Examination of intestinal ultrastructure, bowel wall apoptosis and tight junctions in the early phase of sepsis.

Gut hyperpermeability can be caused by either apoptosis of the intestinal epithelium or altered status, permeability or porosity of tight junctions. This project aims to elucidate these mechanisms in the early phase of sepsis. Eighteen male wild type mice were randomized to two groups. All mice received one single gavage of fluorescein isothiocyanate (FITC) dextran 30 min before intervention. One group (n = 10) underwent cecal ligation and puncture to induce sepsis. The other group (n = 8) was sham operated. Septic animals exhibited significantly increased permeability for FITC 8 h post-operatively. Significantly increased serum interleukin-6, tumor-necrosis-factor-alpha and interleukin-1-beta confirmed sepsis. Septic animals showed significant bowel wall inflammation of ileum and colon samples. PCR revealed significantly increased expression of claudin-2 and decreased expressions of claudin-4, tight-junction-protein-1 and occludin-1 resembling increased permeability of tight junctions. However, these alterations could not be confirmed at the protein level. Light microscopy revealed significant dilatation of intercellular spaces at the basal sections of intestinal epithelial cells (IEC) in septic animals confirmed by increased intercellular spaces at the level of tight junctions and adherens junctions in electron microscopy (TEM). In small angle X-ray scattering no increase in number or size of nanopores could be shown in the bowel wall. HOECHST staining and PCR of ileum samples for apoptosis markers proofed no relevant differences in intestinal epithelial cell apoptosis between the groups. Intestinal hyperpermeability in septic animals was most likely caused by alterations of the intercellular contacts and not by apoptosis or increased size/number of nanopores of intestinal epithelial cells in this murine model of early sepsis.

Bone-implant degradation and mechanical response of bone surrounding Mg-alloy implants.

In the present paper, first results of the influence of the degradation of biodegradable materials on the hardness of the bone are presented in detail. For this purpose, different materials (Mg, Ti and biopolymers) were implanted into the femora of growing rats and bone cross sections were examined for the micro-hardness (MH). The aim of the present paper was to examine the mechanical response of the bone areas surrounding the implant at defined sites and at specified periods after implantation. A special focus was set on Mg alloys. In earlier in-vitro and in-vivo studies, an accumulation of Magnesium in the vicinity of the implant was detected by using different techniques. Therefore, micro-hardness measurements were performed, and the mechanical strength of bone was correlated with the exchange of Magnesium and Calcium in Hydroxyapatite. After the operation and implantation, the micro-hardness values became temporarily lower, but after complete degradation of the implants, the values were identical with those of specimens containing no implants.

The potential of isotopically enriched magnesium to study bone implant degradation in vivo.

This pilot study highlights the substantial potential of using isotopically enriched (non-radioactive) metals to study the fate of biodegradable metal implants. It was possible to show that magnesium (Mg) release can be observed by combining isotopic mass spectrometry and isotopic pattern deconvolution for data reduction, even at low amounts of Mg released a from slowly degrading 26Mg enriched (>99%) Mg metal. Following implantation into rats, structural in vivo changes were monitored by µCT. Results showed that the applied Mg had an average degradation rate of 16±5µmyear-1, which corresponds with the degradation rate of pure Mg. Bone and tissue extraction was performed 4, 24, and 52weeks after implantation. Bone cross sections were analyzed by laser ablation inductively coupled plasma mass spectrometry (ICP-MS) to determine the lateral 26Mg distribution. The 26Mg/24Mg ratios in digested tissue and excretion samples were analyzed by multi collector ICP-MS. Isotope pattern deconvolution in combination with ICP-MS enabled detection of Mg pin material in amounts as low as 200ppm in bone tissues and 20ppm in tissues up to two fold increased Mg levels with a contribution of pin-derived Mg of up to 75% (4weeks) and 30% (24weeks) were found adjacent to the implant. After complete degradation, no visual bone disturbance or residual pin-Mg could be detected in cortical bone. In organs, increased Δ26Mg/24Mg values up to 16‰ were determined compared to control samples. Increased Δ26Mg/24Mg values were detected in serum samples at a constant total Mg level. In contrast to urine, feces did not show a shift in the 26Mg/24Mg ratios. This investigation showed that the organism is capable of handling excess Mg well and that bones fully recover after degradation. STATEMENT OF SIGNIFICANCE: Magnesium alloys as bone implants have faced increasing attention over the past years. In vivo degradation and metabolism studies of these implant materials have shown the promising application in orthopaedic trauma surgery. With advance in Mg research it has become increasingly important to monitor the fate of the implant material in the organism. For the first time, the indispensible potential of isotopically enriched materials is documented by applying 26Mg enriched Mg implants in an animal model. Therefore, the spatial distribution of pin-Mg in bone and the pin-Mg migration and excretion in the organism could be monitored to better understand metal degradation as well as Mg turn over and excretion.

