in SMC may explain, in part, the age-associated transition to plaque instability in humans with atherosclerosis.

Thrombin regulates vascular smooth muscle cell growth and heat shock proteins via the JAK-STAT pathway.

The growth-stimulating effects of thrombin are mediated primarily via activation of a G protein-coupled receptor, PAR-1. Because PAR-1 has no intrinsic tyrosine kinase activity, yet requires tyrosine phosphorylation events to induce mitogenesis, we investigated the role of the Janus tyrosine kinases (JAKs) in thrombin-mediated signaling. JAK2 was activated rapidly in rat vascular smooth muscle cells (VSMC) treated with thrombin, and signal transducers and activators of transcription (STAT1 and STAT3) were phosphorylated and translocated to the nucleus in a JAK2-dependent manner. AG-490, a JAK2-specific inhibitor, and a dominant negative JAK2 mutant inhibited thrombin-induced ERK2 activity and VSMC proliferation suggesting that JAK2 is upstream of the Ras/Raf/MEK/ERK pathway. To elucidate the functional significance of JAK-STAT activation, we studied the effect of thrombin on heat shock protein (Hsp) expression, based upon the following: 1) reports that thrombin stimulates reactive oxygen species production in VSMC; 2) the putative role of Hsps in modulating cellular responses to reactive oxygen species; and 3) the presence of functional STAT1/3-binding sites in Hsp70 and Hsp90beta promoters. Indeed, thrombin up-regulated Hsp70 and Hsp90 protein expression via enhanced binding of STATs to cognate binding sites in the Hsp70 and Hsp90 promoters. Together, these results suggest that JAK-STAT pathway activation is necessary for thrombin-induced VSMC growth and Hsp gene expression.