µXRF Elemental Mapping of Bioresorbable Magnesium-Based Implants in Bone.

This study investigated the distribution of the elemental constituents of Mg-based implants at various stages of the degradation process in surrounding bone tissue, with a focus on magnesium (Mg), as the main component of the alloy, and yttrium (Y), due to its potential adverse health effects. The measurements were performed on the implant-bearing thin sections of rat bone in a time series of implant degradation between one and 18 months. Micro X-ray fluorescence analysis (µXRF) with a special spectrometer meeting the requirements for the measurements of low-Z elements was used. It was found that the migration and accumulation behaviour of implant degradation products is element-specific. A sharp decrease in Mg was observed in the immediate vicinity of the interface and no specific accumulation or aggregation of Mg in the adjacent bone tissue was detected. By contrast, Y was found to migrate further into the bone over time and to remain in the tissue even after the complete degradation of the implant. Although the nature of Y accumulations must still be clarified, its potential health impact should be considered.

Comparative biomechanical and radiological characterization of osseointegration of a biodegradable magnesium alloy pin and a copolymeric control for osteosynthesis.

Magnesium alloys offer great advantages as degradable implant material for pediatric fracture fixation and hold the potential to overcome certain critical shortcomings inherent to currently used degradable (co)polymers. Besides good biocompatibility and appropriate degradation kinetics, sufficient implant anchorage in host bone is critical to prevent implant failure. Bone-implant anchorage of biodegradable magnesium alloys, however, has not yet been related and compared to that of copolymers, their degradable counterparts currently in clinical use. The aim of this study, therefore, was to comparatively assess bone-implant interface strength and the amount of peri-implant bone of a biodegradable magnesium alloy pin (Mg-Y-Nd-HRE) and a self-reinforced copolymeric control (85/15 poly(l-lactic-co-glycolic acid)). To this purpose, push-out testing, microfocus computed tomography (µCT), histological and scanning electron microscopic examination was performed after 4, 12 and 24 weeks of transcortical implantation in 72 rats. Biomechanical testing revealed significantly higher ultimate shear strength for the magnesium alloy pins than for the copolymeric controls at all 3 timepoints (P≤0.001 for all comparisons). As evaluated by µCT, the amount of bone present near the interface and in a wider radius (up to 0.5mm) around it was higher in the magnesium alloy implants at 4 weeks, without significant differences at 12 and 24 weeks. Histological examination confirmed direct bone-to-implant contact for both implant types. In vivo degradation of implants did not induce any noticeable local or systemic inflammation. This data suggests that the investigated degradable magnesium alloy rod exhibits markedly superior bone-implant interface strength and a greater amount of peri-implant bone than a self-reinforced copolymeric control currently in use; thus it fulfills a crucial prerequisite for its successful clinical deployment as an alternative degradable orthopedic implant material. Further studies, however, are warranted to evaluate the long-term degradation behavior and biocompatibility of the investigated degradable magnesium-based alloy.

Influence of environment on the fatigue crack growth behaviour of 12% Cr steel.

In the present work, the influence of different environments on the fatigue crack growth behaviour of 12% Cr steam turbine blade steel is investigated. Fatigue crack growth rates (FCGRs) in the near threshold regime are measured with ultrasonic fatigue testing technique. Fatigue tests are performed in vacuum, air and different aqueous environments with defined chloride and oxygen content. Furthermore, the influence of different stress ratios is investigated. It is found that crack propagation is not necessarily enhanced with increasing corrosiveness. In the aqueous environments, the FCGRs below 10⁻8 m/cycle are lower than in air. The threshold stress intensity factor ranges are higher or equal. Observation of the fracture surfaces shows oxide formation and partly intergranular fracture for specimens tested in aqueous environments. Crack closure effects seem to be responsible for this unexpected behaviour.

PHB, crystalline and amorphous magnesium alloys: promising candidates for bioresorbable osteosynthesis implants?

In this study various biodegradable materials were tested for their suitability for use in osteosynthesis implants, in particular as elastically stable intramedullary nails for fracture treatment in paediatric orthopaedics. The materials investigated comprise polyhydroxybutyrate (PHB), which belongs to the polyester family and is produced by microorganisms, with additions of ZrO2 and a bone graft substitute; two crystalline magnesium alloys with significantly different degradation rates ZX50 (MgZnCa, fast) and WZ21 (MgYZnCa, slow); and MgZnCa bulk metallic glasses (BMG). Push-out tests were conducted after various implantation times in rat femur meta-diaphysis to evaluate the shear forces between the implant material and the bone. The most promising materials are WZ21 and BMG, which exhibit high shear forces and push-out energies. The degradation rate of ZX50 is too fast and thus the alloy does not maintain its mechanical stability long enough during the fracture-healing period. PHB exhibits insufficient mechanical properties: it degrades very slowly and the respective low shear forces and push-out energy levels are unsatisfactory.

Fracture tolerance of reaction wood (yew and spruce wood in the TR crack propagation system).

The fracture properties of spruce and yew were studied by in-situ loading in an environmental scanning microscope (ESEM). Loading was performed with a micro-wedge splitting device in the TR-crack propagation direction. The emphasis was laid on investigating the main mechanisms responsible for a fracture tolerant behavior with a focus on the reaction wood. The fracture mechanical results were correlated with the features of the surface structure observed by the ESEM technique, which allows loading and observation in a humid environment. Some important differences between the reaction wood and normal wood were found for both investigated wood species (spruce and yew), including the formation of cracks before loading (ascribed to residual stresses) and the change of fracture mode during crack propagation in the reaction wood. The higher crack propagation resistance was attributed mainly to the different cell (i.e. fiber) geometries (shape, cell wall thickness) and fiber angle to the load axis of the reaction wood, as basic structural features are responsible for more pronounced crack deflection and branching, thus leading to crack growth retardation. Fiber bridging was recognized as another crack growth retarding mechanism, which is effective in both wood species and especially pronounced in yew wood.

Bone-implant interface strength and osseointegration: Biodegradable magnesium alloy versus standard titanium control.

Previous research on the feasibility of using biodegradable magnesium alloys for bone implant applications mainly focused on biocompatibility and corrosion resistance. However, successful clinical employment of endosseous implants is largely dependent on biological fixation and anchorage in host bone to withstand functional loading. In the present study, we therefore aimed to investigate whether bone-implant interface strength and osseointegration of a novel biodegradable magnesium alloy (Mg-Y-Nd-HRE, based on WE43) is comparable to that of a titanium control (Ti-6Al-7Nb) currently in clinical use. Biomechanical push-out testing, microfocus computed tomography and scanning electron microscopy were performed in 72 Sprague-Dawley rats 4, 12 and 24 weeks after implantation to address this question. Additionally, blood smears were obtained from each rat at sacrifice to detect potential systemic inflammatory reactions. Push-out testing revealed highly significantly greater maximum push-out force, ultimate shear strength and energy absorption to failure in magnesium alloy rods than in titanium controls after each implantation period. Microfocus computed tomography showed significantly higher bone-implant contact and bone volume per tissue volume in magnesium alloy implants as well. Direct bone-implant contact was verified by histological examination. In addition, no systemic inflammatory reactions were observed in any of the animals. We conclude that the tested biodegradable implant is superior to the titanium control with respect to both bone-implant interface strength and osseointegration. These results suggest that the investigated biodegradable magnesium alloy not only achieves enhanced bone response but also excellent interfacial strength and thus fulfils two critical requirements for bone implant applications.

Micromotion at the fracture site after tibial nailing with four unreamed small-diameter nails--a biomechanical study using a distal tibia fracture model.

BACKGROUND: Intramedullary nailing is the treatment of choice in tibia fractures allowing for closed fracture reduction and internal fixation. Small-diameter nails that preserve the endosteal blood supply act as load-sharing devices after proximal and distal locking. Despite fracture healing is influenced by movements at the fracture gap, no data are available reporting on the micromovements at the fracture site if small-diameter nails were used. METHODS: Using a Sawbone distal tibia fracture model, we assessed offset, elastic, plastic, permanent, and overall deformation at the fracture site for four small-diameter tibia nails (Expert, Synthes, Saluburg, Austria; Connex, ITS Spectromed, Lassnitzhöhe, Austria; Versanail, DePuy, Vienna, Austria; T2, Stryker, Vienna, Austria) after mechanical testing with a servohydraulic material testing machine. Cyclic loading was performed with a sinusoidal load of 700 N (+/-600) for 40,000 cycles representing 6 weeks of full weight bearing. RESULTS: Offset deformation was significantly higher for the Connex nail when compared with other nails (p < 0.001). Regarding elastic deformation, no significant difference was recorded between the implants. Plastic deformation was significantly lower if the Connex nail was used (0.134 [+/-0.053] mm; p < 0.001). Elastic deformation did not exceed 0.7 mm and plastic deformation did not exceed 0.4 mm. Regarding permanent and overall deformation, no significant difference between the implants was recorded. CONCLUSIONS: Considerable deformation at the fracture gap can be assumed even after partial weight bearing with 100 N. Despite comparable material properties, differences in axial micromotion were recorded among the nail types used in this series. The number of distal locking screws (three or four) did not substantially influence the axial movements at the fracture gap.

Factors influencing interlocking screw failure in unreamed small diameter nails--a biomechanical study using a distal tibia fracture model.

BACKGROUND: Unreamed tibia nails with small diameters are increasingly used for fracture fixation. However, little is known about the fatigue strength of proximal and distal interlocking screws in those nails. To date, no data are available reporting on mechanical differences of solid compared to cannulated tibial nails. The aim of this study was to assess the fatigue strength of proximal and distal interlocking screws of solid and cannulated small diameter tibia nails. METHODS: We created a distal tibia fracture model (AO/OTA 43 A3) using 16 Sawbones. After fracture stabilization with one of four different nail types (Expert Tibial Nail, VersaNail, T2 Tibial Nailing System, Connex), mechanical testing was performed in three loading series (40,000 cycles each) with incremental loads. Timing and type of interlocking screw failure were assessed. FINDINGS: Interlocking screw failure was observed significantly earlier (after a mean interval of 57,042 cycles) in cannulated tibial nails (VersaNail, T2) compared to solid nails (after a mean interval of 88,415 cycles; P < 0.001). Proximal interlocking screw failure was recorded if oblique screws were used proximally (VersaNail, T2, Connex). No distal interlocking screw failure was recorded in the Connex nail. Two- and three-part fractures of proximal or distal interlocking screws were observed in all specimen. INTERPRETATION: Proximal and distal interlocking screw failure has to be considered in small diameter nails in case of delayed fracture healing. To support our results, further experimental studies and clinical series are necessary.

Cell-wall recovery after irreversible deformation of wood.

The remarkable mechanical properties of biological materials reside in their complex hierarchical architecture and in specific molecular mechanistic phenomena. The fundamental importance of molecular interactions and bond recovery has been suggested by studies on deformation and fracture of bone and nacre. Like these mineral-based materials, wood also represents a complex nanocomposite with excellent mechanical performance, despite the fact that it is mainly based on polymers. In wood, however, the mechanistic contribution of processes in the cell wall is not fully understood. Here we have combined tensile tests on individual wood cells and on wood foils with simultaneous synchrotron X-ray diffraction analysis in order to separate deformation mechanisms inside the cell wall from those mediated by cell-cell interactions. We show that tensile deformation beyond the yield point does not deteriorate the stiffness of either individual cells or foils. This indicates that there is a dominant recovery mechanism that re-forms the amorphous matrix between the cellulose microfibrils within the cell wall, maintaining its mechanical properties. This stick-slip mechanism, rather like Velcro operating at the nanometre level, provides a 'plastic response' similar to that effected by moving dislocations in metals. We suggest that the molecular recovery mechanism in the cell matrix is a universal phenomenon dominating the tensile deformation of different wood tissue types.